Hospital for Children and Adolescents Jorvi Hospital Skin and Allergy Hospital University of Helsinki, Finland

Children Born Very Prematurely at School Age:

Lung Function and Outcome after

Antenatal and Neonatal Dexamethasone Treatment

SUVI MIESKONEN

Academic dissertation

To be publicly discussed with the permission of the Medical Faculty of the University of Helsinki, in the Niilo Hallman Auditorium of the Hospital for Children and Adolescents, on January 21st, 2005, at 12 noon.

Helsinki 2005

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Supervised by: Professor Mikko Hallman Department of Pediatrics University of Oulu Oulu, Finland Docent Markku Turpeinen Skin and Allergy Hospital University of Helsinki Helsinki, Finland Reviewed by: Docent Kirsti Heinonen Department of Pediatrics University of Kuopio Kuopio, Finland Kirsi Timonen, MD, PhD Department of Clinical Physiology and Nuclear Medicine University of Kuopio Kuopio, Finland Opponent: Docent Outi Tammela Department of Pediatrics University of Tampere Tampere, Finland Cover photo: Suvi Mieskonen Anniina held by her mother (warm thanks) ISBN 952-91-8184-1 (paperback) ISBN 952-10-2261-2 (pdf) Yliopistopaino Helsinki 2005

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To Elsa, Alpo and Sara

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ABSTRACT

Glucocorticoids enhance the maturation of premature lung. Antenatal glucocorticoids reduce

the risk of respiratory distress syndrome and neonatal death in preterm infants, and postnatal

dexamethasone (DEX) facilitates weaning from the ventilator. However, these drugs may

disturb the growth and development of lung and brain. To evaluate the long-term effects of

antenatal or postnatal DEX treatment, we studied 49 children aged 7.3-9.2 years, born very

prematurely at 24-30 gestational weeks and with 600-1575 g of birth weight in 1989-1991,

who had participated in randomized placebo-controlled trials of neonatal DEX therapy

administered to ventilated infants at high risk for bronchopulmonary dysplasia (BPD) (n=16)

or/and antenatal DEX therapy in conjunction with exogenous surfactant (n=35).

Cardiorespiratory morbidity, neurological outcome and the family history of smoking or

atopy were evaluated using hospital records and questionnaires. Physical examination, skin

prick tests, impulse oscillometry, flow-volume spirometry, pulmonary diffusing capacity test

and whole-body plethysmography were carried out, and the oscillometric method was

compared with the conventional lung function methods. Lung inflammation was studied by

exhaled nitric oxide (eNO) measurement and cardiac outcome by chest X-ray,

electrocardiogram and echocardiography. The control group consisted of 18 non-atopic

children born at term and tested for lung function.

Antenatal or neonatal DEX had no negative effect on cardiopulmonary outcome. On the

contrary, the children born after antenatal placebo showed greater bronchodilator

responsiveness (∆FEV1) than those born after antenatal DEX.

Severe or moderate disability was common in our prematurely born children (29%).

Neurosensory outcome did not differ between the neonatal DEX and placebo groups.

However, antenatal DEX treatment associated with a higher rate of normal school attendance

and less severe or moderate disability.

The children born prematurely, particularly boys, were shorter for age than the controls.

The BPD children weighed less adjusted for height than the non-BPD children. Antenatal or

postnatal DEX treatment had no significant effect on weight or height.

Half of the children born prematurely reported frequent respiratory symptoms during the

preceding year, with no association with BPD or either DEX treatment. Poorer exercise

tolerance and more common use of inhaled medication were reported in the BPD group.

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Atopy was rare in the children with BPD. Atopy associated with significantly increased

eNO concentrations. In non-atopic children, eNO levels correlated neither with the values of

lung function measurements nor with BPD. Thus, eNO was not an inflammatory marker of

BPD at school age, but it may be valuable in studying atopic asthma even in children born

prematurely.

Airway obstruction, bronchial hyperreactivity and decreased gas exchange were evident in

the children born very prematurely, especially those with BPD. Smoking in the family

associated with decreased gas exchange, and these children should therefore be strongly

advised against smoking or passive smoking. Impulse oscillometry yielded lung function

information concordant with the conventional methods and served well in studying even

children with neurological handicap or small lung volumes.

To conclude, abnormal lung function, somatic growth and neurosensory development

were common at school age in children born very prematurely. However, we found no

negative effect associated with antenatal or neonatal DEX treatment. Less strenuous lung

function methods, such as impulse oscillometry, help to assess the pulmonary disease of

disabled or young children who cannot cooperate in conventional lung function studies.

Despite the few abnormal findings, cardiac follow-up is important, as the chronic lung disease

of prematurity may later in life lead to cardiac sequelae or mask the cardiac symptoms. If

possible, very prematurely born children should also be protected from additional risk factors

for impaired lung function, such as infections or smoking.

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LIST OF ORIGINAL PUBLICATIONS

This thesis is based on the following publications, which are referred to in the text by their

Roman numerals (I to IV).

I Malmberg LP, Mieskonen S, Pelkonen A, Kari A, Sovijärvi AR, Turpeinen M.

Lung function measured by the oscillometric method in prematurely born children

with chronic lung disease. Eur Respir J 2000 Oct; 16(4): 598-603.

II Mieskonen ST, Malmberg LP, Kari MA, Pelkonen AS, Turpeinen MT,

Hallman NMK, Sovijärvi AR. Exhaled nitric oxide at school age in prematurely

born infants with neonatal chronic lung disease. Pediatr Pulmonol 2002 May;

33(5): 347-355.

III Mieskonen S, Eronen M, Malmberg LP, Turpeinen M, Kari MA, Hallman M.

Controlled trial of dexamethasone in neonatal chronic lung disease: an 8-year

follow-up of cardiopulmonary function and growth. Acta Paediatr 2003 Aug;

92(8): 896-904.

IV Mieskonen S, Kari MA, Eronen M, Malmberg LP, Turpeinen M, Hallman M.

Randomized trial of antenatal dexamethasone before preterm birth:

cardiopulmonary function, growth, and neurosensory outcome at 8 years.

Submitted for publication.

In addition, some unpublished data are presented.

All refers to data of all study children born very prematurely.

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ABBREVIATIONS AT acceleration time ACTH adrenocorticotrophic hormone ASD atrial septal defect BPD bronchopulmonary dysplasia C compliance CLD chronic lung disease COPD chronic obstructive

pulmonary disease CP cerebral palsy CPAP continuous positive airway

pressure DEX dexamethasone ∆FEV1 percentage change in FEV1

from baseline DLCO pulmonary diffusing capacity ∆Rrs5 percentage change in Rrs5

from baseline ∆Xrs5 percentage change in Xrs5

from baseline ECG electrocardiogram ELBW extremely low birth weight,

<1000 g eNO exhaled nitric oxide FEF50 forced expiratory flow at 50%

of FVC FEF25-75 forced expiratory flow during

the middle half of FVC FEV1 forced expiratory volume in

one second FiO2 fraction of inspiratory oxygen FOT forced oscillation technique FRC functional residual capacity fres resonance frequency FVC forced vital capacity

IMV intermittent mandatory

ventilation IOS impulse oscillometry IVH intraventricular hemorrhage KCO diffusing coefficient for

carbon monoxide LBW low birth weight, <2500 g MEF maximal expiratory flow NEC necrotizing enterocolitis NO nitric oxide NOS nitric oxide synthase PAP pulmonary arterial pressure PDA patent ductus arteriosus PEF peak expiratory flow PVL periventricular leukomalacia Raw airway resistance RBBB right bundle branch block RDS respiratory distress syndrome ROP retinopathy of prematurity Rrs5 respiratory resistance at 5 Hz RV residual volume RVET right ventricular ejection time SGA small for gestational age, birth

weight <-2 SD from the mean birth weight for each gestational week

SGaw specific conductance in the airways

TGV thoracic gas volume TLC total lung capacity VC vital capacity VLBW very low birth weight,

<1500 g Xrs5 respiratory reactance at 5 Hz

Gaw airway conductance

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CONTENTS ABSTRACT 1 LIST OF ORIGINAL PUBLICATIONS 3 ABBREVIATIONS 4 CONTENTS 5 INTRODUCTION 8 REVIEW OF LITERATURE 11 1. Human lung growth and development............................................................................................. 11 Factors affecting lung growth and development 2. Respiratory distress syndrome (RDS)............................................................................................. 12 Clinical presentation and pathology

Pulmonary surfactant Surfactant therapy Incidence and prevention of RDS

3. Bronchopulmonary dysplasia (BPD)............................................................................................... 14 �Old� BPD

�Changing� BPD �New� BPD

3.1. Pathogenesis of BPD.............................................................................................................. 16 Oxygen toxicity

Barotrauma - volutrama Inflammatory injury Vascular injury Surfactant dysfunction Genetic predisposition

3.2. Incidence of BPD.................................................................................................................... 20 3.3. Prevention and treatment of BPD........................................................................................... 21 Optimization of oxygen administration Antioxidant therapy Mode of ventilation Infection/inflammation Pulmonary vascular resistance Nutrition and growth Improving respiratory mechanics, prevention of oedema Glucocorticoids 4. Respiratory morbidity in children born prematurely....................................................................... 24 5. Tests of lung function in infancy and childhood............................................................................. 26

Measures of forced expiratory flow Measures of lung volume Measures of compliance and resistance Measures of gas exchange

6. Lung function in children born prematurely.................................................................................... 28 Infancy School age

7. Other indices associated with lung disease...................................................................................... 32 Immune response Exhaled nitric oxide (eNO)

8. Follow-up of growth, neurodevelopment and cardiac function in children born prematurely........ 34 Growth Neurosensory development Cardiac function

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9. Pharmacologic effects of glucocorticoids........................................................................................ 37 Effects on the pulmonary defence system Effects on lung structure and growth

Effects on other tissues Brain development

10. Trials on glucocorticoids during the antenatal - neonatal period.................................................... 40 10.1. Glucocorticoid treatment of preterm infants......................................................................... 40 Early postnatal glucocorticoid treatment

Moderately early postnatal glucocorticoid treatment Delayed postnatal glucocorticoid treatment Neurodevelopmental outcome after postnatal dexamethasone treatment Growth after postnatal dexamethasone treatment Respiratory morbidity and lung function after postnatal dexamethasone treatment Cardiovascular findings after postnatal dexamethasone treatment Recommendations and practice of postnatal dexamethasone treatment

10.2. Antenatal glucocorticoid treatment...................................................................................... 47 Prevention of neonatal diseases Neurodevelopmental outcome after antenatal glucocorticoids Growth after antenatal glucocorticoids Respiratory morbidity and lung function after antenatal glucocorticoids Cardiovascular findings after antenatal glucocorticoids

AIMS OF THE STUDY 52 PATIENTS AND METHODS 53 1. Patients............................................................................................................................................. 53 2. Neonatal and follow-up data collection........................................................................................... 54 3. Study protocol.................................................................................................................................. 55 4. Lung function methods.................................................................................................................... 56

Flow-volume spirometry Impulse oscillometry Pulmonary diffusing capacity test Whole-body plethysmography

5. Exhaled nitric oxide......................................................................................................................... 58 6. Cardiological measurements............................................................................................................ 58 7. Statistics........................................................................................................................................... 59 RESULTS 61 1. Population characteristics................................................................................................................ 61

Children born very prematurely Children with BPD Atopic children Children in the postnatal dexamethasone study Children in the antenatal dexamethasone study

2. Growth............................................................................................................................................. 63 Children born very prematurely Children with BPD Children after postnatal dexamethasone treatment Children after antenatal dexamethasone treatment

3. Respiratory symptoms and medication............................................................................................ 64 Children born very prematurely Children with BPD Children after postnatal dexamethasone treatment

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Children after antenatal dexamethasone treatment 4. Smoking in the family..................................................................................................................... 66

Influence on growth Influence on respiratory symptoms Influence on lung function

5. Atopy............................................................................................................................................... 67 Children born very prematurely Children with BPD Children after postnatal dexamethasone treatment Children after antenatal dexamethasone treatment

6. Lung function.................................................................................................................................. 69 Oscillometric method Children born very prematurely Children with BPD Children after postnatal dexamethasone treatment Children after antenatal dexamethasone treatment Effect of smoking

7. Exhaled nitric oxide........................................................................................................................ 75 Children born very prematurely Influence of atopy, familial smoking or glucocorticoid inhalation Non-atopic children with BPD

8. Cardiac function.............................................................................................................................. 76 Children after postnatal dexamethasone treatment Children after antenatal dexamethasone treatment

9. Neurological follow-up................................................................................................................... 78 Children after postnatal dexamethasone treatment Children after antenatal dexamethasone treatment

DISCUSSION 81 1. Methodological aspects................................................................................................................... 81 Definition of bronchopulmonary dysplasia 2. Oscillometric method...................................................................................................................... 83 3. Exhaled nitric oxide ........................................................................................................................ 84 4. Growth............................................................................................................................................. 86 5. Atopy............................................................................................................................................... 87 6. Respiratory symptoms and medication........................................................................................... 88 7. Smoking in the family..................................................................................................................... 89 8. Lung function.................................................................................................................................. 89

Prematurely born children with or without BPD Glucocorticoids versus lung function Lung function after postnatal dexamethasone treatment Lung function after antenatal glucocorticoids

9. Cardiovascular findings................................................................................................................... 93 Cardiovascular findings after postnatal dexamethasone treatment Cardiovascular findings after antenatal glucocorticoids

10. Neurosensory development............................................................................................................. 95 Neurosensory development after glucocorticoid treatment Neurosensory development after postnatal dexamethasone treatment Neurosensory development after antenatal glucocorticoids

CONCLUSIONS 100 ACKNOWLEDGEMENTS 102 REFERENCES 104 ORIGINAL PUBLICATIONS 119

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INTRODUCTION

Prematurity is a major cause of mortality and chronic morbidity in childhood. The outcome of

preterm birth (<37 gestational weeks) depends on time and place. In developed countries, the

survival of very low birth weight (VLBW, <1500 g) infants has increased, and their initial

outcome has improved along with the advances in perinatal care (Lemons et al 2001,

Saugstad 2001, Koivisto et al 2004). However, their infant mortality rate, i.e. death until one

year of age per 1000 live births, is more than 100-fold compared to infants with normal birth

weight. Neonatal deaths during the first four weeks of life account for two thirds of overall

infant mortality, and half of the deaths take place during the first week of life. (Mathews et al

2003) Neonatal mortality of VLBW infants is highest during the first few days of life, mainly

due to respiratory distress syndrome (RDS), infections and immaturity (Stevenson et al 1998).

In a recent Finnish study of extremely low birth weight (ELBW, <1000 g) infants, neonatal

mortality was 38%. Perinatal mortality (death from the 20th week of gestation to seven days of

age) was 55%, accounting for 39% of all perinatal deaths. (Tommiska et al 2001) In a large

multicenter study of VLBW infants in 1989-1990, 78% survived until discharge (Hack et al

1995); in 1995-1996, 84% survived (Lemons et al 2001). In Finland, 834 VLBW infants were

born alive in 1987 and 611 in 2003, accounting for 1.4% and 1.0% of live births, respectively

(Stakes 2004).

The clinical course and definitions of bronchopulmonary dysplasia (BPD), also called the

chronic lung disease (CLD) of prematurity, have changed over time, but with no clear decline

in incidence (Parker et al 1992). Besides BPD, defined as a need for supplemental oxygen at

36 postconceptional weeks in the present study (Shennan et al 1988), periventricular brain

damage, including intraventricular-periventricular haemorrhage (IVH) and periventricular

leukomalacia (PVL), necrotizing enterocolitis (NEC) and retinopathy of prematurity (ROP)

cause severe morbidity in VLBW infants. Pulmonary problems persist well beyond the

neonatal period or infancy. They may be associated with disturbances in growth, cardiac

function and neurodevelopment that may continue until school age or adulthood.

The pathogenesis of BPD is multifactorial and incompletely understood. Immaturity, oxygen

toxicity, baro/volutrauma and other perinatal factors, such as infection, promote lung injury.

Inflammation plays an important role in the progress of initial changes into a chronic lung

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disease. In the less mature survivors, BPD is characterized by an arrest in lung development,

including minimal alveolarization and impaired vascular growth. (Jobe 1999)

Dexamethasone (DEX) and betamethasone are synthetic glucocorticoids with multiple acute

beneficial effects on immature lung (Ballard & Ballard 1995). In 1972, Liggins and Howie

showed that maternal treatment with betamethasone before preterm birth reduced the rate of

RDS and neonatal death. This result was confirmed more extensively in multiple trials during

the following years (Crowley 1995), and antenatal glucocorticoid therapy is now

recommended for women at risk of delivery at 24-34 gestational weeks (National Institutes of

Health, NIH 1995).

An early trial with hydrocortisone as treatment of RDS revealed no significant benefits

(Baden et al 1972). In the 1980´s, DEX treatment for severe BPD was found to decrease the

time on mechanical ventilation and supplemental oxygen (Mammel et al 1983, Avery et al

1985). With growing evidence of the inflammatory basis of BPD, DEX was considered as an

early treatment of very high-risk infants (Yeh et al 1990). However, treatment after birth was

associated with severe side effects, and animal experiments provided evidence of abnormal

brain growth and development after corticosteroids (Whitelaw & Thoresen 2000). The

widespread use of glucocorticoids postnatally and even antenatal use, particularly as multiple

courses, evoked concern about long-term adverse effects. After the initiation of our study,

early treatment with postnatal DEX was shown to associate with an increased risk of cerebral

palsy (CP) and a delay in neurodevelopment (Yeh et al 1998, O´Shea et al 1999, Shinwell et

al 2000). New guidelines limited the use of DEX to exceptional circumstances, i.e. infants on

maximal ventilatory and oxygen support (American Academy of Pediatrics, AAP 2002), but

the final role of postnatal corticosteroid is still undefined (Halliday 2003). Although antenatal

DEX treatment was associated with serious adverse consequences, long-term data after

antenatal or neonatal glucocorticoid trials are sparse or completely missing.

The lung function of prematurely born children cannot usually be assessed until school age,

since the conventional tests require considerable cooperation. Even then, the examinations

may be hampered by neurological disabilities and small lung volumes. The forced oscillation

technique (FOT) and impulse oscillometry (IOS), a modification of FOT, enable the

measurement of lung function during normal quiet tidal breathing. They therefore require

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only minimal cooperation and are suitable at preschool age. Scant FOT data are available on

prematurely born children (Duiverman et al 1988).

Inhaled glucocorticoid treatment of low birth weight (LBW) children had no effect on the

baseline lung function or the airway response to histamine at school age in one study, arguing

against an inflammatory basis of airway hyperresponsiveness (Chan & Silverman 1993).

However, changes in T lymphocyte subsets, with an association with bronchial lability,

suggested that the abnormal immunological alteration may be related to BPD (Pelkonen et al

1999). Measurement of the exhaled nitric oxide (eNO) concentration has been proposed as a

non-invasive method to assess airway inflammation in asthma (Alving et al 1993, Lundberg

et al 1996) and disease activity in chronic obstructive pulmonary disease (COPD) (Maziak et

al 1998). ENO has been associated with eosinophilic inflammation in the lungs (Piacentini et

al 1999, Mattes et al 1999), but the role in non-atopic asthma remains undefined. No reports

exist of eNO in school-aged children with BPD.

In the present follow-up study of two randomized placebo-controlled trials, we evaluated the

effect of antenatal or neonatal DEX on cardiopulmonary function, growth and neurosensory

development at school age in children born very prematurely. Elaborate non-invasive methods

were used to measure lung function, exhaled NO and cardiac function. The children born

very prematurely were born very preterm (<31 gestational weeks) as well as with a very low

birth weight (<1575 g) in 1989-1991.

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REVIEW OF LITERATURE

1. Human lung growth and development

Normal lung development during organogenesis can be divided into five stages (Burri 1997,

Kotecha 2000). During the embryonic stage (3-7 weeks of gestation), the lung develops as an

outgrowth of the ventral wall of the primitive foregut endoderm. Epithelial cells from the

endoderm invade the surrounding mesoderm to form the proximal structures of the respiratory

tract: the trachea, main bronchi, five lobes and 18 major lobules. The pulmonary arteries

develop to accompany the branching airways. During the pseudo-glandular stage (7-16

weeks), the epithelial tubules are surrounded by thick mesenchymal tissue, and the conducting

airways, terminal bronchioles and primitive acinus are formed. The pseudo-stratified

columnar epithelium is progressively replaced by tall columnar cells in the proximal airways

and cuboidal cells in the distal acinar structures. During the canalicular stage (16-26 weeks),

primary acini develop, consisting of respiratory bronchioles, alveolar ducts and rudimentary

alveoli. The intra-acinar capillaries derive from the surrounding mesenchyme. Many cells in

the distal epithelium differentiate into lamellar body-containing cells resembling type II

pneumocytes. During the saccular stage (24-36 weeks), the peripheral airways enlarge as the

acinar tubules dilate and the walls become thinner, resulting in increased gas-exchanging

surface area. Lamellar bodies increase, and terminal differentiation of type II cells into type I

pneumocytes takes place. Capillaries closely associated with type I cells reduce the distance

between the future air-blood interface. During the alveolar stage (32 weeks - 10 years), the

secondary alveolar septa are formed. Mesenchymal cells proliferate to deposit the

extracellular matrix, epithelial cells (type I and type II pneumocytes) line the alveolar walls,

and endothelial cells grow into the secondary septa and, after remodelling, form a single

capillary loop. This maturation results in a marked increase in gas-exchanging surface area, as

the estimated 20 to 50 million alveoli present at birth increase to 300 million by adulthood.

Factors affecting lung growth and development

Data on lung growth are mainly derived from studies of animals with differences in timing of

lung maturation. Different mechanisms and a number of hormones, growth factors, cytokines

and transcription factors are involved in normal lung growth, differentiation and repair

processes after lung injury. (Kotecha 2000)

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During the canalicular stage, pulmonary hypoplasia may be secondary to oligohydramnion

as a result of prolonged rupture of membranes, to the restriction of fetal breathing movements

or to space-occupying abdominal organs in diaphragmatic hernia. Severe pulmonary

hypoplasia leads to severe respiratory distress because of poorly developed peripheral airways

and small lung size. On the other hand, overdistention of the lung by fluid seems to improve

lung growth by releasing many growth factors. Severe malnutrition decreases lung volumes

without interfering with maturation, whereas postnatal food restriction may exacerbate severe

lung injury and resistance against toxic agents, such as hyperoxia. (Kotecha 2000)

Glucocorticoids are involved in both lung growth and differentiation. A genetic lack of

glucocorticoid receptor in fetal mice results in hypoplastic immature lungs and impaired gas

exchange at term birth (Cole et al 1995). Glucocorticoids bind with high affinity to

cytoplasmic receptors, and the activated steroid-receptor complex binds to several response

elements in the promoters of a number of genes profoundly influencing transcription.

Glucocorticoid activity partly controls the synthesis and secretion of alveolar surfactant.

Furthermore, the increase in compliance and maximal lung volume, the decrease in vascular

permeability, the maturation of the parenchymal structure and the clearance of lung water are

controlled by glucocorticoids, adrenergic agents and other agonists. (Ballard & Ballard 1995,

Mallampalli et al 1997, Jobe & Bancalari 2001)

2. Respiratory distress syndrome (RDS)

Clinical presentation and pathology

RDS in the premature infant presents with an acute, severe respiratory distress and

requirement for supplemental oxygen shortly after birth. The chest radiograph shows

increased density in the lung fields with fine granularity, air bronchograms and elevation of

the diaphragm, indicating generalized atelectasis and lung oedema. Traditionally, the severity

increases for three days, and normal lung function is restored within a week. Nearly all infants

die with supportive care only, and about 50% die with the administration of supplemental

oxygen alone. In infants <32 gestational weeks, who rarely survive without ventilation using

continuous distending pressures, the condition is usually prolonged and commonly involves

pulmonary complications, such as pneumothorax, pulmonary interstitial emphysema and

infection. Immaturity of the lung and the cardiovascular system (leaky pulmonary capillaries

shortly after birth, immature pulmonary connective tissue, compliant chest cage, patency of

ductus arteriosus, immature host defence against microbes and other external noxious agents)

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further predispose these infants to lung injury. RDS is a major risk factor of BPD and IVH

and involves significant mortality. (Bancalari & Bidegain 1999)

Pulmonary surfactant

In 1959, Avery and Mead discovered evidence of deficient surface activity in lung lavage of

infants dying of hyaline membrane disease. These findings marked the beginning of

systematic research leading to the introduction of surfactant therapy for RDS and further

therapeutic applications. Lung surfactant is needed to decrease surface tension and to prevent

the collapse of the small airways. The composition of surfactant, as reviewed by Jobe (1993a)

and Hallman et al (2001), consists of specific phospholipids (85-95%), neutral lipids (3-8%)

and specific proteins (5-10%). Of the four identified surfactant-associated proteins, SP-A

improves surface properties and has roles in pulmonary host defence. The lipophilic proteins

SP-B and SP-C facilitate rapid adsorption and spreading of surfactant phospholipids to the

gas-liquid interface. The timetable of the maturation of the human lung and surfactant system

is highly variable. Generally, it takes place before term, increasing towards term.

RDS is caused by a deficiency in alveolar surfactant due to immaturity of alveolar type II

epithelial cells. Many premature infants have sufficient quantities of surfactant in the airways

and do not develop RDS, even when born after only 65-80% of the pregnancy has lapsed.

(Hallman et al 2001) The antenatal development of surfactant secretion corresponds closely to

the risk of RDS, and the suspectibility to RDS is influenced by variations in the SP-A and SP-

B genes, race, sex and perinatal complications. Besides providing alveolar stability, individual

components of surfactant have important roles in the innate immune response and in the

defence against microbes. (Marttila et al 2003b, Marttila 2003a, Hallman et al 2002)

Surfactant therapy

The first pilot study of exogenous surfactant given to severely ill preterm infants in 1980

(Fujiwara et al) was soon followed by randomized clinical trials on exogenous surfactant

from human amniotic fluid, from mammalian lungs or from broncho-alveolar lavage fluid and

on synthetic surfactant (Merritt & Hallman 1988). Surfactant therapy ameliorates or prevents

the expiratory failure dramatically when administered into the airways in RDS or at very

premature birth. Neonatal mortality decreased by 45-55% and serious pulmonary air leak by

50-80%. Since the early 1990´s, administration of animal-based or synthetic surfactant into

the airways has been an approved treatment of RDS, and it has since been accepted into

common use as effective and life-saving treatment. (Hallman et al 2001) As more and more

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very immature infants survive, there has been no significant decrease in the incidence of BPD

or patent ductus arteriosus (PDA) or IVH (Jobe 1993a).

Incidence and prevention of RDS

Despite surfactant and antenatal glucocorticoid treatments (Chapter 10.2.), RDS remains a

major cause of mortality and morbidity, as the survival of more immature infants increases. In

Finland, the incidence of RDS increased during the 1960-1970´s, but remained stable from

1990-1995 to 1996-1999 (8.7-7.6 per 1000 livebirths). The gestational-age-specific incidence

of and mortality from RDS decreased, but still accounted for 15% of neonatal deaths.

(Koivisto et al 2004) In the USA, in an inborn cohort of 4438 infants in 14 centres (1995-

1996), RDS occurred in 50% of VLBW infants and in 78% of those with 501-750 g birth

weight (Lemons et al 2001). In a Finnish national study (1996-1997), the neonatal mortality

of ELBW infants was 38%, and in 39% of the cases the reported leading cause of death was

RDS. In that study, of the infants surviving >12 hours, 76% had RDS, and 74% of those <30

weeks of gestational age received surfactant. (Tommiska et al 2001)

Expectant management of high-risk pregnancies, analysis of fetal lung maturity,

avoidance of near-term caesarean sections without labour and glucocorticoid treatment in

imminent preterm birth (<34 gestational weeks) are practices that decrease the risk of RDS

(Hallman 1992a, Crowley 2000).

3. Bronchopulmonary dysplasia (BPD)

�Old� BPD

BPD was first described by Northway et al in 1967 as a severe lung injury in mechanically

ventilated large preterm infants, who required 80-100% oxygen and had a prolonged course of

severe respiratory distress. The clinical, radiological and pathological progression of the

primary lung disease had several stages. The initial RDS (stage I, days 1-3) was characterized

by severe diffuse lung disease. It was followed by increasing opacification of the lungs,

corresponding to pathological findings of exudation in the airways and patchy bronchiolar and

alveolar epithelial necrosis (stage II, days 4-10). Stage III (days 10-20) was a transition to

chronic lung disease with small, round areas of radiolucency alternating with areas of sponge-

like irregular density in chest radiographs, representing emphysematous alveoli adjacent to

atelectatic alveoli, and pulmonary fibrosis. At stage IV (after 30 days), chronic lung disease

with hyperexpansion and larger cysts became evident in the basal lung fields. This stage was

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frequently accompanied by right ventricular hypertrophy and cardiomegaly with cor

pulmonale. Pathologically, emphysema with thickened lung interstitium, dysplastic airways

and lung vasculature were frequently evident. (Northway et al 1967, Northway 1990a)

�Changing� BPD

In the 1970´s, BPD was defined as a respiratory failure in the neonatal period requiring

assisted ventilation for at least three days, with persistent respiratory symptoms and

radiological findings and oxygen dependency at four weeks of age (Bancalari et al 1979,

Tooley 1979, Edwards 1979a). In the 1980´s, with improvements in the ventilation techniques,

the survival of infants <1000 g increased, and oxygen concentrations and peak inspiratory

pressures decreased, modifying the radiological changes into a fine reticular pattern, with or

without hyperexpansion (Northway 1990a). O´Brodovich & Mellins (1985) reviewed BPD

and suggested it to represent a non-specific reaction of the lung to slowly resolving acute

injury. Hyperinflation was seen as the predominant radiographic abnormality along with

chronic respiratory difficulties, prolonged and recurrent hospitalization, neurodevelopmental

disabilities and growth restriction associated with BPD. In VLBW infants, apart from

classical coarse reticulation, a milder type with homogenous or patchy opacification,

particularly perihilar, was present in radiographs (Hyde et al 1989), and the pathological

progression of lung injury differed from that originally seen: a largely parenchymal picture

with marked increases in interstitial elastic tissue, less severe squamous metaplasia and poor

development and simplification of the terminal respiratory unit (Chambers & van Velzen

1989).

With the survival of increasingly immature infants, the criterion of supplemental oxygen

at 28 days became insufficient in anticipating long-term morbidity. In 1988, Shennan et al

proposed that the need for supplemental oxygen at 36 gestational weeks was a better predictor

of abnormal outcome during the first two years of life. This criterion was adopted into wide

use, and it is often also referred to as chronic lung disease (CLD).

Palta et al (1998) compared different criteria, such as supplemental oxygen and/or

radiograph or a severity score based on oxygen need, ventilatory variables, blood gases and

radiograph at 30 days of age, and supplemental oxygen at 36 gestational weeks. All criteria,

especially the radiographic ones, were predictive of long-term outcome at five years of age,

but the combination of radiograph and supplemental oxygen at 36 weeks was proposed.

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prominent findings, accompanied by early vascular lesions, right ventricular hypertrophy and

cardiomegaly with cor pulmonale. Infants dying of BPD in the era of surfactant treatment had

fewer and larger alveoli with less fibrosis and inflammation than in the past (Husain et al

1998, Jobe & Bancalari 2001), and alveolar hypoplasia was accompanied by bronchial

smooth muscle hypertrophy and bronchial gland hyperplasia, contributing to airflow

limitation (Margraf et al 1991). Preterm baboons (75% of gestation) exposed to 100% oxygen

and mechanical ventilation, used to model the �old� BPD, had a lower alveolar number at the

age of 33 weeks compared to those ventilated with clinically appropriate oxygen (Coalson et

al 1995). Similar pathology was seen in baboons delivered extremely preterm (67% of

gestation) after prenatal glucocorticoid, surfactant treatment and mechanical ventilation with

even less than 50% oxygen (Coalson et al 1999). The �new� BPD is characterized by an

arrest of lung development with minimal alveolarization, less airway epithelial disease and

less interstitial fibrosis (Jobe 1999), with dysmorphic capillary configurations (Coalson 2003).

The predominant pathogenetic factor in BPD is a recurrent inflammatory reaction,

whether provoked by an intrauterine infection or inflammation that persists after birth or

caused by neonatal damage to the airways and airspaces due to hypoxia,

barotrauma/volutrauma or infection (Groneck & Speer 1995, Watterberg et al 1996, Speer

1999, Hallman et al 2001). Other antenatal or postnatal factors, such as intrauterine growth

retardation, preeclampsia, drugs or malnutrition may also affect the risk of BPD (Kotecha

2000).

Oxygen toxicity

Preterm infants are predisposed to changing levels of supplemental oxygen and mechanical

ventilation soon after birth, as reviewed by Barrington & Finer (1998), Kotecha & Silverman

(1999) and Jobe & Bancalari (2001). According to the traditional view, BPD is caused

primarily by oxidant- and ventilation-mediated injury. In animal studies, oxygen can arrest the

septation of lungs at the saccular stage (Coalson et al 1995), with more impairment at higher

oxygen levels (Coalson et al 1999).

High concentrations of oxygen induce lung inflammation and may lead to chronic fibrotic

and destructive changes, as the production of reactive oxygen species and the release of

chemotactic factors lead to the release of inflammatory mediators and proteolytic enzymes.

The low levels of antiproteases and antioxidant enzymes in the serum and lung of preterm

infants result in increased susceptibility to oxygen toxicity and to BPD. (Barrington & Finer

1998)

16

Barotrauma - volutrauma

Multiple factors act additively or synergistically to promote lung injury. The initiation of

mechanical ventilation causes a proinflammatory response. In preterm baboons, mechanical

ventilation even without high levels of oxygen may result in the pathologic lesion of BPD

(Coalson et al 1999). Although the exact mechanisms of lung injury are unknown, attempts to

reduce barotraumas are suggested to reduce the incidence of BPD (Kotecha & Silverman

1999). The avoidance of intubation and mechanical ventilation by administering continuous

positive airway pressure (CPAP) in the delivery room has been associated with a lower

incidence of BPD (Avery et al 1987, Van Marter et al 2000).

Inflammatory injury

Pulmonary inflammation is maximal at 7-10 days in infants who develop BPD (Vyas &

Kotecha 1997), as evidenced by the increase in neutrophils and multiple proinflammatory and

chemotactic factors present in the airspaces of ventilated preterm infants (Merritt et al 1983,

Groneck & Speer 1995, Vyas & Kotecha 1997). Infants exposed to chorioamnionitis had IL-

1β more frequently present in their tracheal fluid from the first day, which associated with the

development of BPD, despite the lower risk of RDS (Watterberg et al 1996). Colonization of

the airways of preterm infants with Ureaplasma urealyticum associated with acute respiratory

insufficiency (Ollikainen et al 1993); others found protection against RDS, but an increase in

BPD (Hannaford et al 1999). Chemotaxis of inflammatory cells and release of cytokines and

other mediators may cause lung injury and initiate the repair process. The same factors may

be released in nosocomial pneumonia. (Barrington & Finer 1998)

In very preterm born baboons, significant elevations of certain proinflammatory cytokines

(TNF-α, IL-6, IL-8, but not IL-1β and IL-10) in tracheal aspirate fluids were present at

various times during ventilatory treatment, supporting a role for mediator-induced

autoinflammation; IL-8 was elevated mostly in the presence of significant lung infection

(Coalson et al 1995). Preterm lung contains very few macrophages or granulocytes.

Granulocytes appear in the lung soon after the initiation of ventilation in animals (Carlton et

al 1997), which correlates with pulmonary oedema and the appearance of early indicators of

injury and occurs parallel to a decrease in circulating granulocytes. This decrease in

granulocytes at one hour of age associates with an increased risk of BPD (Ferreira et al 2000).

Proteases produced by activated white blood cells in the lung may contribute to the

progression of lung injury.

17

Vascular injury

Pulmonary vascular resistance is often increased in infants with BPD, and decreased vascular

development may be important in the pathopsysiology of BPD (Jobe & Bancalari 2001). The

vascular injury may result from direct toxicity of stimuli, such as oxygen and mechanical

ventilation, to the developing vasculature or from the release of toxic inflammatory mediators.

Damage to the developing alveolar epithelium is suggested to interrupt the signalling between

vascular endothelial growth factor (VEGF) and its receptors and to result in abnormal

pulmonary capillary growth and function. Converse theories suggest that the primary

disruption of pulmonary vascular development impairs alveolarization and may result in

histologic changes similar to BPD. (Parker & Abman 2003) Low VEGF levels in tracheal

aspirates have been found in preterm infants who later develop BPD (Lassus et al 1999),

while high levels have been seen in infants treated with DEX (D´Angio et al 1999).

Surfactant dysfunction

The recovery of surfactant phospholipids during the first week of life among infants who

develop BPD is not strikingly different from those who have an uncomplicated course of

RDS. The most clearly detectable difference, low surfactant phosphatidylinositol in infants

developing BPD, was eliminated by inositol supplementation at birth. (Hallman et al 1992b)

Infants developing BPD also had very low SP-A levels in airway specimens, but according to

recent evidence, surfactant dysfunction in infants developing BPD is not prevented by

surfactant therapy (Hallman et al 2001).

Genetic predisposition

Genetics may contribute to BPD at multiple levels (Marttila 2003a). According to an early

study, HLA-A2 is associated with susceptibility to BPD (Clark et al 1982). The

alleles/genotypes of SP-A, SP-C or SP-D associate with the risk of inflammatory lung and

airway diseases. Rare mutations in SP-B and SP-C cause serious, often fatal, lung diseases

(Hallman et al 2002, Marttila 2003a, Hallman 2004). Thus far, one allelic variant of the

surfactant protein B gene (intron 4 deletion variant) has been shown to associate with an

increased risk of BPD (Rova et al 2004).

18

3.2. Incidence of BPD

The original BPD occurred in 11-21% of mechanically ventilated infants and in 1-8% of all

infants with RDS (Northway 1979), showing a relationship with low birth weight: 38% of

survivors of RDS with birth weights of 750-1500 g had BPD compared with less than 10% of

those >1500 g (Tooley 1979). Even after controlling for birth weight, race and sex, the

incidence at 28-30 days varied among intensive care units, correlating with differences in

treatment (Avery et al 1987, Horbar et al 1988, Kraybill et al 1987, Kraybill et al 1989). In

ELBW infants after mechanical ventilation, the most powerful predictors of BPD were male

sex and lower PaCO2 at 48 hours, indicating that it might be possible to reduce the risk by

adjusting ventilation (Kraybill et al 1989). The incidence of BPD at 28 days in VLBW infants

increased from 11% in the late 1970´s to 33% in the late 1980´s, mostly but not exclusively

due to improved survival. The increase persisted even with the criterion of oxygen at 36

gestational weeks for infants <32 gestational weeks at birth. Gestational age was a significant

predictor of BPD after adjustment for birth weight. (Parker et al 1992) In a Finnish study of

VLBW infants born in one hospital in 1990-1994, 31% of the surviving infants had BPD, i.e.

supplemental oxygen with radiographic changes, at 28 days and 13% at 36 gestational weeks.

Besides low birth weight and gestational age, early weight gain, preeclampsia and possibly

intrauterine growth retardation were important risk factors for BPD at 36 weeks. (Korhonen et

al 1999b)

BPD is currently rare in infants with >1200 g or >30 gestational weeks at birth, as gentler

ventilation techniques, antenatal glucocorticoids and surfactant treatment have minimized

severe lung injury. However, BPD is the most common complication in ELBW infants (Jobe

1999, Jobe & Bancalari 2001), and it may also occur in infants with minimal or no lung

disease initially (Charafeddine et al 1999), some of them exposed to chorioamnionitis

(Watterberg et al 1996). In the NICHD multicentre study (1995-1996), the incidence of BPD

at 36 weeks was 23% in surviving infants with birth weights of 501-1500 g, with a slight

increase over five years (19-23%, survival 80-84%); the incidence increased markedly with

decreasing gestational age, and nearly all survivors with birth weight of 400-500 g had BPD

(Lemons et al 2001). In comparison, of the VLBW infants born in 1989-1990, 78% survived

and 8% of the survivors had BPD (Hack et al 1995). Thus, no evidence suggests a decline in

the incidence of BPD.

19

3.3. Prevention and treatment of BPD

The prevention of preterm births and various maternal conditions harmful for the fetus is a

continuing challenge, with major effects on the development of BPD. O´Brodovich & Mellins

(1985) presented that the means to treat BPD are identical to those initiating the injury.

Despite the advances in perinatal care, the means to prevent and treat BPD are still

unsatisfactory, as reviewed by Barrington & Finer (1998) and more recently by Korhonen

(2004a).

Optimization of oxygen administration

As far as possible, high oxygen concentrations should be avoided in the treatment of preterm

infants. However, the saturation target range is still controversial. (Kotecha & Allen 2002, Tin

2004) Exogenous surfactant should be administered to intubated VLBW infants with

significant lung disease prior to two hours of age, to reduce the need for oxygen and

ventilatory support, even though no clear effect is seen in the incidence of BPD (Jobe &

Bancalari 2001). In BPD infants, avoidance of hypoxia is important, to alleviate the

progressive muscularisation of small pulmonary arteries and enhanced vasoreactivity (Parker

& Abman 2003). According to the present proposal, O2 saturations of 94-96% should be

aimed at for infants with established BPD who are not at risk of further progression of ROP

(Kotecha & Allen 2002).

Antioxidant therapy

Hyperoxic lung injury may be ameliorated in animals by the administration of exogenous

antioxidants, such as recombinant superoxide dismutase. In humans, no difference in the

primary outcome of death and/or BPD was found, but IVH and PVL were less severe.

(Barrington & Finer 1998) Nor did N-acetylcysteine prevent BPD (Ahola et al 2003). Specific

nutrients, such as vitamins and cofactors, may also function in antioxidant defence (Kotecha

& Silverman 1999). Despite initial suggestions, vitamin E supplementation failed to show

benefits in reducing BPD (Jobe & Bancalari 2001).

Mode of ventilation

Different modes of ventilation have been used to minimize baro- and volutrauma, but none of

them have proven superior to the others (Henderson-Smart et al 2004). In 1963, the first

infant survived intubation and ventilation without air leak or cerebral abnormalities

20

(McGettigan et al 1998). Since 1971, CPAP has been used to treat infants with RDS (Mariani

& Carlo 1998), and it is considered to lower the risk of BPD by reducing barotrauma (Avery

et al 1987). Since that time, ventilation with continuous distending pressures has been the

basis of neonatal ventilatory support. During intermittent mandatory ventilation (IMV),

asynchronous breathing may result in irregular systemic and cerebral flow patterns, air leak

and neurological complications. In 1986, patient-triggered ventilation was first described:

synchronous IMV (SIMV), assist control and pressure support ventilation were further

developed. However, conventional mechanical ventilation still induced severe lung injury.

High-frequency oscillatory ventilation, using very low tidal volumes, is used to rescue

patients and to avoid lung injury in severe respiratory distress. (McGettigan et al 1998)

Assisted ventilation is recommended to be used with the lowest possible pressures required

for adequate gas exchange, in the most gentle ways available, followed by weaning and

extubation as soon as possible, allowing moderate hypercarbia. Nasal CPAP at birth or after a

brief period of ventilation is currently studied as a non-invasive method of ventilation. (Avery

et al 1987, Kraybill et al 1989, Barrington & Finer 1998)

Infection / inflammation

Infection may contribute to the development of inflammation with an adverse effect on lung

growth and pulmonary defence functions (Watterberg et al 1996). The prevention and

treatment of perinatal and nosocomial infections have been studied in relation to BPD.

Prevention of later infections by immune globulins, including those against respiratory

syncytial virus, and vaccinations are considered to reduce further risks for the lung in BPD.

Anti-inflammatory agents have been used in treatment (sodium cromoglycate,

glucocorticoids). (Barrington & Finer 1998)

Pulmonary vascular resistance

In infants with severe RDS, vasodilators, such as inhaled nitric oxide, can be used along with

oxygen therapy to decrease pulmonary vascular resistance. Inhalation of NO may improve

oxygenation and decrease the risk of BPD by minimising intra- and extrapulmonary shunting

and by decreasing inflammation. (Jobe & Bancalari 2001)

Nutrition and growth

Nutrition plays a supportive role in normal lung development and maturation. General

undernutrition, especially insufficient protein intake, may increase the vulnerability to

21

oxidant-induced lung injury. (Jobe & Bancalari 2001) Inositol (Hallman et al 1992b), vitamin

A (Tyson et al 1999) and retinoic acid have been studied in experimental models of lung

injury (Massaro & Massaro 2000). Supplementation of inositol (Hallman et al 1992b) or

vitamin A (Tyson et al 1999) to small preterm infants may reduce the risk of BPD, although

the evidence is not convincing. Nutrition should be optimized early after birth but probably

with some delay in intravenous lipid administration until pulmonary vascular resistance has

decreased (Barrington & Finer 1998).

Growth failure is common in infants with severe BPD (Northway 1979, Markestad &

Fitzhardinge 1981,Yu et al 1983), possibly because of their increased energy expenditure and

high metabolic rate. Hypoxemia, gastroesophageal reflux, poor feeding with inadequate

nutritional intake, heart failure, neurodevelopmental handicap and socioeconomic factors

further predispose BPD infants to unsatisfactory growth (Kotecha & Silverman 1999).

However, data on the ability of optimal growth and nutrition to improve long-term outcome

are sparse.

Improving respiratory mechanics, prevention of oedema

Bronchodilating agents improve lung mechanics in ventilator-dependent infants, but their

long-term effects are unknown. Methylxanthines (theophylline) are used to prevent apnoea in

preterm infants, but they also function as bronchodilators and diuretics. Increased fluid intake

may aggravate pulmonary oedema directly or by increasing the incidence of PDA (Kotecha &

Silverman 1999). Relative fluid restriction in very preterm infants at risk for BPD decreased

both mortality and BPD rates (Tammela & Koivisto 1992). Interstitial lung water can be

reduced by loop diuretics (furosemide), which decrease pulmonary resistance and increase

dynamic compliance. Non-loop diuretics (hydrochlorothiazide and spironolactone) improve

lung mechanics, but with no consistent long-term benefit. In BPD, furosemide and inhaled

bronchodilator agents are considered to allow weaning and extubation of ventilator-dependent

infants. (Barrington & Finer 1998)

Glucocorticoids

Glucocorticoids serve as anti-inflammatory and antioedematic agents. They accelerate the

differentiation of immature lung by increasing the synthesis and secretion of lung surfactant

and the activity of antioxidants. However, they also interfere with alveolarization and

angiogenesis (See Chapter 9 and 10). Antenatal glucocorticoids should be used in imminent

preterm delivery to attenuate initial respiratory distress (Crowley 1995, NIH 1995). Inhaled

22

glucocorticoids have been used to shorten the duration of mechanical ventilation (Cole et al

1999, Shah et al 2003). Postnatal glucocorticoid treatment may be considered in severely ill,

ventilator-dependent infants to facilitate weaning (AAP 2002). However, no clear decline has

been seen in the incidence of BPD after either modality of glucocorticoid treatment.

4. Respiratory morbidity in children born prematurely

The survivors of �old� BPD born in the 1960-1970´s frequently had persistent respiratory

symptoms at school age (Smyth et al 1981) and more wheezing, upper respiratory tract

infections, pneumonia, limitations of exercise capacity and long-term medication than full-

term controls at school age (Bader et al 1987) or preterm born adolescents with no BPD

(Northway et al 1990b).

In the late 1970´s, mechanically ventilated VLBW infants had tracheobronchial

hypersecretion and frequent lower respiratory infections during the first year of life. The need

for hospitalization was associated with male gender, CPAP, mechanical ventilation and

oxygen therapy. (Wong et al 1982a) Myers et al (1986) used regular physical examination,

respiratory illness score and viral cultures until the age of one year to show that LBW infants

(<2000 g) with or without RDS did not have an increased risk for respiratory illness compared

with full-term controls. However, the RDS survivors, particularly those with BPD,

experienced their lower respiratory tract infections earlier and with more severe symptoms

than the controls.

A prospective cohort study compared LBW children (<2000 g) with unselected

schoolchildren using questionnaires. Wheezing (16%) and troublesome cough (17%) were

common in healthy schoolchildren. There were no differences in maternal smoking (40%) or

the family history of asthma (12%) between the LBW and control children. However,

whooping cough, previous chest infection and previous admission into hospital for chest

illness were twice as common in the LBW cohort than in the controls. Oxygen

supplementation, mechanical ventilation and BPD increased the rate of chest infections during

the first two years (30% in the BPD group) and also increased admission into hospital for

respiratory infection (40% in the BPD group). In the subgroup of VLBW children after

neonatal respiratory treatment, frequent and troublesome cough was more common at seven

years than in unselected controls. No excess of wheezing, but a weak correlation between

wheezing and the oxygen score, or absence from school due to respiratory symptoms was

23

found in LBW children. Wheezing associated with maternal smoking. Of the VLBW

children, those with BPD had a high prevalence of symptoms, but they did not differ

significantly from the non-BPD group. (Chan et al 1989a)

Another study confirmed the previous findings of excess respiratory morbidity associated

with mechanical ventilation in children born very prematurely, particularly during the first

two years, but found dyspnea to be uncommon at school age. BPD children needed

adenoidectomy and tympanostomy tubes more commonly. (Hakulinen et al 1990) At

preschool age, BPD children had an increased risk for respiratory infections compared to

controls born at term, with no significant difference compared to non-BPD children

(Korhonen et al 1999a). In a prospective study, the persistence of respiratory symptoms into

the second year of life was associated with mechanical ventilation, supplemental oxygen, air

leak and increased resistance in body plethysmography at six months (Yuksel & Greenough

1992).

In ELBW survivors, clinical respiratory health at the age of 14 years, based on wheezing

requiring bronchodilators or readmissions into hospital for pneumonia or asthma, was found

to be similar to normal birth weight controls, despite differences in lung function (Doyle

2000a).

The studies of VLBW children born in the 1980´s have mainly used the criterion of oxygen at

36 weeks for BPD. In a Finnish study, half of the VLBW children had had dyspneic

symptoms, and two out of 29 had suffered from prolonged coughing (>3 weeks) during the

previous year. Atopy was diagnosed in 17% of the BPD and 29% of the non-BPD children; in

their families, atopy (41% vs 50%) and smoking (50%) were common. (Pelkonen et al 1997)

Others found atopy less often in VLBW children (15%) at ten years than in full-term controls

(31%) (Siltanen et al 2001), and the lifetime prevalence of wheezing was 43% vs 17%,

respectively (Siltanen et al 2004). After severe BPD requiring oxygen at home, the children

had a higher incidence of activity limitation because of their respiratory symptoms than non-

BPD controls at the age of 11 years (Jacob et al 1998). Children born very preterm (<32

gestational weeks), especially those with BPD, required more hospitalizations during the first

two years than their full-term controls. Wheezing and chronic cough were common at school

age, with no differences between the BPD and non-BPD groups, but the BPD children used

more bronchodilators. (Gross et al 1998)

24

5. Tests of lung function in infancy and childhood

Lung function can be assessed in infancy and childhood by measuring forced expiratory flow,

lung volume, compliance and resistance, respiratory pattern, chest wall motion and gas

mixing, as reviewed by the American Thoracic Society (ATS)/ European Respiratory Society

(ERS) (1993), Sovijärvi et al (1994), Kotecha & Silverman (1999) and Sly et al (1999).

Measures of forced expiratory flow

Maximal expiratory flow (MEF) on the descending portion of the MEF-volume curve is

independent of effort and reflects airway caliber peripheral to the segment subjected to flow

limitation. The measurement of MEF is a standardized and sensitive test of abnormalities in

the intrathoracic airways. Forced expiratory flow-volume relationship can be measured by the

forced deflation technique in intubated infants and by the rapid thoracoabdominal impression

method in non-intubated infants, and it can also be used with bronchodilating or

bronchoconstricting agents. Near school age, the measurements become more accurate.

Ventilatory capacity is influenced by lung volume, airway caliber, compliance and elasticity

of the lungs and chest and respiratory muscle function. In flow-volume spirometry, forced

expiratory volume in one second (FEV1) (or in 0.5 or 0.75 s) is the best indicator of

ventilation capacity and represents mainly the larger airways, as does peak expiratory flow

(PEF). Forced vital capacity (FVC) measures the dynamic volume of the lungs. Forced

expiratory flow at 50% (FEF50) and during the middle half of FVC (FEF25-75) represent the

airflow in medium-sized and small airways and the compliance of lung tissue as well.

Spirometry is used to differentiate between obstructive and restrictive dysfunction and,

combined with bronchodilators, bronchoconstrictors or exercise, to assess hyperreactivity.

Measures of lung volume

Functional residual capacity (FRC), i.e. the volume of air in the lungs and airways at end-

expiration, is a balance between the outward recoil of the chest wall and the inward recoil of

the lungs. FRC is the only lung volume that can be measured accurately and repeatedly in

infants, by the whole-body plethysmography or gas dilution techniques (helium dilution and

nitrogen washout). In whole-body plethysmography, a child, placed on a rigid container,

makes respiratory efforts against an occlusion at the airway opening. The changes in pressure

at the airway are related to the changes in volume, and thoracic gas volume (TGV) can be

calculated by applying Boyle´s law. Plethysmography measures all gas present in the thorax,

25

including that not in direct communication with the airway (gas trapping). In healthy infants,

TGV measurements tend to be reproducible. The helium dilution technique is based on the

principle of gas equilibration between an unknown lung volume and a known volume

containing helium as an indicator gas. Gas mixing is produced by ventilatory movements, and

assuming a mass balance, the unknown lung volume can be derived from the change in

helium concentration. The nitrogen washout technique measures the volume of nitrogen

washed out of the lungs when the infant rebreathes nitrogen-free gas. Gas dilution techniques

may underestimate FRC with gas trapping due to airway obstruction. At preschool age, lung

volumes, such as total lung capacity (TLC), residual volume (RV) and RV/TLC ratio, can be

measured by whole-body plethysmography with the patient sitting in a cabine. The static lung

volumes [vital capacity (VC), TLC, RV/TLC] can be estimated by the single-breath method

of the pulmonary diffusing capacity, even if the presence of air trapping can underestimate

TLC.

Measures of compliance and resistance

The intrinsic elastic properties of the lungs and the chest can be assessed by measuring

compliance and resistance during maturation. Dynamic techniques, such as body

plethysmography, forced oscillation and the interrupter technique, are applied during

spontaneous breathing or mechanically assisted ventilation. Passive techniques, such as

multiple occlusion, passive flow-volume and weighted spirometry, are done after silencing of

the respiratory muscles. All require measurements of airflow, volume and pressure;

transpulmonary pressure is approximated by measuring the difference between the airway and

esophageal pressures. Compliance (C) is calculated as the quotient of the change in volume

divided by the pressure applied. As it is proportional to changes in lung volume, specific

compliance (corrected for FRC) can be used instead. Respiratory system resistance, i.e. the

change in pressure divided by flow, is dominated by the upper airways, and the measured

values hence do not reflect modest changes in lower airway pressure. Therefore, only

relatively large changes in airway resistance (Raw) due to bronchodilators can be detected. At

preschool age, plethysmographic measurements of Raw and specific conductance (SGaw) can

be performed with the patient sitting in a cabin; Gaw is the reciprocal of Raw, and SGaw is

related to volume [SGaw = 1/(Raw xTGV)].

Forced oscillation technique (FOT) was introduced by DuBois et al in 1956 as a method to

characterize respiratory impedance and its two components, resistance (Rrs) and reactance

(Xrs), over a wide range of frequencies. Flow oscillations generated by a loudspeaker are

26

applied to the mouth and superimposed on normal breathing: FOT uses pseudorandom noise,

and its user-friendly commercial modification, impulse oscillometry (IOS), uses rectangular

pulse signals containing harmocis up to 35 Hz or higher (Vogel & Smidt 1994, Hellinckx et al

2001). The output pressure and flow signals are analyzed for their amplitude and phase

difference, to determine the Rrs and Xrs of the total respiratory system and even to study

airway mechanics during quiet tidal breathing. The oscillometric method has been used to

describe the mechanical properties of the lung in children with asthma, cystic fibrosis and

chronic respiratory symptoms (Solymar et al 1984, Lebecque et al 1987, Lebecque &

Stănescu 1997, Timonen et al 1997, Hellinckx et al 1998) as early as the age of 2-4 years

(König et al 1984, Duiverman et al 1985, Bisgaard & Klug 1995, Klug & Bisgaard 1996).

Scant data are available on preterm children (Duiverman et al 1988). Recently, the values of

IOS and FOT were shown to be similar, but not identical (Hellinckx et al 2001).

Measures of gas exchange

The function of the lung parenchyma can be assessed by measuring the pulmonary diffusing

capacity, i.e. the diffusion of the inspired gas from the lungs into the circulation. At preschool

or school age, a single-breath method, using an inhalation of a known mixture of an inert gas

(helium) and a diffusing gas, can be used for the analysis of the total pulmonary diffusing

capacity (DLCO) and the diffusing coefficient for carbon monoxide (KCO). (Salorinne 1994)

6. Lung function in children born prematurely

Infancy

During the neonatal intensive care, due to difficulties in performance and interpretation,

measurements of lung function have been mostly done as part of trials of medication or

ventilation techniques. More studies have been done on infants, but the interpretation of data

is influenced by sedation, considerable short-term variability and normalization procedures,

such as weight. Forced expiratory techniques have improved the accuracy of measurement

and made it possible to define the degree of dysfunction, the responses to specific therapy and

the onset of and recovery from respiratory problems. Both static and dynamic compliances are

low during neonatal intensive care and largely influenced by the ventilation technique and the

distending pressures applied. Compliance measurements have been used to investigate the

effect of surfactant in neonates and the effect of glucocorticoids in young children with

interstitial lung disease. Studies with bronchodilators have shown bronchoconstriction to

27

occur even in preterm infants during the first weeks of life. (ATS/ERS 1993, Kotecha &

Silverman 1999) In infants, chest wall compliance is 3-fold compared to lung compliance,

and it may be even higher in preterm infants (5-7-fold greater), making the respiratory pump

less efficient in moving the tidal volume (Gerhardt & Bancalari 1980, ATS/ERS 1993). Raw

values are high at birth but decrease rapidly during the first year of life. After birth,

conductance increases much more slowly than FRC, resulting in a rapid drop of SGaw

(Gerhardt et al 1987a). The age-specific changes in Raw can be only evaluated in relation to

changes in lung volume, as by using SGaw.

The survivors of �old� BPD in the 1960-1970´s had increased Raw, increased airway

reactivity, increased FRC, high arterial carbon dioxide tension, low arterial oxygen tension,

right or/and left ventricular hypertrophy and pulmonary and systemic hypertension in infancy

(Northway et al 1990b).

In VLBW children born before the surfactant era in the late 1970´s, TGV was higher, and

dynamic compliance and pulmonary and airway conductance were lower at the age of six

months and normalized by the end of the first year in those ventilated for RDS compared with

non-RDS infants, regardless of whether or not normalized for body weight or TGV. The

disturbances of lung function were particularly severe in infants with lower respiratory tract

infection. (Wong et al 1982a)

In VLBW infants born in the 1980´s, FRC was in the normal range, but compliance

decreased and Raw increased after both intermittent mandatory ventilation and high-frequency

oscillatory ventilation at two months of age (Gerhardt et al 1989). In larger preterm infants

with RDS, pulmonary mechanics were almost similar during early infancy after surfactant or

placebo treatment but, at the age of one year, total pulmonary resistance and the resistive

work of breathing were lower and expiratory airflow higher in surfactant-treated individuals,

suggesting decreased residual bronchopulmonary damage (Abbasi et al 1993).

In BPD, the flow-volume curves of tidal breathing generated by the compression

technique were clearly abnormal, exhibiting severe flow restrictions (Tepper et al 1986).

Respiratory system compliance was low during the first months, returning to normal by 1-3

years. Specific compliance was also low, indicating that other factors apart from lung volume

were making the lung stiff. Resistance values were mostly high, indicating the severity of the

disease. SGaw was low (60% of predicted) in infancy and increased only slightly (to 70%)

during the first three years of life. FRC was reported to be low during the first six months of

life, exceeding the normal by 12 months, measured as nitrogen washout. (Gerhardt et al

28

1987b) Plethysmographic lung volumes, which represent all gas present in the chest, tended to

be higher, while FRC and the FRC/TGV ratio were lower in symptomatic than in

asymptomatic infants, suggesting true hyperinflation, as shown in LBW infants at 12 months

of age (Yuksel & Greenough 1991).

School age

The survivors of �old� BPD continued to have airway obstruction, bronchial hyperreactivity

and hyperinflation in lung function studies at school age (Smyth et al 1981, Bader et al 1987)

and even in adolescence (Northway et al 1990b). Catch-up improvement in abnormal lung

function during the early school years was also reported (Blayney et al 1991). Of the original

BPD group, 24% had fixed airway obstruction, and 52% had reactive airway disease in

adolescence; the BPD children had a higher Raw and a lower SGaw than those with no BPD

(Northway et al 1990b). Limited exercise tolerance, defined as reported symptoms or

measured abnormal oxygen saturation, was seen (Bader et al 1987, Northway et al 1990b).

DLCO was found to be normal (Smyth et al 1981) or slightly decreased (Northway et al

1990b). The duration of both endotracheal intubation and mechanical ventilation associated

significantly with values of lung function (RV/TLC, FRC). Lung injury in BPD was

suggested to have a role in the pathogenesis of later pulmonary dysfunction, as the reduction

of airway growth during the rapid postnatal phase of lung growth could contribute to

disproportionate undergrowth of the airway lumen, resulting in increased airway resistance.

(Northway et al 1990b)

A cohort of LBW children born in the late 1970´s, irrespective of neonatal respiratory illness,

were shown to have lower expiratory flow indices with well preserved VC than controls

representing local school children. Airway function correlated strongly with birth weight and,

to a lesser extent, with gestational age, maternal smoking, male sex and cough. The BPD

subgroup also had reduced FVC. (Chan et al 1989b) Four-week inhalation of beclomethasone

had no effect on symptoms or lung function in the 44% of the children who showed increased

airway responsiveness in histamine challenge (Chan & Silverman 1993). A Finnish cohort

study of children born very prematurely in the same period compared BPD, ventilated non-

BPD, non-ventilated and full-term control children. Despite few clinical problems at school

age, the BPD children had markedly lower SGaw and larger RV than the term controls,

explained by increased Raw, air trapping and reduced elasticity of the lungs. The non-

ventilated children did not differ from the controls. Contrary to the study of Chan et al,

29

neonatal oxygen treatment associated with more severely impaired SGaw, RV and FEV1.

(Hakulinen et al 1990) VLBW children showed increased airway reactivity, irrespective of

RDS, at 10-13 years of age, and DLCO was decreased in the non-RDS group (Galdès-Sebaldt

et al 1989). Later studies showed that VLBW children had a significant change towards

normal in FEV1 and RV in follow-up at 8-14 years. More improvement was seen in ELBW

children, who still had significantly more impaired values reflecting airflow, but not lung

volume and air trapping at age 14 years compared with term controls. Mechanical ventilation,

oxygen supplementation and BPD at 28 days had no effect on changes over time, but asthma

and exposure to smoking decreased the improvement. (Doyle et al 1999, Doyle 2000a)

Nikolajev et al (2002) assessed the lung function of discordant twins with LBW. They

showed that bronchial hyperreactivity was common in challenge tests at school age, but

small-for-gestational-age (SGA) was not associated with major impairment in lung function.

The increasing survival of more immature infants in the 1980´s focused the lung function

studies on VLBW, and oxygen supplementation at 36 weeks was mainly used as a definition

of BPD. These children had reduced values of airway flow, lower SGaw and lower DLCO than

controls, with associations with birth weight, suggesting structural changes in lung tissue due

to prematurity itself (Hakulinen et al 1996). BPD children had even more severely impaired

expiratory flow values and were more responsive to histamine than non-BPD children, which

was of clinical significance, as the latter finding associated significantly with respiratory

symptoms (Pelkonen et al 1997). In another study, impaired lung function was confirmed in

BPD, but not in non-BPD children born very prematurely, but their mean birth weight was

higher than in the former studies (Gross et al 1998). After severe BPD with a need for home

oxygen therapy, those with the most severely impaired expiratory flow had seriously

decreased diffusing capacity at the age of 11 years. Pulmonary dysfunction correlated with the

time of oxygen supplementation. Symptoms correlated with the reversible component of

airway obstruction rather than the fixed component. (Jacob et al 1998) In children born with

<800g of birth weight, low oxygen consumption in a treadmill test suggested a lower level of

fitness. Even with such a low birth weight and without apparent pulmonary abnormalities, the

children with BPD differed from the non-BPD group in the frequency of obstructive

abnormalities in lung function. (Kilbride et al 2003) After an exogenous surfactant trial,

spirometric parameters were better in children treated with surfactant compared to placebo,

suggesting that even in small preterm infants, advances in initial intensive care can improve

the pulmonary outcome (Pelkonen et al 1998).

30

7. Other indices associated with lung disease

Immune response

In asthma, the degree of inflammation is associated with the severity of basic lung function

impairment or the reactivity to bronchoconstrictors or exercise (Bousquet et al 1990, Venge

1994). The soluble markers of allergic inflammation have been reviewed by Venge (1994)

and the biopsy markers by Jeffery et al (2000b). Eosinophil-mediated damage of the

respiratory epithelium emerged as a major pathogenetic mechanism in asthma (Bousquet et al

1990), whereas COPD in adults involved a prominence of neutrophils (Lacoste et al 1993).

Merritt et al (1981) found a prominence of neutrophils and evidence of inflammatory injury in

the airways of newborn infants developing BPD. A secretion product of activated eosinophils,

eosinophil cationic protein, was elevated in the serum of asthma patients (Venge 1994).

Myeloperoxidase, secreted by activated neutrophils, was increased in bronchoalveolar lavage

fluid in COPD (Lacoste et al 1993). Lymphocytes were shown to be important in the

pathogenesis of both asthma (Robinson et al 1993, Azzawi et al 1990) and COPD

(O´Shaughnessy et al 1997, De Jong et al 1997), CD4 T-lymphocytes predominating in

asthma with eosinophils and CD8 T-lymphocytes in COPD with neutrophils and

macrophages, as reviewed by Jeffery (2000a).

Data on the inflammatory basis of lung function abnormalities at school age in children born

very prematurely are sparse. Treatment of low birth weight children with inhaled

glucocorticoids had no significant effect on baseline lung function or the airway response to

histamine, arguing against an inflammatory basis of the airway hyperresponsiveness in these

children (Chan & Silverman 1993). However, schoolchildren born very preterm were shown

to have a low CD4:CD8 lymphocyte ratio in peripheral blood in contrast to a normal

CD4:CD8 ratio found in asthma, bronchial lability associated with specific T-cell subsets and

elevated serum eosinophil cationic protein. The changes in lymphocyte subsets suggested the

immunological abnormalities to be related to BPD. (Pelkonen et al 1999)

Examination of induced sputum differential cell counts were a non-invasive method to

evaluate airway inflammation (Pavord et al 1997), but it is not an ideal routine test for

children, as it requires much work of skilled personnel and good cooperation of the patients.

31

Exhaled nitric oxide (eNO)

Nitric oxide (NO) gas is produced by various cells within the respiratory tract from L-arginine

in synthesis catalyzed by NO synthase (NOS) enzymes, as reviewed by Barnes & Belvisi

(1993) and Morris & Billiar (1994). There are at least three NOS isoenzymes: NOS1 (nNOS

or neuronal NOS), NOS2 (iNOS or inducible NOS) and NOS3 (eNOS or endothelial NOS)

(Yates et al 1995). All of the three isoenzymes have been detected in fetal lung and are

individually regulated (Sherman et al 1999, Aikio et al 2002). NO derived from constitutive

NOS (NOS1 and NOS3) is involved in the physiological regulation of airway function,

whereas NO derived from inducible NOS (in macrophages, endothelial, smooth muscle and

epithelial cells, fibroblasts and neutrophils) has an important role in respiratory inflammatory

reactions and host defence. NO is a potent vasodilator of bronchial circulation, a regulator of

pulmonary circulation and an important neurotransmitter in the neural bronchodilator system.

In 1991, Gustafsson et al first described NO in the exhaled air (eNO) of guinea pigs, rabbits

and human. Soon thereafter, elevated levels of eNO were reported in asthma patients (Alving

et al 1993, Kharitonov et al 1994, Persson et al 1994). The cellular origin of eNO was still

unclear. High concentrations were detected in the nasal air of normal subjects (Alving et al

1993), but when nasal contamination was prevented by expiration against resistance, eNO

was shown to originate from the lower respiratory tract (Kharitonov et al 1997). Oral

glucocorticoids reduced eNO in asthmatic patients, but not in healthy subjects (Yates et al

1995, Baraldi et al 1997). In patients with atopic asthma, inhaled glucocorticoids decreased

eNO (Kharitonov et at 1996), but did not prevent the increase in acute exacerbations in

children (Baraldi et al 1997). Viral respiratory infections raised (Kharitonov et al 1997) and

smoking reduced eNO (Persson et al 1994). In chronic obstructive disease (COPD), normal or

low eNO values were measured during the stable phase, but elevated levels might serve in

monitoring disease activity (Maziak et al 1998). In cystic fibrosis, low nasal and normal

exhaled NO values were reported (Lundberg et al 1996, Balfour-Lynn et al 1996). In a recent

study, eNO was superior to baseline lung function and bronchial responsiveness in identifying

preschool children with asthma (Malmberg et al 2003).

Infants with acute wheezy bronchitis had low eNO levels, while premature infants without

respiratory disease did not differ from term infants (Ratjen et al 2000). In ventilated preterm

infants, bronchoalveolar lavage fluid nitrate was similar in BPD, RDS and control children

during the first week of life, but remained high in those who later developed BPD (Vyas et al

1999). ENO levels were high during the second day in ventilated preterm infants. Despite the

32

absence of any significant association with the respiratory variables, the infants with the

highest levels manifested most severe BPD (Aikio et al 2002). Recently, high eNO

concentrations were related to BPD in face mask measurements at term (Leipälä et al 2004).

In evaluating the degree of airway inflammation in asthmatic children, eNO concentration and

sputum eosinophil counts were concordant. Thus, eNO was proposed as an easy-to-measure

marker of eosinophilic airway inflammation. (Piacentini et al 1999, Mattes et al 1999)

8. Follow-up of growth, neurodevelopment and cardiac function in children born

prematurely

Growth

In BPD children born in 1970-1980´s, growth retardation was common in early observations

at three years (Northway 1979) and in a follow-up at two years of age (Markestad &

Fitzhardinge 1981); in the latter study, it associated with severe and prolonged respiratory

dysfunction. In adolescence or early adulthood, those with BPD were significantly shorter and

weighed less than non-BPD individuals or term controls (Northway et al 1990b). Others found

no impairment of growth in severe BPD at the age of 10 (Bader et al 1987).

At school age, heights in relation to age among 20 children with BPD at 28 days were

significantly lower than those of term controls, and weights adjusted for height were also

lower. The non-BPD children did not differ from term controls. (Hakulinen et al 1996) The

children with BPD at 36 gestational weeks were shorter for age than those with no BPD or

controls (Pelkonen et al 1997). In a cohort study, VLBW children were shorter and lighter

and had smaller head circumferences at 14 years than those with normal birth weight, with

catch-up growth at 8-14 years. There were no differences in the stage of puberty. (Ford et al

2000) In that study, the ELBW girls were 4.5 cm and the ELBW boys 5.8 cm shorter at 14

years than the controls (Doyle 2000a), but no similar difference was detected at age 11 by

others (Kilbride et al 2003). In a 20-year follow up of VLBW children, boys were shorter and

lighter than controls at eight years, whereas girls were only lighter. Catch-up growth in

weight, height and body mass index occurred during 8-20 years of age among females; the

males remained smaller at 20 years, especially those who were SGA. (Hack et al 2003)

In the 1990´s, Barker presented a hypothesis of the fetal origin of coronary heart disease

(1995), and further studies consistently supported this finding of an association between low

33

birth weight, especially thinness at birth, catch-up growth in childhood and coronary disease

in adulthood (Forsén et al 1999). In a follow-up of growth, most VLBW infants born in the

surfactant era had not achieved the median birth weight of a fetus at the same gestational age

on hospital discharge. Appropriate-for-gestational-age infants, who did not develop BPD,

IVH, NEC or late sepsis, gained weight faster. (Ehrenkranz et al 1999) In a prospective study

of longitudinal growth, infants born at <29 gestational weeks had an initial period of poor

growth and a catch-up period at 4-5 years and mainly achieved normal weight and height by

seven years (Niklasson et al 2003).

Neurosensory development

Among the survivors of the original BPD group, grade repetition at school and abnormalities

in coordination, gait and muscle tone were reported; five out of 26 had CP, three had hearing

loss and one ROP compared with none of the controls (Northway 1990a). In a follow-up of 26

infants with BPD, 25% had low Bayley scores (<85) at 18 months, related to perinatal and

neonatal events rather than BPD (Markestad & Fitzhardinge 1981). In a population-based

Swedish follow-up cohort, the live birth prevalence of CP, especially spastic diplegia,

increased during the 1970-1980´s, reflecting the improved survival of VLBW infants

(Hagberg et al 1989). Later, no further increase was seen, but a slight decrease in preterm CP.

In 1991-1994, the prevalence was 81 per 1000 for infants of 1000-1499 g birth weight.

(Hagberg et al 2001) IVH and PVL were powerful predictors of CP (Hagberg et al 1996,

Salokorpi et al 1999). Despite the improved survival of more immature infants, the overall

incidences of CP and IVH have not increased. However, the risk of abnormal neurological

outcome increases sharply with decreasing birth weight. In a multicenter study (1995-1996),

30% of VLBW infants with brain ultrasound examinations (89%) had IVH, 11% had grade 3-

4 and 26% of the infants with 501-750 g birth weight had severe IVH. Cystic PVL was noted

in 5%. (Lemons et al 2001)

Besides IVH and CP, VLBW infants have an increased risk for other neurosensory

impairments, as reviewed recently by Tommiska [2003 (ELBW) and 2004a]. The families of

BPD children reported more need for physiotherapy and occupational therapy and more

concern for the child and impact of the child´s health on family life compared to non-BPD

children (Korhonen et al 1999a). At age 11, ELBW children were judged as occasionally

active or inactive by their parents, which differed from controls (Kilbride et al 2003).

34

Cardiac function

The severe �old� BPD was frequently accompanied by right ventricular hypertrophy and

cardiomegaly with cor pulmonale in electrocardiogram (Northway et al 1967, Markestad &

Fitzhardinge 1981, Northway 1990a) and by elevated right ventricular end-diastolic volume

and increased right ventricular wall thickness in echocardiography as late as school age

(Smyth et al 1981). After severe BPD with a need for home oxygen therapy, recurrent

episodes of heart failure were found in the children with most severely impaired lung function

(Jacob et al 1998). Pulmonary hypertension with increased pulmonary vascular resistance and

intrapulmonary shunting was demonstrated at cardiac catheterisation in children with severe

BPD at 3 months - 5 years of age associated with poor prognosis (Abman et al 1985, Berman

et al 1986, Bush et al 1990).

Pulmonary arterial pressure (PAP) can be assessed non-invasively by Doppler

echocardiography by measuring tricuspid valve regurgitation or pulmonary flow indices. PAP

is inversely related to the ratio between acceleration time (AT) and right ventricular ejection

time (RVET) (Kosturakis et al 1984). The VLBW children with BPD at 28 days born in the

late 1980´s had a slower drop in PAP than those without BPD, followed by an increase during

14-28 days of age (Gill & Weindling 1993). Furthermore, the children with BPD had lower

Doppler-mode-derived variables of pulmonary flow (AT, AT/RVET, pulmonary flow

velocity) at two years of age than those without BPD. The children with clinically severe BPD

and the most severe peripheral airway obstruction had the shortest acceleration time. (Farstad

et al 1995) In infants with RDS born in the 1990´s, the initially raised PAP remained

persistently elevated in a one-year follow-up in those who developed BPD. PAP was

inversely related to gestational age and corrected age at examination in preterm born infants

and to the duration of supplemental oxygen administration in infants with BPD. (Subhedar &

Shaw 2000) Previously, early findings of pulmonary hypertension were independently

associated with BPD at 36 weeks and suggested to be a sensitive marker of pulmonary injury

(Subhedar et al 1998). The cardiac findings in BPD have been recently reviewed by

Korhonen (2004a).

In children born in the late 1980´s or early 1990´s, no cardiac hypertrophy (Farstad et al 1995,

Korhonen et al 2004b) or significant pulmonary hypertension was seen at 2-7 years of age

(Korhonen et al 2004b).

35

9. Pharmacologic effects of glucocorticoids

Synthetic glucocorticoids, DEX and betamethasone, have a greater affinity for glucocorticoid

receptor than cortisol (7.1- and 5.4-fold higher, respectively), which additionally possesses

mineralocorticoid activity (Ballard & Ballard 1995, Jobe & Bancalari 2001).

Effects on the pulmonary defence systems

The initial studies in lambs suggested a response of the surfactant system to antenatal

glucocorticoids (Liggins 1969). Thereafter, all known components of surfactant have been

induced by treatment with glucocorticoids, presumably resulting in an increased content of

surfactant of normal composition and function, the effects depending on dosage and the stage

of lung development (Ballard & Ballard 1995, Vyas & Kotecha 1997). In the original

perinatal trial of our study population, after antenatal DEX, the requirements for surfactant

treatment were lower (Kari et al 1994), but no significant differences were seen in tracheal

fluid phosphatidylcholine despite the lesser need for ventilatory support (Kari et al 1995).

Another study showed an increase in surfactant proteins after postnatal glucocorticoid,

associated with improved initial lung function (Wang et al 1996).

In animals, glucocorticoids cause precocious increases in the activity of the antioxidant

enzymes superoxide dismutase, glutathione peroxidase and catalase. They regulate a

relatively small subset of proteins in lung cells, which act as important mediators of hormonal

effects. They induce both sodium-potassium-APTase and a subunit of the sodium channel

involved in the clearance of lung fluid from airspaces. (Ballard & Ballard 1995) Recently, in

preterm infants, the expression of epithelial sodium channel subunits was low but it was

increased by therapeutic doses of glucocorticoids (Helve et al 2004).

Glucocorticoids are among the most potent anti-inflammatory agents, but their role in lung

inflammation is poorly understood. DEX treatment has been shown to decrease nearly all

studied inflammatory indicators (Groneck & Speer 1995). In ventilated infants with BPD

(Yoder et al 1991) or RDS (Wang et al 1997), neutrophils and other anti-inflammatory

mediators were reduced in tracheal fluid, associated with improved pulmonary function, and

recently, early in RDS, downregulation of polymorphonuclear leukocytes and monocyte

activation was seen (Nupponen et al 2002). In rats, antenatal DEX has been shown to enhance

the abundance of endothelial NOS protein expression in lungs at birth (Lin et al 2001).

Recently, in ventilator-dependent infants, high eNO levels were shown to decrease after DEX

treatment (Williams et al 2004).

36

Effects on lung structure and growth

In animal studies, glucocorticoid treatment decreased the number of alveoli, which appeared

larger and more mature with reduced septal mesenchyme (Vyas & Kotecha 1997). In rats,

changes in the capillaries within the septa impaired the future formation of further alveoli

(Tschanz et al 1995, Tschanz et al 2003). Low-dose long-lasting postnatal DEX therapy

resulted in larger and fewer alveoli with reduced septal mesenchyme, leading to an

emphysematous condition, but a high-dose short-term treatment accelerated lung maturation,

followed by a step-back after discontinuation, with almost complete recovery, indicating that

both dosage and duration might be critical in terms of lung response (Tschanz et al 2003).

After antenatal glucocorticoids, the small but mature rat lungs at birth showed a growth

rebound and normal later development (Adamson & King 1988); others reported impaired

postnatal lung growth (Okajima et al 2001).

Retinoids are essential developmental mediators, and particularly all-trans retinoic acid

plays a key role in promoting septation. In neonatal rats, it also reversed the glucocorticoid-

induced inhibition of septation. (Massaro & Massaro 2000)

In sheep, single or repetitive maternal, but not fetal, betamethasone treatment caused

growth retardation persisting to term. Fetal treatment also improved lung function less,

suggesting that maternal glucocorticoids induce factors participating in lung maturation.

Repetitive treatment improved lung function more effectively than single treatment, but

concerns arose about decreased growth and neurodevelopment. (Jobe et al 1998) A review of

animal studies suggested that multiple courses of antenatal glucocorticoids improve lung

function, but decrease birth and lung weight and restrict brain growth (Aghajafari et al 2002a).

Scant data exist on human lung growth, mainly derived from lung function tests: no adverse

effect of antenatal DEX was seen in VLBW infants with little or no neonatal lung disease

during the first year of life (Wong et al 1982b).

Effects on other tissues

Glucocorticoids stimulate cytodifferentiation and cause precocious changes in different

tissues, accelerating the rate of differentiation without altering the sequence of developmental

events. They take part in glycogen metabolism, bile canaliculus formation, hematopoesis,

enzyme induction, intestinal maturation and ion exchange in kidneys and enhance the insulin

response to glucose and the adrenocorticotrophic hormone (ACTH) responsiveness of the

adrenal cortex, resulting in maturation of the catecholamine responses to birth in preterm

animals. The increased activity of myocardial adenylyl cyclase accounts for the improved

37

respiratory, cardiovascular and metabolic responses seen after treatment. (Ballard & Ballard

1995) The modulating effects on tissue differentiation involve interactions with other fetal

hormones, i.e. prolactin, thyroid hormones and catecholamines, insulin, EGF, interferon-γ and

testosterone. In humans, antenatal therapy with glucocorticoid and thyrotropin-releasing

hormone did not affect the incidence of RDS or BPD more than corticosteroid alone (Ballard

et al 1998), despite previous results (Ballard & Ballard 1995). Antenatal DEX decreased

plasma catecholamines, attenuating the birth-related increase (Kallio et al 1998, Banks et al

1999); others reported normal responses in tests stimulating human corticotrophin release at

one and two weeks after >8 courses and mild suppression in only some (Ng et al 1999). In

sheep, antenatal low-dose DEX suppressed the pituitary-adrenal function but augmented the

glycemic response to hypoxia (Fletcher et al 2000).

In human, concentrations of amniotic fluid cortisol and corticoid conjugates, primarily

derived from fetal cortisol, increase several-fold during the third trimester. Many infants with

RDS begin their recovery 48-72 hours after birth, which timing is consistent with the

accelerated production of surfactant in response to the elevation of cortisol that occurs in this

disease. Postnatal administration of hydrocortisone to infants with RDS did not improve their

clinical course, suggesting the postnatal elevation in endogenous levels to be sufficient to

induce lung maturity (Baden et al 1972). Thus, antenatal steroid is believed to mimic the

intrauterine and postnatal exposure to endogenous glucocorticoid (Ballard & Ballard 1995).

Very preterm infants may lack the capacity to produce enough cortisol to respond to

extrauterine stress, and infants who develop BPD were seen to have low cortisol levels and a

decreased response to ACTH after birth (Watterberg & Scott 1995). Low-dose hydrocortisone

increased survival without BPD in ELBW infants, when instituted shortly after birth,

particularly in those with chorioamnionitis (Watterberg et al 1999).

Brain development

Despite differences in the timing of the peak brain growth, the general sequence is similar in

animals and humans. Animal experiments have provided ample evidence of corticosteroid-

induced decrease in brain growth and abnormal development of the immature brain, but

depending on dosage and timing, glucocorticoids may either enhance or reduce brain injury

after a hypoxic insult (Huang et al 1999, Whitelaw & Thoresen 2000, Aghajafari et al 2002a).

In general, neurones in the human brain have stopped dividing by the third trimester, but

those in the dentate gyrus continue to divide long after full term and are thus vulnerable to

adverse influences. Despite the small molecular difference between betamethasone and

38

dexamethasone, their biological effects may differ. Among mice followed up into adulthood,

the antenatally betamethasone-treated group showed enhancement of memory compared with

the placebo group, whereas DEX treatment resulted in a decrement. (Rayburn et al 1997)

10. Trials on glucocorticoids during the antenatal - neonatal period

10.1. Glucocorticoid treatment of preterm infants

After initial discouraging attempts to prevent BPD in children with RDS by using

hydrocortisone (Baden et al 1972), DEX was adopted into use to facilitate the weaning of

VLBW infants from the ventilator (Mammel et al 1983, Avery et al 1985, Harkavy et al

1989, Kazzi et al 1990, Ohlsson et al 1992). The usual initial dose of DEX was 0.5 mg/kg/d

administered for 2-42 days with a different tapering schedule. In most studies, �open� steroids

were allowed afterwards. In a small study, a 42-day course at two weeks of age in infants

dependent on supplemental oxygen and mechanical ventilation was superior to an 18-day

course or placebo with respect to the pulmonary and neurodevelopmental outcome

(Cummings et al 1989). One study on oxygen-dependent infants at four weeks of age showed

more rapid improvement of ventilation in the DEX group, but no difference in the duration of

oxygen supplementation; new periventricular abnormalities on brain ultrasound scans were

reported (Noble-Jamieson et al 1989). A multicentre international trial, organized by

Collaborative Dexamethasone Trial Group (CDTG) (1991), studied the effect of 7-day

treatment on chronic oxygen dependency at three weeks of age in 287 VLBW infants born in

1986-1989. In the DEX group, the duration of further assisted ventilation was reduced, and

the infants with no mechanical ventilation at entry tended to need oxygen supplementation for

a shorter time. Hypertension and hyperglycemia were encountered. The Finnish study, in

which our study children participated, aimed to prevent BPD in infants dependent on

mechanical ventilation by earlier treatment at ten days of age. A shorter duration of

supplemental oxygen in DEX-treated girls, but not in all infants, and short-lived suppression

of basal cortisol were reported. Infants requiring ventilator at the age of ten days were at high

risk of developing BPD, as evaluated at the age of four weeks, and were thus not treated early

enough for the treatment to be truly prophylactic. (Kari et al 1993)

Apart from chronic oxygen dependency, with growing evidence of the inflammatory basis

of BPD (Yoder et al 1991, Groneck & Speer 1995), DEX was increasingly given soon after

birth to infants on mechanical ventilation for severe RDS. Pulmonary mechanics had already

39

been shown to improve and extubation by two weeks of age to increase among infants treated

within 12 hours after birth in an early study before the surfactant era (Yeh et al 1990). In an

Israeli multicentre trial of 248 preterm infants on mechanical ventilation with more than 40%

oxygen for RDS, treated with rescue surfactant, early postnatal DEX did not reduce the

incidence of BPD at either 28 days or 36 gestational weeks of age. The lesser need for

mechanical ventilation at the age of three days was not sustained at seven days of age.

Gastrointestinal hemorrhage associated with DEX. (Shinwell et al 1996) However, 43 VLBW

infants on mechanical ventilation showed significant improvement in respiratory compliance

and tidal volume, with further increase until seven days of age: the fraction of inspiratory

oxygen (FiO2) was reduced, weaning was facilitated, and there was less BPD at 36 weeks

(DEX 10%, placebo 47%). As the decreasing beneficial effect of surfactant increases the

susceptibility to barotrauma in the second week of life, the initiation of glucocorticoids during

this time could have minimized the lung injury and thus explained these results. (Durand et al

1995)

The initial goal was to rescue severely ill VLBW infants from prolonged mechanical

ventilation and to prevent BPD. With the increasing use in less severe disease or as

prophylaxis, there was growing concern of the long-term sequelae. Ng (1993) reviewed ten

postnatal DEX trials: hyperglycemia, hypertension, adrenal suppression and growth

impairment were common but usually reversible. Gastrointestinal bleeding, pulmonary air

leak, osteopenia, bone fracture, immunosuppressive effects and hypertrophic cardiomyopathy

were seen. Although no major neurodevelopmental risk was found in studies of antenatal

DEX, attention was raised to identify all treatable causes preventing successful weaning

before starting steroid treatment, such as anemia, PDA and sepsis.

Bhuta & Ohlsson (1998) reviewed ten studies of postnatal DEX treatment before 14 days of

life. The risk of BPD at 28 days or 36 gestational weeks was significantly reduced, especially

in infants treated at 7-14 days of age, in whom mortality was reduced as well. In this group,

only three children would need to be treated to prevent one infant from developing BPD.

More recent meta-analyses reviewed separately trials in which glucocorticoid, usually DEX,

was started either early (<96 hours), moderately early (7-14 days) or delayed (>3 weeks) after

birth (Halliday & Ehrenkranz 2000a, 2000b, 2000c).

40

Early postnatal glucocorticoid treatment

A meta-analysis of early postnatal corticosteroids for preventing BPD covered the following

studies: Baden et al 1972 (hydrocortisone), Yeh et al 1990, Sanders et al 1994, Shinwell et al

1996, Rastogi et al 1996, Suske et al 1996, Subhedar et al 1998, Wang et al 1997, Yeh et al

1997, Mukhopadhyay et al 1998, Tapia et al 1998, Garland et al 1999, Kopelman et al 1999,

Lin et al 1999, Romagnoli et al 1999, Soll et al 1999, Stark et al 1999, Watterberg et al 1999

(hydrocortisone) and Sinkin et al 2000 (Halliday & Ehrenkranz 2000a). Surfactant use was

common in most studies. DEX treatment did not reduce mortality, but facilitated weaning and

reduced the risk of PDA and pulmonary air leaks. Death or BPD and BPD at 28 days or 36

weeks were decreased, and the recipients of DEX required less steroids during the later

respiratory course. The risks of hyperglycemia, hypertension and growth failure,

gastrointestinal bleeding, intestinal perforation and hypertrophic cardiomyopathy were

increased, but no increase of NEC, infection or pulmonary hemorrhage was detectable. The

risk of severe IVH, PVL or severe or any ROP was not affected, either. The infants who

underwent cranial ultrasound scans showed a borderline increase in the risk of PVL (Soll et al

1999). In one study of a 3-day course in surfactant-treated RDS infants with a high risk for

BPD, IVH of any grade was more common in the placebo group (Garland et al 1999). Two

studies presented neurological follow-up data with increased risks of abnormal neurological

examination, developmental delay, CP and death or CP (Yeh et al 1998, Shinwell et al 2000).

A pilot study of low-dose hydrocortisone for 12 days in 40 ELBW infants revealed increased

survival without BPD at 36 weeks and reductions in the duration of oxygen supplementation

and mechanical ventilation with no detectable difference in side effects (Watterberg et al

1999).

In a recent study, early postnatal DEX was given as a 3-day course at a lower dose of 0.15

mg/kg/d, followed by 7-day tapering, to 220 ELBW infants on mechanical ventilation. The

risks of death and BPD at 36 weeks did not differ, but the DEX-treated infants were less

likely to receive supplemental oxygen at 28 days and open DEX later. Spontaneous

gastrointestinal perforation, especially with indomethacin use, was more common in the DEX

group, as were lower weight and smaller head circumference at 36 gestational weeks. (Stark

et al 2001)

Moderately early postnatal glucocorticoid treatment

A meta-analysis of moderately early postnatal corticosteroids for preventing BPD included

the following studies: Cummings et al 1989, Kari et al 1993, Brozanski et al 1995, Durand et

41

al 1995, Kovacs et al 1998, Papile et al 1998 and Romagnoli et al 1998 (Halliday &

Ehrenkranz 2000b). DEX treatment reduced mortality at 28 days, death or BPD and BPD at

28 days or 36 weeks and the need for later steroids and facilitated extubation, but had no

effect on the risk of pneumothorax or NEC. Adverse effects included hyperglycemia,

hypertension, gastrointestinal bleeding, hypertrophic cardiomyopathy, adrenal suppression

(Kari et al 1993) and infection. One study showed significant differences in pulmonary

mechanics and oxygen need at two days after DEX treatment with less BPD at 36 weeks

(Durand et al 1995). In a study of 3-day treatment followed by 18 days of nebulized

budesonide, the steroid group required less ventilatory support and supplemental oxygen at

the second week of age and had better pulmonary compliance at ten days, but the

improvements were not maintained, and the incidence of BPD at 36 weeks was not decreased

(Kovács et al 1998). There was no effect on the incidence of severe IVH or severe ROP, but

one study presented, apart from diminished weight gain, slowing down of head growth during

DEX treatment in both 2-week and 4-week groups (Papile et al 1998). Only one long-term

follow-up was available, with no increase in adverse neurological outcome (Cummings et al

1989).

Delayed postnatal glucocorticoid treatment

A meta-analysis of delayed postnatal corticosteroids for chronic lung disease included the

following studies: Ariagno et al 1987, Avery et al 1985, Harkavy et al 1989, Noble-Jamieson

et al 1989, Kazzi et al 1990, CDTG 1991, Ohlsson et al 1992, Vincer et al 1998 and Kothadia

et al 1999 (Halliday & Ehrenkranz 2000c). Surfactant was not used in most of the studies.

DEX treatment improved respiratory compliance, facilitated extubation and reduced the need

for oxygen supplementation and later steroids, but had no effect on survival or the duration of

hospitalization. Only one study reported the incidence of BPD at 36 weeks to be decreased

with borderline significance and death or BPD at 36 weeks to be decreased (Kothadia et al

1999). Poor weight gain was seen in the DEX group, but no increase in the risk of infection,

NEC or gastrointestinal bleeding. Abnormal neurological findings were increased in the

DEX-treated children, but moderate to severe neurological impairment was not, either overall

or in survivors. The increased risk of CP in survivors was of borderline significance, but in

keeping with the findings of early postnatal treatment. No increase in blindness was detected.

42

Neurodevelopmental outcome after postnatal dexamethasone treatment

A non-significant increase in cranial ultrasound abnormalities was reported after DEX

treatment (Noble-Jamieson et al 1989, O´Shea et al 1999, Shinwell et al 2000), and an �early�

trial of ELBW infants was discontinued because of adverse outcome, particularly PVL (Soll

et al 1999, the Vermont Oxford Network Steroid Study Group 2001). On the contrary, one

study reported more IVH in the placebo group (Garland et al 1999). In a study of ELBW

infants, postnatal but not antenatal DEX exposure was associated with abnormal brain

ultrasound findings and worse neurodevelopmental outcome at 18 months (LeFlore et al

2002). Poorer head growth was detected at the age of 30 days in infants treated moderately

early with 14-day DEX, compared to a 7-day early course (Romagnoli et al 1999). Later, even

an early 3-day course at a lower dose of 0.15 mg/kg/d associated with a smaller head

circumference at 36 weeks of gestational age in ELBW infants (Stark et al 2001).

Follow-up studies of randomized trials have been few in number. In the trial of Cummings et

al (1989), less abnormal neurodevelopmental outcomes (abnormal examination or Bayley

developmental index) were seen at 15 months of age after a 42-day course at two weeks (2

infants out of 9) compared to a 18-day course (7 out of 9) or placebo (3 out of 5), but another

small study of a 12-day course failed to reveal any difference in Bayley scores at two years of

age (Bhuta & Ohlsson 1998).

In the follow-up of 209 children oxygen-dependent at three weeks in the CDTG trial

(1991), no clear difference was evident at the age of three years in CP (20% vs 17%) or any

other neurological disability after a 7-day course, but less hearing loss was suggested in the

DEX group. Their disability score was described in our methods. Altogether 19% of the

infants had CP, 8% had visual loss, and 16% suffered from hearing loss; 19% were classified

as severely disabled and 35% as severely or moderately disabled; 18% were expected to need

special school education and 13% extra help at school. The need for physiotherapy (42%),

speech therapy (32%), occupational therapy (16%) and special schooling was common, but

similar in both groups. Underestimation of disability was suggested to be due to the methods

used. Neurological impairment was more common in the subgroup dependent on a ventilator

at recruitment than those on oxygen only. (Jones et al 1995)

In another trial of 118 infants, a 42-day course at 15-25 days of age increased the risk of

CP (DEX 25% vs placebo 7%) and abnormal neurological examinations, such as hypotonia or

possible or definite CP (45% vs 16%) at one year of age. Of the DEX-treated infants with CP,

44% had normal neonatal brain ultrasound scans. (Kothadia et al 1999, O´Shea et al 1999)

43

Similarly, in two trials of early DEX, a 3-day course increased the incidence of CP (49%

vs 15%), particularly spastic diplegia (28% vs 6%), and developmental delay (55% vs 29%) at

2-6 years of age (Shinwell et al 1996, Shinwell et al 2000), and a 4-week course increased

neurological abnormality, especially diplegia and hypotonia, at two years (40% vs 17%) (Yeh

et al 1997, Yeh et al 1998). In the study of Shinwell et al, all infants with PVL, 38% of IVH

grade 1-2 and 53% of IVH grade 3-4 developed CP; 22% of the children with CP had normal

neonatal ultrasound scans, all treated with DEX. In the study of Yeh et al, more fast activity

in electroencephalograms (EEG) was seen in the DEX group, with no correlation with the

outcome, but the overall abnormalities in EEG, brain stem auditory evoked potential and

visual evoked potential did not differ between the groups. The incidence of CP was twofold in

children treated with DEX compared to those with placebo. No difference was seen in either

visual (Yeh et al 1998, Shinwell et al 2000) or hearing problems (Shinwell et al 2000).

The differences in neurological outcome were not confirmed in a recent small follow-up

study at three years (Romagnoli et al 2002).

Barrington (2001) reviewed eight randomized controlled trials with neurodevelopmental

follow-up data of 679 survivors out of 1052 participants. According to the meta-analysis,

postnatal glucocorticoid was associated with an increase in CP and neurodevelopmental

impairment. The studies with less than 30% contamination with open steroid later showed a

greater steroid effect, and the relative risk of CP was 2.86 (95% confidence interval: 1.95-

4.19). The conclusion was to abandon the use of steroids for this indication in the absence of

evidence of any long-term benefits.

Growth after postnatal dexamethasone treatment

Poor growth and weight gain are common findings associated with DEX, but usually

reversible (Ng 1993, Halliday & Ehrenkranz 2000ac). Diminished weight gain was shown

during 2-week and 4-week courses (Papile et al 1998) and after a 2-week moderately early

and a one-week early course, but catch-up occurred soon (Romagnoli et al 1999). A recent

study of ELBW infants reported a significantly lower weight even after a low dose (Stark et al

2001).

In the follow-up of the CDTG trial (1991), no statistical differences emerged in growth at

the age of three years after 7-day postnatal treatment with DEX or placebo. Height, weight

and head circumference were below the average in both groups. (Jones et al 1995) After early

treatment, in a study of a 4-week course, which initially decreased the high need for oxygen,

44

somatic growth did not differ at two years of age in girls, but the DEX-treated boys were

shorter and weighed less than the controls (Yeh et al 1998), while a 7-day course had no

effect on growth at three years of age (Romagnoli et al 2002).

Respiratory morbidity and lung function after postnatal dexamethasone treatment

The short-term benefits of DEX to respiratory morbidity are evident and have been described

previously. The initial improvement of pulmonary mechanics (Yeh et al 1990, Durand et al

1995) was concomitant with the acute suppression of pulmonary inflammation (Yoder et al

1991), but resulted in only a modest decrease in the incidence of BPD at 36 gestational weeks.

In the follow-up of the CDTG trial (1991), despite improved weaning from the ventilator,

no beneficial or adverse effects of a 7-day course on respiratory morbidity were evident at the

age of three years. In all, based on questionnaires, 38% had frequent episodes of coughing or

wheezing on exercise, at night or with common cold, and 48% had had at least one admission

to hospital. In the DEX group, consultations for respiratory problems were more common and

an excess of cot deaths were reported, even though late mortality was similar in both groups.

(Jones et al 1995) However, a 4-week course of early DEX, which decreased the initially high

need for oxygen and the incidence of PDA, associated with a lower incidence of upper

respiratory infections and rehospitalization for respiratory reasons at the age of two years

(Yeh et al 1998); frequent rehospitalizations until the age of one year were also less common

after a delayed 42-day course (O´Shea et al 1999). The follow-up studies of randomized trials

did not assess lung function after treatment.

Cardiovascular findings after postnatal dexamethasone treatment

Hypertension, as a common finding, and cardiomyopathy or myocardial hypertrophy were

encountered after DEX treatment, but were usually reversible (Werner et al 1992, Ng 1993,

Evans 1994, Yeh et al 1997). However, DEX-induced fatal cardiomyopathy has been reported

(Riede et al 2001).

In the cardiac follow-ups during randomized trials of a 42-day (Kothadia et al 1999) and a

7-day course (Romagnoli et al 1999), DEX-associated ventricular septal hypertrophy and left

ventricular ourflow obstruction were usually reversible within one month, but had clinical

significance in some infants (Bensky et al 1996, Zecca et al 2001). Long-term follow-up has

not been reported.

45

Recommendations and practice of postnatal dexamethasone treatment

Despite the short-term benefits seen in infants treated moderately early, due to the initial

findings on neurological outcome, early postnatal treatment is not indicated as a treatment of

respiratory failure. Corticosteroids have been suggested to be reserved for infants not weaning

from mechanical ventilation and studies made on lower doses, other steroids, alternative

routes and late outcomes among survivors of randomized trials. (Halliday & Ehrenkranz

2000abc) Others proposed to abandon the use of steroids for this indication, as no evidence of

long-term benefits exists. In 2002, The American Academy of Pediatrics published guidelines

for the use of postnatal steroid treatment and stated that it should not be routine and should be

limited to �exceptional clinical circumstances�, such as infants on maximal ventilatory and

oxygen support. The publication of concerns reduced the use of postnatal DEX: in Israel, 22%

of infants were treated with open DEX after the age of seven days in 1994, compared with 6%

of similarly high-risk infants in 2001 (Shinwell et al 2003). Still, the Vermont-Oxford

Network data showed that 40% of ELBW infants in the participating institutions received

postnatal DEX by the age of 28 days in 1999 (Barrington 2001), and 48% of European units

also used DEX for non-intubated infants (Truffert et al 2003). The possible indications,

quality, dosage, duration and time of administration of corticosteroid for the prevention or

treatment of BPD remain one of the most controversial issues in neonatal medicine (Halliday

2003).

10.2. Antenatal glucocorticoid treatment

Prevention of neonatal diseases

After their introduction for fetal maturation 30 years ago, antenatal glucocorticoids were

slowly incorporated into clinical practice, due to the variable interpretations of the evidence

leading to conflicting advise, fear of long-term adverse effects and advances in the care of

VLBW infants. A meta-analysis of 15 randomized controlled trials showed them to reduce

mortality, RDS, IVH and NEC (overall reduction in the odds about 40%, 50%, 60% and 70%,

respectively) in infants born before 34 gestational weeks: Liggins & Howie 1972, Block et al

1977, Morrison et al 1978, Papageorgiou et al 1979, Schutte et al 1979, Taeusch et al 1979,

Doran et al 1980, Teramo et al 1980, Collaborative Group on Antenatal Steroid Therapy

(CGAST) 1981, Schmidt et al 1984, Morales et al 1986, Carlan et al 1991, Garite et al 1992,

Kari et al 1994 (Crowley 1995). The benefits were greater when the glucocorticoids were

administered 24 hours - 7 days before birth or even earlier, additively to those derived from

46

surfactant, and in case of premature rupture of membranes. Even treatment <24 hours before

birth decreased IVH (Kari et al 1994). Their effect on infection was equivocal, and no effect

was found on PDA (Eronen et al 1993, Kari et al 1994) or BPD. The few available follow-up

studies did not reveal adverse neurological outcomes (MacArthur et al 1982, CGAST 1984,

Smolders-de Haas et al 1990); on the contrary, protective effects were suggested (Crowley

1995). A study of an observational database of 35 000 VLBW infants supported the reduction

in IVH but not in NEC and reported a decrease in PDA and an increase in infection (Wright et

al 1995). In 1994, based on the meta-analysis, the NIH Consensus Development Conference

recommended antenatal glucocorticoids for women at risk of premature delivery (24-34

gestational weeks), as cost-saving therapies to reduce mortality and morbidity: either two 12-

mg doses of betamethasone given intramuscularly 24 hours apart or four 6-mg doses of DEX

12 hours apart, preferably 24 hours - 7 days before birth, but even later. The role in high-risk

conditions, such as hypertension and diabetes, was not established. Treatment was

recommended even with the available surfactant, as the latter alone had little or no impact on

the incidence of IVH or PDA. (NIH 1995)

Thereafter, maternal administration increased, even as repeated courses after seven days

for those who remained undelivered. Multiple courses were not recommended, as no

additional short-term benefits were seen in non-randomized studies, but instead, there was

increased mortality with lower gestational age (Banks et al 1999) and a trend towards more

BPD (French et al 1999). Randomized or quasi-randomized trials of 3700 infants showed a

consistently significant reduction in mortality, RDS and IVH with no adverse consequences

of a single course of betamethasone, DEX or hydrocortisone, but treatment >7 days before

delivery was not proved effective (Crowley 2000), as initially suggested by Liggins & Howie

(1972). In a further analysis of three randomized controlled trials of 551 infants (Guinn et al

2001, Aghajafari et al 2002b, McEvoy et al 2002), repeated doses decreased the risk of any

severe lung disease (Guinn et al 2001) and the need for surfactant (Guinn et al 2001, McEvoy

et al 2002). No difference was found in perinatal death, IVH or PVL. (Crowther & Harding

2003)

Neurodevelopmental outcome after antenatal glucocorticoids

Antenatal glucocorticoids prevent IVH (Crowley 1995, Wright et al 1995, Crowley 2000),

and the first follow-up studies also suggested protection against adverse neurological outcome

(MacArthur et al 1982, CGAST 1984, Smolders-de Haas et al 1990, Crowley 1995, Salokorpi

et al 1997). Data on animal studies (Jobe et al 1998, Huang et al 1999, Whitelaw & Thoresen

47

2000, Aghajafari et al 2002a) and postnatal glucocorticoids (Yeh et al 1998, Shinwell et al

2000) aroused concern about possible long-term effects on neurodevelopmental outcome. In a

prospective cohort study, repeated courses were associated with a 4% reduction in head

circumference at birth, representing a nearly 11% reduction in cranial volume (French et al

1999) and presenting an increased risk for abnormal neurodevelopment (Hack et al 1991). In

a retrospective study of 883 infants, DEX (11%) but not betamethasone (4.4%) was

associated with an increased risk of cystic PVL, a major cause of CP, compared with no

glucocorticoid treatment (8.4%) (Baud et al 1999). In the original trial of our study, abnormal

brain ultrasound findings, including PVL, were decreased (Kari et al 1994), and in a

prospective study of ELBW infants, they were not increased after antenatal DEX on hospital

discharge (LeFlore et al 2002). No association was found between brain weight and antenatal

glucocorticoids in preterm infants who died (Murphy 2001).

Few long-term follow-up results of randomized antenatal glucocorticoid trials continued until

school age or adulthood are available. In a Dutch follow-up trial of 81 adults born at 27-40

gestational weeks in the 1970´s, those treated with a single course of betamethasone did not

differ from placebo-treated controls in CP or cognitive function at the age of 20 years

(Schutte et al 1980, Dessens et al 2000). Previous follow-up studies of 250 out of 304

survivors after antenatal betamethasone (Liggins & Howie 1972, MacArthur et al 1982) and

406 out of 739 infants after DEX trial (CGAST 1981, 1984), revealed no negative

neurological outcome at six years or three years of age, respectively. In the previous

Auckland trial, betamethasone-treated girls scored better than control girls in school

behaviour. In the Collaborative study, mental and psychomotor development was assessed at

nine and 18 months and cognitive abilities at 36 months of age.

In an Australian cohort study, teenagers who had been exposed to antenatal betamethasone

in the early 1980´s showed better cognitive functioning at 14 years of age compared to

VLBW controls and tended to have less CP (Doyle et al 2000b). The cohort study of French et

al (1999) assessed 327 infants at three years of age: CP was found in 7.8% of the non-

corticosteroid group, 5.1% of the single-course group, and none in the group given repeated

courses. The disability rate did not differ, despite initial differences in head growth, but was

small in all groups.

48

Growth after antenatal glucocorticoids

Decreased birth weight has been associated with antenatal DEX (Bloom et al 2001) and with

multiple courses of glucocorticoids (Banks et al 1999). In the study of French et al (1999),

repetitive courses impaired fetal growth, but no difference persisted at the age of three years.

In the follow-up studies of randomized trials, no harmful effects on growth, i.e. height,

weight or head circumference, were reported at 3-20 years of age (MacArthur et al 1982,

CGAST 1984, Smolders-de Haas et al 1990, Dessens et al 2000). After antenatal DEX, body

weight and height were slightly greater at three years of age (CGAST 1984), and girls given

betamethasone were taller and heavier at six years (MacArthur et al 1982) than the controls in

the placebo group. The Dutch study reported a delay in puberty in boys (Smolders-de Haas et

al 1990).

In a non-random cohort study, the children exposed to antenatal betamethasone were

significantly taller than those not exposed, with no difference in sexual development (Doyle et

al 2000b).

Respiratory morbidity and lung function after antenatal glucocorticoids

Antenatal glucocorticoids halved the risk of RDS, but had no consistent effect on BPD at 28

days (Crowley 1995, Wright et al 1995, Crowley 2000). Nor did a case-referent study of 1454

VLBW infants born in the surfactant era show any change in BPD at 36 gestational weeks

(Van Marter et al 2001). Repeated doses have been shown to decrease the risk of severe lung

disease and the need for surfactant, but there are no data on long-term safety (Crowther &

Harding 2003).

In studies of lung function, infants treated with a single course of betamethasone were

shown to have higher FRC and better respiratory system compliance than matched untreated

infants within 36 hours, prior to surfactant treatment. No significant difference was found in

specific compliance or Raw, suggesting that there was recruitment of alveoli without

significant overdistension (McEvoy et al 2001). Compliance was higher after multiple courses

(McEvoy et al 2000).

Respiratory morbidity is infrequently assessed in follow-up studies of randomized trials.

More hospital admissions because of infections or respiratory diseases during the first year

were reported after betamethasone (Smolders-de Haas et al 1990) and more respiratory

complications at 18 months of age after DEX (CGAST 1984).

Data on the long-term effects of DEX treatment on lung function are sparse. Follow-up

studies of 75 children 10-12 years after a betamethasone trial (Smolders-de Haas et al 1990)

49

and of 20 children six years after a DEX trial (Wiebicke et al 1988) revealed no negative

effects of glucocorticoid on lung function. In a cohort study of betamethasone, no significant

differences were seen in lung function, assessed by spirometry and body plethysmography, at

the age of 14 years between those with and without exposure to glucocorticoids (Doyle et al

2000b).

Cardiovascular findings after antenatal glucocorticoids

In randomized trials comparing the fetal effects of antenatal glucocorticoids, betamethasone

reduced and DEX increased fetal heart rate variation (Mulder et al 1997), both induced

profound suppression of heart rate acceleration and variability at 48 hours, with a more

marked decrease after betamethasone (Rotmensch et al 1999), and the reversible fall in

variability was preceded by an initial increase with no differences between the groups (Subtil

et al 2003). All changes were reversible. In the meta-analyses of randomized studies, no effect

was found on PDA (Crowley 1995), but the observational database showed a decrease in

incidence (Wright et al 1995). No overall effect, but a beneficial effect on ductal closure in

infants born at <30 gestational weeks was reported (Eronen et al 1993).

Antenatal DEX did not appear to affect cardiac dimensions and blood flow velocity in

preterm infants during the first month of age (Skelton et al 1998). No studies exist on the

long-term effects of antenatal glucocorticoid on heart function or growth.

50

AIMS OF THE STUDY

In this follow-up study of two randomized trials, we investigated the effects of antenatal or

neonatal dexamethasone treatment on cardiopulmonary function, somatic growth and

neurosensory development at school age in children born very prematurely, by using different

lung function methods, cardiac evaluation, hospital data and questionnaires. Furthermore, we

wanted to assess the possible role of inflammation in bronchopulmonary dysplasia.

The specific aims of the present study:

I To assess the oscillometric method in detecting abnormalities of lung function

in children with or without bronchopulmonary dysplasia, compared with

conventional lung function tests.

II To study the possible inflammatory basis of abnormal lung function in

bronchopulmonary dysplasia by measuring exhaled nitric oxide concentrations.

III To evaluate cardiopulmonary function, growth and neurosensory development

after neonatal dexamethasone treatment for persistent respiratory failure.

IV To investigate cardiopulmonary function, growth and neurosensory

development after antenatal dexamethasone treatment for imminent preterm birth.

51

PATIENTS AND METHODS

1. Patients

The 49 children in the present study were born very prematurely in 1989-1991, at gestational

ages of <31 weeks or birth weights of <1500 g, and they were treated in the Neonatal

Intensive Care Unit of the Hospital for Children and Adolescents, Helsinki University Central

Hospital. They were included in one of two randomized double-blind placebo-controlled

trials: postnatal DEX therapy in VLBW infants at risk for BPD (Trial 1) (Kari et al 1993) or

prenatal DEX therapy in conjunction with exogenous surfactant therapy (Trial 2) (Kari et al

1994). They represent the major arms of multicentre trials, where the patients in Helsinki

were randomized as separate groups. Three children participated in both trials. Additionally,

one twin sister of a Trial 1 participant was included in the present study (I and II).

The entry criteria for Trial 1 were: 1) birth weight <1500 g; 2) gestational age >24 weeks; 3)

dependence on mechanical ventilation at ten days of age; and 4) no signs of ductus arteriosus,

sepsis, gastrointestinal bleeding or major malformation. A total of 23 infants received either

DEX (Oradexon®, Organon, Oss., The Netherlands) 0.5 mg/kg per day, divided into two

doses, for one week (DEX group) or an equivalent volume of saline (placebo group). Open-

label DEX therapy was allowed after 28 days of age at the discretion of the attending

neonatologist. During the initial hospitalization, three infants died: one boy in the DEX group

and one boy and one girl in the placebo group. The 20 surviving children (DEX 10, placebo

10) were invited to participate in this follow-up study: one child was excluded because of

neurological handicap, one family refused to participate and two could not be traced. Thus, 16

(80%) children participated in the present study (I, II and III). In addition, data on the

neurological outcome of all non-participants were collected from hospital records up to the

age of 5-7 years (III).

In Trial 2, 112 pregnant women with imminent preterm delivery before 32 weeks of gestation

were recruited and received either four doses of DEX (Oradexon®, Organon, Oss., The

Netherlands), 6 mg at 12-hour intervals intramuscularly (DEX group) or an equivalent volume

of saline (placebo group). 138 live-born babies were delivered (DEX 74, placebo 64).

Fiftynine infants were born at <31 weeks of gestation (DEX 37, placebo 22), of whom 14 died

52

during the initial hospitalization and one at five months of age (DEX 6, placebo 9). The 44

surviving children (DEX 31, placebo 13) were invited to participate in this follow-up study:

nine of them could not be evaluated, but data on their outcome were collected from hospital

records. A total of 35 (80%) children participated in the present study (DEX 87%, placebo

62%) (I, II and IV). Besides, the limited data available on the neurological outcome of 43

children at school age were analyzed (DEX 100%, placebo 92%) (IV).

Eighteen healthy non-atopic children born at term were recruited as controls for the lung

function tests. The study was approved by the institutional ethics committees. Written

informed consent was obtained from the parents.

2. Neonatal and follow-up data collection

Data on the initial hospitalization and supplemental oxygen requirements were collected in

order to characterize the severity of the infants´ primary respiratory and neurosensory disease.

The fraction of inspiratory oxygen (FiO2) was recorded at the postnatal ages of 2, 6, 12, 18,

24, 48, 72, 96 and 120 hours. The mean FiO2 requirements during 120 hours were calculated

on the basis of the areas under the curve. The duration of supplemental oxygen, FiO2 >0.4

requirements and dependence on supplemental oxygen at the ages of 28 days and 36 and 40

gestational weeks were recorded, as were the duration of ventilator treatment and initial

hospitalization. BPD was defined as a requirement for supplemental oxygen at a postnatal

gestational age of 36 weeks (Shennan et al 1988). The length of open-label DEX therapy after

28 days was recorded. The diagnoses of IVH (Papile et al 1978), PVL (Bejar et al 1986) and

ROP (International classification 1984) were established using standard methods. Abnormal

brain ultrasound findings were defined as IVH or PVL. PDA was regarded as significant if it

required indomethacin treatment or surgical closure. Small-for-gestational-age (SGA) infants

weighed <-2 SD from the mean birth weight for each gestational week (Pihkala et al 1989).

All hospital records were reviewed. Secondary hospitalization during infections, history of

wheezing and use of glucocorticoids or bronchodilators were regarded as signs of respiratory

morbidity. Major neurological problems and neuropsychological status at the age of five

years, major visual and hearing deficits and the conclusions drawn from school maturity

assessments were recorded. The diagnosis of CP was based on findings of abnormal muscle

tone, exaggerated tendon reflexes and a positive Babinsky sign and on persistent or

exaggerated primitive reflexes, dyskinesia or ataxia. The degree of disability was classified

53

according to Jones et al (1995): severe disability was defined as CP, severe global delay or

sensory or other impairment necessitating special school attendance. Moderate disability was

defined as mild CP, severe deafness, moderate global delay (extra help needed at school,

assessment of global retardation or language problems) or home oxygen use beyond three

years of age, but not severe enough to require education at a special school. Possible

cardiovascular medication or intervention was recorded.

3. Study protocol

All of the 49 children enrolled in the study were serially evaluated during 1998-1999 in the

Outpatient Department of Allergic Diseases, Helsinki University Central Hospital. At the first

visit, the principal investigator carried out the physical examination. The transformation of

body height into standard deviations (SDs) was based on the Finnish growth standards, with

standard curves constructed using group mean values and SDs for height at certain ages and

according to gender (Sorva et al 1990). Relative weight was calculated as the percentage

deviation from the median weight of Finnish children of the same height and gender.

Moderate or severe neurological impairment was recorded. The questionnaires focused on

respiratory symptoms and medication used during the preceding year and the history of

smoking or atopy in the family. Frequent respiratory symptoms were defined as dyspnea,

prolonged coughing for >2 months or recurrent coughing during exercise or at night. Impulse

oscillometry, flow-volume spirometry and pulmonary diffusing capacity test were done in the

Lung Function Laboratory of the Division of Allergy, Helsinki University Central Hospital.

The principal investigator, unaware of the neonatal history of each child at the time of testing,

supervised the lung function tests, supported the children to succeed in the test performance

and evaluated their co-operation for further lung tests. The children were examined clinically

when free of respiratory infections or allergic symptoms. Skin tests were performed by skin

prick with ten common environmental allergen extracts (birch, timothy grass, meadow fescue

and mugwort pollens, cat, dog, horse and cow epithelial danders, house dust mite Dermato-

phagoides pteronyssimus and spores of Cladosporium herbarum), negative vehicle and

positive histamine hydrochloride (10 mg/ml). Atopy was defined as at least one wheal

reaction >3 mm in diameter in the absence of a response to the negative control solution.

During the second visit, exhaled nitric oxide measurement and whole-body plethysmography

were performed in the Division of Clinical Physiology, Helsinki University Central Hospital.

54

The children were to have been clinically free of respiratory infections or allergic symptoms

for at least three weeks. Children on glucocorticoid inhalation therapy during infections were

examined more than three weeks after the discontinuation of their glucocorticoid, and those

on regular glucocorticoid inhalation therapy had a 24-hour glucocorticoid-free interval before

the measurement. β2-sympathomimetic agents were not allowed for 12 hours before the

measurement. Exposure to tobacco smoke and physical exercise were avoided on the same

morning.

During the third visit, a chest X-ray was taken, echocardiography was performed, and

electrocardiogram (ECG) readings were analysed by a pediatric cardiologist at the Division of

Pediatric Cardiology, Helsinki University Central Hospital.

.

4. Lung function methods

Flow-volume spirometry (I to IV)

Flow-volume spirometry was performed with a pneumotachometer-based spirometer

(Spirotrac III, Vitalograph Ltd, Buckingham, UK). A nose clip was used, and a visual

computer program helped to optimize the forced exhalations. At least three acceptable forced

expiratory curves were recorded according to the reproducibility criteria of the American

Thoracic Society (ATS) (1995). The curve with the highest sum of forced expiratory volume

in one second (FEV1) and forced vital capacity (FVC) was selected for the analysis. FVC,

FEV1 and forced expiratory flow at 50% of FVC (FEF50) were recorded, and the results were

expressed as percentages of predicted value (Polgar & Promadhat 1971). Spirometric

parameters were measured before and 15 min after the inhalation of 0.3 mg of salbutamol

(Ventoline®, Glaxo, Greenford, UK) via a large-volume plastic spacer (Volumatic®). The

bronchodilator response was expressed as the percentage change in FEV1 from baseline

(∆FEV1). Abnormal basic spirometric finding was defined as FVC<80%, FEV1<80% or

FEF50<62% of predicted values, and ∆FEV1>15% was considered as an increased

bronchodilator response.

Impulse oscillometry (I, III and IV)

Impulse oscillometry (Jaeger GmbH, Würzburg, Germany) (Vogel & Smidt 1994) was

performed, when the child was sitting and breathing quietly through a mouthpiece. A nose

clip was used, and the cheeks were supported by the investigator, to minimize pressure loss

55

through the upper airway shunt. The impulse interval was set to 0.3 seconds, and by using

Fast Fourier transformation, the resistance (Rrs) and reactance (Xrs) of the total respiratory

system were calculated as functions of an oscillation frequency of 5�35 Hz. The

measurements were repeated at least three times, and three acceptable, visually uniform

recordings were used to calculate a mean value for Rrs at 5, 10 and 20 Hz (Rrs5, Rrs10,

Rrs20) and for Xrs at 5 and 10 Hz (Xrs5, Xrs10) and the resonance frequency (fres; the

frequency where reactance has the value of zero). The results were expressed as SDs of

reference values (Duiverman et al 1985). IOS measurements were repeated 15 min after the

inhalation of 0.3 mg of salbutamol. The bronchodilator response was expressed as the

percentage change in Rrs5 from baseline (∆Rrs5). Abnormal IOS findings were defined as

Rrs5>1.65 SD or Xrs5<-1.65 SD, and ∆Rrs5>30% was considered as an increased

bronchodilator response.

Pulmonary diffusing capacity test (I to IV)

Pulmonary diffusing capacity and static lung volumes were measured using the single-breath

method (Cotes et al 1993) (PFT, Jaeger GmbH, Würzburg, Germany). A volume of 90% of

inspired vital capacity was used for breath holding; the washout volume was preset at 500 ml

and the sample for expiratory gas fraction was 750 ml. Mean values of three acceptable

measurements were used for the analysis of pulmonary diffusing capacity (DLCO) and the

diffusing coefficient (KCO) for carbon monoxide. The results were expressed as percentages of

the predicted values of Polgar & Promadhat (1971) for static lung volumes (I) and for

diffusing capacity (II), and as percentages of the predicted values of Cotes et al (1979) for

diffusing capacity (I, III and IV).

Whole-body plethysmography (I)

Whole-body plethysmography (Body-Screen II, Jaeger) was performed for those who showed

sufficient cooperation in previous lung function studies. The mean values of three acceptable

measurements of total lung capacity (TLC), residual volume (RV), airway resistance (Raw)

and specific conductance (SGaw) were recorded, and the RV/TLC ratio was recorded to assess

hyperinflation. The reference values of Polgar & Promadhat (1971) were used for the

analysis.

56

5. Exhaled nitric oxide (II)

Exhaled nitric oxide was measured using a chemiluminescence analyzer (Sievers 270B,

Boulder, CO) (Kharitonov et al 1997) with a rapid-response time (<200 ms) and an accuracy

of +1 ppb (parts per billion). In addition, the analyzer also measured expiratory flow and

exhaled volume in real time. The sampling rate through the reaction chamber of the analyzer

was 250 mL/min for all measurements. The analyzer was calibrated daily, using NO-free

oxygen to set the absolute zero and certified NO gas of 184 ppb NO in nitrogen (AGA

Edelgas, Germany).

For eNO measurements, after inhaling NO-free oxygen, the children exhaled slowly from

TLC for 15 seconds through a flow resistor (Hans Rudolp, Model 7100R, 50cmH20/L/s, flow

range 0-0.5 L/s), which created a positive pressure necessary to close the soft palate. Due to

the small lung volumes of the children, the target flows recommended by the European

Respiratory Society (ERS) (Kharitonov et al 1997) (0.16-0.25 L/s) appeared to be unsuitable,

as they presumed a volume of at least 2.4-3.7 L for 15-second exhalation. Therefore, lower

target flows were used in this study (0.10-0.20 L/s). A visual computer program was used to

help to maintain the flow as fairly constant. A nose clip was used, and the cheeks and lips

were supported to prevent any air leak. During a steady exhalation, eNO reaches a plateau

after 5-10 seconds, while the exhaled volume continues to increase. The mean value of the

last three seconds of the end-expiratory plateau of NO concentration was taken for analysis.

The results of the analyses were computed and graphically displayed on a plot showing eNO

levels, flow and volume against time and expressed as eNO concentration (ppb) and NO

output (pmol/s). At least three successive recordings were made, with a coefficient of

variation of <10%. The mean value of the measurements was recorded on the case record

form.

6. Cardiological measurements (III and IV)

ECG criteria for right ventricular hypertrophy were defined as R wave >20 mm or >S wave at

lead V1. Incomplete right bundle branch block (RBBB) was registered. Cardiac volume was

derived from the size of the heart on plain chest x-ray. The anatomy and function of the heart

and the large vessels were studied using two-dimensionally guided, pulsed, continuous-wave

and colour Doppler equipment (Acuson 128 XP machine, Mountain View, CA, USA) with

2.5, 3.5 and 5.0 MHz probes and recorded on videotape. Pulmonary arterial pressure was non-

57

invasively assessed by measuring tricuspid valve regurgitation flow (Farstad et al 1995,

Subhedar et al 2000). In the main pulmonary artery, peak systolic velocities (cm/s) and

acceleration time (ms) were registered.

7. Statistics

The results of lung function tests were expressed as percentages or SDs of predicted values. In

Study I, ANOVA with Bonferroni/Dunn´s application for multiple comparisons was used to

compare the lung function test results of the thre groups. Pearson´s correlation coefficient was

calculated for the relationship between the different lung function tests. Kappa (κ) statistics

were used to describe the agreement between spirometry and the IOS method, and

McNemar´s test to compare the agreement of oscillometric variables with spirometry. A p-

value <0.05/3=0.017 was considered significant in group comparisons, otherwise p<0.05. In

the Studies II to IV and in supplemental analyses of the whole population, the Kruskall-Wallis

non-parametric test was used to determine the significances of the differences between the

three groups. The Mann-Whitney test and the χ2 test with continuity correction were used for

the comparisons between two groups. P<0.05 was considered significant. Spearman´s rank

correlation was used to evaluate the association between individual values of neonatal data,

lung function and cardiological measurements.

59

RESULTS

1. Population characteristics

Children born very prematurely (All, I to IV)

The basic perinatal and neonatal data of the study children and the different subgroups are

presented in Table 2. The children were born at 24.1 to 30.9 gestational weeks with birth

weights of 600-1575 g (I to IV). 45% of them were male and 12% SGA. 55% had received

antenatal DEX. All except one child required mechanical ventilation after birth, which was

provided by a pressure-limited Baby Bird ventilator (Bird Corporation, Palm Springs, CA,

USA). 84% had RDS, and 43% received surfactant for treatment. 6% had air leak syndrome.

73% required indomethacin and 6% surgical ligation for PDA. At the age of 28 days, 29%

were on ventilator support and 63% on supplemental oxygen. 33% were on supplemental

oxygen at 36 gestational weeks and 22% at 40 gestational weeks. 33% received postnatal

DEX before two months of age for weaning from the ventilator. 22% had IVH grade 1-4, 8%

IVH grade 3-4 and 28% abnormal brain ultrasound findings. 6% had NEC, 12% had ROP,

and 8% needed cryocoagulation (not shown in the original studies).

Children with BPD (All, I and II)

Apart from the more long-term need for supplemental oxygen, the BPD children had

significantly lower gestational age (I) and birth weight and a longer need for ventilator

treatment and initial hospital care than those with no BPD (I and II). Subsequent hospital data

on oxygen need caused one child to be re-classified into the BPD group after the completion

of Study I, and therefore, in the subsequent studies, 16 children had BPD and 33 did not.

Atopic children (II)

The difference in gestational age was not significant, but the atopic children had higher birth

weight and a shorter need for initial respiratory support and hospital care (p<0.01) than the

non-atopic children.

Children in the postnatal dexamethasone study (III)

The DEX and placebo groups did not differ in basic neonatal data. One child in each group

had received antenatal DEX, and 3 out of 16 received surfactant therapy for RDS. Half of the

61

children in the placebo group and 25% in the DEX group received open-label DEX treatment

after the trial. Half of the children in each group had BPD.

Children in the antenatal dexamethasone study (IV)

Abnormal brain ultrasound findings were more common in the placebo group than in the

DEX group (p=0.02), which was the only significant difference in the neonatal data. 33% of

the infants in the DEX group and all in the placebo group received indomethacin for PDA

(p=0.06); nor was the difference in surfactant therapy (DEX 52%, placebo 88%) or ROP

(DEX 7%, placebo 25%) significant. All of the 6 SGA infants were in the DEX group. 11% in

the DEX group and 38% in the placebo group received postnatal DEX treatment. One fourth

in each group had BPD.

2. Growth

Children born very prematurely (All, I to IV)

The demographic data of all study children and the non-atopic control children born at term

are presented in Table 3. The study children did not differ significantly from the controls in

age, height, weight (I and II) or relative weight. However, standardized height was lower in

the prematurely born children than in the controls (p=0.03 in II; p=0.008 in All); this

difference was significant in boys (p=0.007 in All, not shown in the studies), but not in girls.

Children with BPD (All, I)

Despite their lower gestational age and birth weight, the BPD children did not differ

significantly at school age in height, weight (I) or standardized height, but they weighed less

after adjustment for height and gender than the non-BPD children (p=0.03, not shown in the

studies).

Children after postnatal dexamethasone treatment (III)

The children treated with DEX did not differ significantly from the placebo-treated children

or the controls in height, standardized height, weight or relative weight. Standardized height

was lower in the placebo group than in the controls (p=0.006). No significant correlation was

found between the duration of DEX therapy before one year of age and the variables of

growth (not in Tables).

62

Children after antenatal dexamethasone treatment (IV)

Height, standardized height, weight and relative weight did not differ significantly between

the children given antenatal DEX or placebo and the controls. Standardized height showed a

tendency towards a difference (p=0.05) between the three groups; further comparisons proved

it to be lower in the placebo group than in the control group (p=0.03). Adjusted for gender,

this difference was significant in boys (p=0.01) (not shown in the studies).

3. Respiratory symptoms and medication

Children born very prematurely (All, I, III and IV)

Table 4 shows the respiratory symptoms and medication of the study children in the different

subgroups. 33% received glucocorticoid inhalations at 12 months and 37% at 24 months of

age. One child was on oral systemic glucocorticoid until the age of 7.5 years. Adenoidectomy

was performed on 57% of the children, and 39% needed tympanostomy tubes. Regarding the

symptoms during the preceding year, 22% were reported to have asthma, 29% dyspnoea, 35%

prolonged cough for >2 months, 12% cough during the pollen season, 14% on exercise and

18% at night. A total of 49% reported either dyspnoea or prolonged cough. Glucocorticoid

inhalations were used regularly by 5 (10%) and occasionally by 8 (16%) children. Combined

with glucocorticoids, 4% used bronchodilators and 4% chromone inhalations regularly. 41%

used β2-sympathomimetic agents. None had required hospitalization because of respiratory

symptoms during the preceding year. Previous diagnosis of asthma, i.e. a history of asthma

symptoms and regular therapy, correlated significantly with the use of glucocorticoids and

bronchodilators and cough during the pollen season, but not with cough on exercise or at

night, nor was the correlation with frequent symptoms during the preceding year significant.

Dyspnoea or prolonged cough associated with lower age (p=0.02) and shortness for age

(p=0.02) at the time of the study, but not with gestational age or birth weight. The use of

glucocorticoid inhalations associated with lower age (p=0.007), low birth weight (p=0.02) and

need for ventilator treatment (p=0.007), oxygen supplementation (p=0.02) and initial hospital

care (p=0.02) (not shown in the studies). Worse exercise tolerance compared to others and at

school, subjectively assessed by parents, was reported for 14 children: a correlation was found

with shortness for age (p=0.046), low birth weight (p=0.03), ventilator treatment (p=0.03),

oxygen supplementation (p=0.009) and initial hospital care (p=0.01). Only 3 of these 14

children were free from severe or moderate disability (p=0.001) (not shown in the studies).

Two children received medication for epilepsy (I).

64

sympathomimetic agents tended to be (p=0.07) more common in the children with BPD

during the preceding year. 38% of the BPD and 45% of the non-BPD children had had neither

prolonged respiratory symptoms nor inhalation therapy during the preceding year (not shown

in the studies).

Children after postnatal dexamethasone treatment (III)

In the DEX group, 50% received glucocorticoid inhalations at 12 months and 63% at 24

months, and in the placebo group, 50% and 50% did so, respectively; in addition, one child in

the latter group received oral systemic glucocorticoid by the age of 7.5 years. Frequent

respiratory symptoms during the preceding year were reported by 38% in each group. In the

DEX group, none received regular glucocorticoids, although occasional use of inhalations

was common. In the placebo group, one child out of 8 was on regular glucocorticoid

inhalation, and 2 used β2-sympathomimetic agents occasionally. One fourth in each group had

had neither prolonged respiratory symptoms nor a need for inhalations during childhood. No

significant differences were seen in respiratory symptoms or medication associated with

postnatal DEX treatment.

Children after antenatal dexamethasone treatment (IV)

In the DEX group, 26% received glucocorticoid inhalations at 12 months and 26% at 24

months, compared with 25% and 50% in the placebo group. Half of the children in each group

reported frequent respiratory symptoms during the preceding year. In each group, 2 children

were on regular glucocorticoid inhalation (DEX 7%, placebo 25%), and β2-sympathomimetic

agents were used commonly (DEX 44%, placebo 38%). The 2 children on regular inhaled

bronchodilators were in the placebo group (p=0.07). One fourth in each group were free from

frequent respiratory symptoms and inhalations until school age. Antenatal DEX treatment did

not associate with respiratory symptoms or medication.

4. Smoking in the family (I to IV)

Exposure to tobacco smoke was common in this series of children born very prematurely,

defined as smoking during pregnancy (33% from hospital records, 25% reported), maternal

smoking (39%), smoking within the family (45%) or smoking at home (6%) (not shown in the

studies), with no difference between the subgroups in individual studies (III and IV) (I and II,

not shown in the studies).

65

Influence on growth

No significant correlation emerged between maternal smoking or smoking during pregnancy

and growth (III) (in All, not shown in the studies).

Influence on respiratory symptoms

No associations were found between maternal smoking or smoking during pregnancy and

dyspnea or prolonged cough (III and IV). Of the 15 study children with no prolonged

respiratory morbidity and no need for inhalation during childhood, 60% had smoking

mothers, and 47% had mothers who had smoked during pregnancy (not shown in the studies).

Influence on lung function

Maternal smoking or smoking during pregnancy was not associated with basic lung function

as assessed by spirometry or oscillometry. However, maternal smoking or smoking in the

family associated with poorer diffusion capacity at school age (see results in page 73).

5. Atopy

Children born very prematurely (All, II)

30% of the families reported having medically diagnosed atopy and 32% probably atopic

symptoms; 39% had animals. 12 out of 48 (25%) study children were skin prick-positive (not

shown in the studies); one child refused to be tested. The atopic children did not differ

significantly from the non-atopic children in gestational age (p=0.06), but they had higher

birth weight (p=0.001) and a shorter duration of ventilator treatment (p=0.001), oxygen

therapy (p=0.007) and initial hospital care (p=0.001) (II; p<0.01 in All, not shown in the

studies). None of the skin prick-positive children had received postnatal DEX, differing

significantly from the skin prick-negative ones (p=0.01). Prick positivity correlated neither

with respiratory symptoms nor with medication (not shown in the studies).

Children with BPD (All, II)

In Study II, only one out of 12 (8%) BPD children had positive skin prick tests compared with

9 out of 27 (33%) non-BPD children. In the whole group of study children, this finding of less

atopy in the BPD group (1 out of 16) compared with the non-BPD group (11 out of 32) was

statistically significant (p=0.04) (not shown in the studies).

68

Xrs5 were significantly associated with FVC and FEF50 as well. Rrs5 was significantly

associated with Raw (r=0.63, p<0.0001), but showed higher values than Raw at low values of

resistance and lower values at resistance values >1 kPa/L/s. Xrs5 was significantly related to

RV (r=0.43, p<0.01) and RV/TLC (r=0.57, p=0.0004), but not to either DLCO or KCO.

Low FEV1 was associated with increased Rrs5 in 10 and with decreased Xrs5 in 13 out of

18 children; correspondingly, normal FEV1 was associated with normal Rrs5 in 22 and with

normal Xrs5 in 27 out of 30 children. Rrs5 agreed with FEV1 at a low level (κ=0.29, 95%

confidence interval (CI): 0.00-0.57) and Xrs5 at a moderate level (κ=0.64, 95% CI: 0.41-

0.87). Xrs5 yielded significantly more concordant information with spirometry than Rrs5

(p=0.02). ∆FEV1 correlated significantly with the bronchodilator response in Xrs5 (p=0.02)

and inversely with the initial FEV1 (p=0.0002) (not shown in the studies).

Children born very prematurely (I and II)

The children born very prematurely showed significant differences in basic lung function

adjusted for height-dependent predicted values compared with the full-term controls: in

spirometry, the FVC, FEV1 and FEF50 values were lower (I and II) and the bronchodilator

response higher (II); in IOS, Rrs5, Rrs10 and fres were higher and Xrs5 and Xrs10 lower than

in the controls (I); in the diffusing capacity test, DLCO (II), KCO and VC were lower and RV

higher than in the controls (I). Thus, pulmonary function studies showed obstructive

ventilatory defects, increased airway resistance, decreased airway reactance, pulmonary

hyperinflation and decreased diffusing capacity among preterm children compared with full-

term controls. The use of inhaled medication during the preceding year associated with

impaired lung function: both glucocorticoids and β2-sympathomimetics with FEV1 (p=0.0003

for glucocorticoids, p=0.0008 for β2-sympathomimetics), FEF50 (p=0.0001 and p=0.005,

respectively), Rrs5 (p=0.01 and p=0.002), Xrs5 (p=0.003 for both), Raw (p=0.02 for both) and

SGaw (p=0.002 for glucocorticoids, p=0.01 for β2-sympathomimetics), but not with the gas

exchange variables. Frequent respiratory symptoms did not correlate with the values of lung

function tests. Worse exercise tolerance compared to others and at school, assessed by

parents, associated with lower FEV1 (p=0.02) and SGaw (p=0.04) and tended to correlate with

lower FVC (p=0.057) and FEF50 (p=0.055); no correlations were found with the variables of

IOS or diffusing capacity.

Twentythree out of 48 (48%) children had abnormal findings in basic spirometry and 21

out of 48 (44%) in IOS (Figure 2); the bronchodilator response was significant in 8 out of 45

(18%) in FEV1 and in 17 out of 47 (36%) in Rrs5.

71

Children after postnatal dexamethasone treatment (III)

Spirometry and IOS showed significant impairment in pulmonary function in both study

groups compared with the controls´ values (Table 6). FVC was higher in the DEX group

(p=0.04), this being the only significant difference between the DEX and placebo groups. In

IOS, Rrs5 was higher in the placebo group and Xrs5 was lower in the DEX group as signs of

airway obstruction, and the bronchodilator response was increased in both groups compared

with the full-term controls. DEX treatment was not shown to have any negative effect on

pulmonary outcome.

Children after antenatal dexamethasone treatment (IV)

The study children had significantly impaired basic spirometric values and increased ∆FEV1,

suggesting bronchial lability (Table 6). The more pronounced bronchodilator response in the

placebo group was the only significant difference between the study groups in height-adjusted

spirometric values (p=0.01). IOS revealed no significant differences between the DEX and

placebo groups. Compared with the control group, Rrs5 was significantly higher and Xrs5

lower in both study groups. Diffusing capacity test failed in 7 DEX and 3 placebo children

and in 5 controls, thus rendering the comparisons between the preterm groups invalid.

However, the mean DLCO and KCO values were significantly lower in all study children than

in the controls. Antenatal DEX treatment was not shown to have any negative effect on

pulmonary outcome.

Effect of smoking

Basic spirometric, oscillometric and body-plethysmographic values did not correlate with

maternal smoking or smoking during pregnancy (III and IV). Nor did the values of the

diffusing capacity test correlate with smoking during pregnancy. However, in Study IV, the

prematurely born children with smoking mothers had significantly lower DLCO values

(p=0.009) and a slight tendency towards lower KCO values (p=0.08) compared with the

children of non-smoking mothers. Smoking in the family also tended to correlate with lower

DLCO (p=0.08) and KCO (p=0.08) (IV); the difference in KCO was significant in all study

children (p=0.02, not shown in the studies).

74

Influence of atopy, familial smoking or glucocorticoid inhalation

In the prematurely born group, the atopic children had higher mean (SD) eNO values than the

non-atopic ones [14.8 (14.2) vs 6.3 (4.5) ppb, p=0.02] and higher mean (SD) NO outputs of

68.7 (54.6) and 36.8 (33.8) pmol/s, respectively (p=0.04) (Figure 3). The eNO concentrations

and NO output values of the non-atopic study children did not differ significantly from those

of the controls. Gender and exposure to smoking in the family were not significant predictors

of eNO concentrations or output in the study children, nor was the use of inhaled

glucocorticoids during the first two years of life or during the preceding year.

Non-atopic children with BPD

Due to the significant association of atopy with increased eNO in our study and of

glucocorticoids with decreased eNO in previous studies, we excluded the 10 atopic children

and the 3 children on regular glucocorticoid inhalation therapy from the further analyses. Of

the remaining 27 non-atopic study children, 9 had a history of BPD and 18 had no BPD.

These two groups did not differ in gestational age or demographic data at the time of the

study, but the BPD children had lower birth weights and needed more oxygen

supplementation, as by definition, and ventilator treatment (Tables 2 and 3). Despite

significant differences in spirometry, the eNO concentrations did not differ between the BPD,

non-BPD and control children [6.8 (4.6), 5.9 (4.8) and 6.4 (4.3) ppb, respectively], nor did the

eNO output values [48.7 (47.9), 29.5 (23.6) and 30.8 (24.8) pmol/s] (Figure 3). In the non-

atopic children born very prematurely with a history of BPD, we found no evidence of airway

inflammation associated with increased eNO concentration. No association was found

between eNO levels and the severity of chronic lung disease, determined by conventional

lung function tests, either.

8. Cardiac function

Children after postnatal dexamethasone treatment (III)

No significant difference between the study groups was seen in cardiac volumes, Doppler-

mode-derived variables of pulmonary flow (peak velocity or acceleration time) or left

ventricular function (Table 8). Tricuspid valve regurgitation of >30 mmHg was detected in 3

children in the DEX group and one child in the placebo group. The placebo group child had

elevated pulmonary arterial pressure of 42 mmHg in cardiac catheterization at three years of

age and was treated with digoxin and nifedipine. In the follow-up at eight years of age,

76

cardiomyopathy was seen. One girl in the DEX group had a tiny PDA 1.5 mm in diameter. No

murmur was heard, and no closure was needed. None required cardiovascular medication.

51% of the children born prematurely presented with RBBB in ECG. These children had

more severely impaired values of FEV1 (p=0.03), Rrs5 (p=0.02) and Xrs5 (p=0.02) than those

without RBBB. However, RBBB did not associate with BPD or PDA.

9. Neurological follow-up

Children after postnatal dexamethasone treatment (III)

At the age of five years, 50% of the children in each study group needed physiotherapy,

occupational therapy or evaluation for the most appropriate type of school because of

neurological impairment or difficulties in speech, learning or comprehension. All were able to

walk without aid or support. All of the children in the DEX group attended normal school,

and 3 of them had CP: one girl with mild, spastic diplegia and mild cognitive difficulties

needed a personally adjusted course in mathematics as well as physiotherapy; one girl with a

shunted hydrocephalus, epilepsy, mild, spastic diplegia and strabismus needed a personal

assistant at school (IVH grade 3-4); one boy with mild, spastic hemiplegia needed

physiotherapy, and one boy with no previous problems needed speech therapy because of

reading difficulties (Table 9). In the placebo group, 7 (88%) children attended normal school,

and 2 of them had CP: one girl who had received oxygen therapy for 4.5 years had a personal

assistant because of moderate spastic diplegia, and she also needed physiotherapy, and one

boy with mild, hypotonic diplegia had a personal assistant and had problems in mathematics

(IVH grade 2-3), but spoke three languages. One girl attended a preparatory class before

entering normal school. Another mentally retarded girl attended a special school for blind

children because of a severe bilateral visual deficit caused by ROP.

Thus, 38% of the children in the DEX group and 25% in the placebo group had CP. In the

DEX group, 38% were moderately disabled, while in the placebo group, 25% were severely

and 25% moderately disabled. The neurodevelopmental outcome at school age did not differ

significantly between the DEX and placebo groups.

The data from the hospital records of the 4 children lost to this follow-up did not add to

any differences between the groups, as no CP was found, all could walk without support and

attended or planned to attend normal school, and one child in each group was moderately

disabled.

78

Thus, 11% of the children in the DEX group and 50% in the placebo group had CP. 22%

of the children in the DEX group and 75% in the placebo group were either severely or

moderately disabled. In the DEX group, the rate of normal school attendance was higher

(p=0.01), and there were fewer instances of severe to moderate disability (p=0.02) and a

tendency towards a lower incidence of CP (p=0.06) than in the placebo group.

The limited data on the neurological outcome of 8 out of the 9 children lost to this follow-

up study (Table 1 in IV) confirmed these results. Furthermore, out of the 59 infants in the

original antenatal DEX study, 26 of the 37 (70%) children in the DEX group survived without

CP compared with 6 of the 21 (29%) in the placebo group (p=0.003).

79

DISCUSSION

1. Methodological aspects

Our study population consisted of VLBW children born very preterm, who participated in one

of the following two randomized placebo-controlled perinatal trials: study of neonatal DEX

given to severely ill ventilator-dependent infants at a high risk of BPD (Trial 1) (Kari et al

1993) or very preterm infants born to mothers who participated in the study of antenatal DEX

in imminent preterm birth (Trial 2) (Kari et al 1994). All infants were treated in the same

hospital during a 3-year period, with no major changes in ventilator treatment. Thus, the

infants in Trial 1 were a relatively homogenous group, with minimal variations in neonatal

treatment. The infants in Trial 2 were born after the anticipation of preterm birth, with the best

possible perinatal care available. The latter setting may underestimate the morbidities and the

long-term sequelae of very preterm birth.

Altogether 49 (79%) of the 62 recruited children participated in this follow-up. About one

third of the non-participants refused, one third were excluded because of severe neurological

problems, and the rest were not traced. The perinatal data of the non-participants did not differ

significantly from those of the study children. The neurological outcome data were extracted

from hospital records in the case of 61 (98%) children. Inclusion of the non-participants in the

neurologic assessment did not change the results (III and IV). Therefore, our study children

can be considered good representatives of the original trials.

Despite occasional severe neurological impairments, all could perform the IOS procedure

and all but one the spirometric measurements. The children with most severe lung disease or

small lung volumes could not perform the diffusing capacity test or the body

plethysmography. This may underestimate the impairment of lung function or conceal the

differences between the subgroups. However, occasional children with severe neurological

handicap, with or without lung disease, also either failed in performance or were

prospectively excluded. Even though the study population was small, the lung function tests

were carefully standardized with low variability (<5% in FVC and FEV1 and <10% in eNO).

All cardiac measurements were performed by the same pediatric cardiologist.

Only non-atopic full-term children were selected as a control group in the lung function

studies. The larger age range in the control group compared to the study groups made the

range of body heights, to which the lung function results were adjusted, very similar in all groups.

80

Compared to IOS, spirometry showed significant concordance, as did also body

plethysmography. Still, the different methods could not be compared in the children with the

most severe lung disease or the smallest lung volumes. One BPD child was misclassified into

the non-BPD group in Study I, but this had no significant effect on the results. (I)

The impairment of lung function was compared with one marker of inflammation, eNO.

To avoid the bronchodilator effect, eNO measurements were performed at another visit than

spirometry, which could influence the associations. (II)

The sample size in Study III was too small to warrant definite conclusions. The power of

this study alone is low, but it will be of value in future meta-analyses. The homogeneity of the

group (Trial 1), the successful lung function tests and the abundant neurological data are a

strength, but the frequent use of DEX after the study could be considered a weakness of the

study.

In Study IV, the size of the placebo group was small, due to the small number of infants

born very preterm and the high mortality (Trial 2). This emphasizes the better outcome in the

original DEX group, if perinatal death is included. This group was heterogenous in perinatal

respiratory morbidity; differences in treatment may individually affect the long-term outcome,

causing bias, or represent the effects of antenatal DEX. This follow-up is important in meta-

analyses of the long-term outcome, especially as new placebo-controlled randomized studies

will hardly follow, because of the well-established short-term benefits of antenatal DEX.

Definition of bronchopulmonary dysplasia

We used a simple criterion for BPD, i.e. supplemental oxygen at 36 gestational weeks.

Despite the variations in treatment strategies, discontinuation of oxygen is one clinical

descriptor of infant stability. In our study, the diagnosis of BPD is used more as a tool to

divide the preterm born children into those at a �high risk� and those at a �risk� of long-term

sequelae of prematurity than as a strictly defined disease. The infants in Trial 1 were born

before the introduction of antenatal glucocorticoid and surfactant therapy into common use,

and due to the entry criteria, represent VLBW children with severe lung disease and hence

with a high incidence of BPD (50%). The infants in Trial 2 were born at the beginning of the

era of antenatal glucocorticoid and surfactant therapy, and the incidence of BPD was 23%.

The differences in the definitions of BPD make comparisons of the incidences and long-term

sequelae in studies between different centres and periods extremely rough.

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2. Oscillometric method

In our study, IOS yielded lung function information concordant with conventional methods,

such as spirometry and whole-body plethysmography, and served well in studying

prematurely born children. The findings probably reflect the peripheral or more widespread

airway obstruction and are most valuable in combination with other lung function tests.

Using the oscillometric method, the mechanical properties of the lung have been described

previously in children with asthma (Solymar et al 1984, König et al 1984, Lebecque et al

1987, Bisgaard & Klug 1995, Klug & Bisgaard 1996), cystic fibrosis (Lebecque & Stănescu

1997, Hellinckx et al 1998) and chronic respiratory symptoms (Timonen et al 1997). In

asthma, airway obstruction is associated with increased Rrs, the values measured at the lowest

oscillatory frequencies (2-5 Hz) showing the best sensitivity (Clément et al 1983, Solymar et

al 1984, Bisgaard & Klug 1995). This pattern of abnormality has been similar regardless of

whether the FOT (Clément et al 1983, Solymar et al 1984) or the IOS method has been

applied (Bisgaard & Klug 1995, Klug & Bisgaard 1996). Despite the concordant results in

asthma, the concordance between FEV1 and Raw in cystic fibrosis has been reported to be poor

(Lebecque & Stănescu 1997). This may be due to the effect of dynamic compression of the

airways during forced expiratory manoeuvres, which is lacking when Raw is measured at tidal

breathing. In the present study, a significant but low concordance between Rrs5 and FEV1

was found. However, the concordance was better in Xrs, which delineated the severely

affected children (BPD) more accurately than Rrs. (I) This is in agreement with the previous

findings on asthmatic children, where Xrs has been shown to correlate well with FEV1 and

Raw during challenge tests (Solymar et al 1984, Bisgaard & Klug 1995, Klug & Bisgaard

1996, Bouaziz et al 1996). Compared to an increase in Rrs, a decrease in Xrs has been

reported to be a more sensitive (Buhr et al 1990, Bisgaard & Klug 1995, Klug & Bisgaard

1996) and specific (Bouaziz et al 1996) indicator of thoracic obstruction.

Theoretically, Xrs is mainly determined by the elastic and mass-inertial properties of the

respiratory system; at low frequencies the former predominate and Xrs varies in relation to

dynamic compliance (Solymar et al 1989). The dynamic compliance of the lung is impaired

in both peripheral airway obstruction and disorders causing stiffness of the lungs, such as

interstitial fibrosis, both changes likely to be present in BPD. As in our study, no significant

relationship was found between Xrs and DLCO, the decreased Xrs values are more likely to be

due to airway obstruction than interstitial fibrosis. Rrs5 was closely associated in Raw in body

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plethysmography. At the lower rate of resistance, IOS showed higher values than Raw, since

the method measures total respiratory resistance. At high values, Rrs5 underestimates

resistance (Cauberghs & Van de Woestijne 1989), as in our study.

The data on the range of normality in the bronchodilator response elicited with oscillatory

methods are still sparse (Delacourt et al 2000). In our study, the bronchodilator response in

Xrs5 correlated significantly with ∆FEV1, which was in line with the concordance found

between Xrs5 and FEV1.

To conclude, being easy to perform, IOS measurement alone may produce important

information on the lung function of preschool or school-aged children with neurological

handicap or other disabilities impairing cooperation. The bronchodilator test might be

valuable in examining children unable to produce forced expiration, and it should be further

evaluated. In combination with conventional lung function tests, IOS yields both concordant

and also additive information, as it assesses the quiet tidal breathing instead of forced

expiration. We chose to use the IOS method in our studies of long-term sequelae after the

administration of perinatal DEX (III and IV).

3. Exhaled nitric oxide

In eNO measurement, to avoid nasal NO contamination (Alving et al 1993), we used a

technique of single exhalation through a mouthpiece against a resistance, creating a back

pressure to close the soft palate (Kharitonov et al 1997). The present guidelines do not

recommend the use of a nose clip, but allow it in cases when the subject cannot avoid nasal

inspiration or nasal exhalation (ATS 1999). We used it systematically, as this technique of

breathing was familiar to the children from previous tests. Because neurological disabilities

and small lung volumes affect cooperation, the children were individually supported in the

performance. This technique was considered reliable in all study children, including the

youngest control child, who was 5.3 years old. The eNO concentration is highly flow-

dependent (Silkoff et al 1997). The expiratory target flows recommended at that time by the

ERS appeared unsuitable for us, as they require large lung volumes. Therefore, we used lower

target flows, which were still higher than those subsequently recommended by the ATS

(1999). Some variation of flows had to be accepted because of limited cooperation. Despite

this variation, we achieved recordings with acceptable individual variation in eNO, and the

flows in the different groups did not differ significantly. Even though relatively high flows

83

may mask some differences, they did not mask the differences between the atopic and non-

atopic groups.

In our prematurely born children, the atopic children presented with higher eNO values than

the non-atopic ones, in line with the previous findings (Frank et al 1998, Franklin et al 1999,

Mattes et al 1999, Piacentini et al 1999). Therefore, we excluded the atopic children and those

on regular glucocorticoid inhalations, known to suppress eNO, from the main study of eNO in

BPD. Living with a smoker (45%) had no detectable impact on eNO values either, in

agreement with previous findings (Franklin et al 1999). Despite the evident presence of

significant obstructive and restrictive lung function abnormalities in the study children,

especially those with BPD, we found no significant differences in eNO concentrations and

output between the non-atopic BPD, non-BPD and full-term control children. (II)

While prior studies on the inflammatory characteristics of BPD focused on the early neonatal

period (Merritt et al 1981, Speer 1999), postnatal follow-up data are scanty. Airway

inflammation has been assessed by eNO measurements in adult asthmatics (Alving et al 1993,

Kharitonov et al 1994, Persson et al 1994), in exacerbations of COPD (Maziak et al 1998)

and in asthmatic children (Lundberg et al 1996, Baraldi et al 1997, Nelson et al 1997),

particularly ones with atopy (Frank et al 1998). In asthmatic children, eNO concentration and

sputum eosinophil counts were found to be concordant, and eNO was proposed as an easy-to-

perform marker of eosinophilic airway inflammation (Mattes et al 1999, Piacentini et al

1999). In a recent study, eNO was superior to baseline lung function and bronchial

responsiveness in identifying preschool children with asthma (Malmberg et al 2003).

However, the role in non-atopic asthma was still unclear. In ventilated preterm infants, the

persistence of high bronchoalveolar lavage fluid nitrate associated with the development of

BPD (Vyas et al 1999), and high eNO levels shortly after birth were found in some infants

developing BPD (Aikio et al 2002). Exhaled NO concentrations decreased after DEX

treatment (Williams et al 2004). In face mask measurements at term, high eNO levels were

related to the presence of BPD (Leipälä et al 2004). No previous reports of eNO values in

children born very preterm with BPD exist.

As high eNO in atopic asthma is likely to represent airway inflammation, the lack of eNO

elevation in the school children with BPD in our study is evidence against airway

inflammation. However, high inducible NOS activity in the airway inflammatory cells or high

eNO is not a constant finding in neonatal inflammatory lung disease (Aikio et al 2000, Aikio

84

et al 2003). Non-atopic children born very prematurely showed no evidence of increased

concentrations of eNO. Nor did the eNO levels correlate with the severity of lung disease, as

determined by conventional lung function studies. As our study children with most severe

lung disease were all non-atopic, the eNO measurement did not serve in assessing the degree

of lung disease in a stable phase in these children.

To conclude, in non-atopic children born very prematurely, exhaled NO values did not

correlate with impaired lung function, indicating that eNO was not an inflammatory marker in

established BPD. However, because asthma is a common disease, eNO measurements may be

beneficial in the differential diagnosis of symptomatic children with chronic lung disease of

prematurity and atopic asthma.

4. Growth

In our study, the prematurely born children had significantly lower standardized height than

the controls, but the difference was seen only in boys. This confirms the findings of Hack et

al (2003), who showed that boys were shorter and lighter at the age of eight years, followed

by poorer catch-up growth until 20 years, and that this trend was especially obvious in those

who were SGA. Weight adjusted for height in our study did not differ between the groups.

Our BPD children weighed less adjusted for height than the non-BPD children, but in contrast

to earlier findings (Northway et al 1990b, Hakulinen et al 1996, Pelkonen et al 1997), they

were not shorter for age.

In previous studies, BPD children born during 1970-1980´s commonly showed growth

retardation in infancy (Northway 1979, Markestad & Fitzhardinge 1981), at school age

(Hakulinen et al 1996, Pelkonen et al 1997) or in adolescence (Northway et al 1990b), while

some others found no impairment (Bader et al 1987). Growth impairment was also seen in

VLBW children at preschool or school age or in adolescence, with or without catch-up

growth (Niklasson et al 2003, Hack et al 2003, Korhonen et al 2004c), and in ELBW children

(Doyle 2000a), but this findind has not been confirmed by others (Kilbride et al 2003).

Postnatal DEX may impair growth during treatment, but this effect is usually reversible

(Jones et al 1995, O´Shea et al 1999). Our study did not reveal any significant differences in

growth between the postnatal DEX and placebo groups, but the latter were shorter for age

than the controls. Among boys, the difference persisted. The cumulative dose of postnatal

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DEX during the first year of life did not correlate with the growth parameters, either (III and

IV). Thus, postnatal DEX was not shown to have a negative effect on somatic growth at

school age, in line with the previous findings at 2-3 years (Jones et al 1995, Romagnoli et al

2002). (III)

A single dose of antenatal DEX had no harmful effect on the growth parameters in our

study, as reported in the follow-ups of randomized glucocorticoid trials at 3-20 years of age

(MacArthur et al 1982, CGAST 1984, Smolders-de Haas et al 1990, Dessens et al 2000).

After antenatal DEX, body height and weight were slightly greater at three years (CGAST

1984), and after betamethasone, girls were taller and heavier at six years (MacArthur et al

1982) than after placebo treatment. In our study, the placebo group was shorter for age than

the controls. (IV)

Our DEX studies suggest that the diseases associated with prematurity itself rather than

glucocorticoid caused the growth retardation.

5. Atopy

One fourth of the study children were atopic, i.e. had positive skin prick tests. In Study II,

atopy was less common in the children with BPD (8%) than in those without BPD (33%). The

atopic children had a higher birth weight and needed less initial respiratory care than the non-

atopic study children. Of the children with severe initial respiratory disease in Study III, only

one was atopic, in contrast to 35% of the Study IV participants, even though the rates reported

for familial atopy did not differ. None of the skin prick-positive children had received

postnatal DEX, which differed significantly from the skin prick-negative children.

In a Finnish population-based study, positive skin prick tests were found in 77% of

schoolchildren with asthma or other symptoms from lower airways and in 40% of controls

with no chronic or recurrent respiratory symptoms (Remes & Korppi 1996).

Our results support the proposal that severe neonatal lung disease is not associated with an

increased incidence of atopy (Pelkonen et al 1997). Thus, we propose that the initial

inflammation and inflammatory response, the medications used or the strategies to support

preterm infants may modify the direction of the inflammatory sequence towards skin prick

negativity.

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6. Respiratory symptoms and medication

Half of our study children had glucocorticoid inhalations at 12 or 24 months of age, half

reported prolonged respiratory symptoms, and 10% reported regular use of inhalations. In the

Finnish study of schoolchildren, 7% had asthma and 8% other symptoms from lower airways

at the age of 7-12 years (Remes & Korppi 1996). The excess of morbidity and medication in

our study is in line with the previous studies of preschool or school-aged children born

preterm during the same period (Pelkonen et al 1997, Gross et al 1998, Korhonen et al

1999a). The abundant use of glucocorticoid inhalation by the BPD children at 12 months

might partly reflect, besides their high respiratory morbidity, the strategies to treat and follow

up BPD at that time; the use at 24 months probably reflects more accurately the prevailing

respiratory morbidity. The frequency of reported symptoms did not associate with BPD, as in

other studies (Gross et al 1998, Korhonen et al 1999a), but had a negative association with age

at the time of the study. The use of inhaled glucocorticoid was (Korhonen et al 1999a) and β2-

sympathomimetic agents (Gross et al 1998, Korhonen et al 1999a) tended to be more common

in BPD children. The use of inhaled medication during the preceding year associated with

impaired lung function, and no extra use due to the diagnosis of BPD was suspected at school

age. 27% of our study children had neither had prolonged respiratory symptoms nor used

inhalations until the time of study.

Reports of respiratory symptoms during follow-up after postnatal DEX trials are few: Jones et

al (1995) found no difference at three years, Yeh et al (1998) reported less upper respiratory

infections and rehospitalization at two years, and O´Shea et al (1999) found less frequent

rehospitalizations by the age of one year in the DEX than in the placebo group. Our finding of

no association between a �moderately early� course of postnatal DEX or the length of DEX

therapy during the first year of life and respiratory morbidity and medication are in line with

these previous findings in younger children. One fourth in each group had neither had

prolonged symptoms nor used inhalations by their enrolment in the study. (III)

The follow-up studies of randomized trials infrequently assess respiratory morbidity after

antenatal glucocorticoids: Smolders-de Haas et al (1990) reported more hospital admissions

because of infections or respiratory diseases during the first year after betamethasone

treatment. In our study, the use of antenatal DEX did not associate with respiratory morbidity

or medication at school age. (IV)

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In all, half of our study children with and also without BPD reported prolonged respiratory

symptoms during the preceding year, suggesting that preterm infants even without a �high

risk� for respiratory morbidity may need a careful follow-up.

7. Smoking in the family

Exposure to tobacco smoke was common in our study children, as shown in the children born

very prematurely, but did not differ between the BPD and non-BPD groups or between the

subgroups in the DEX studies.

In contrast to previous findings (Chan et al 1989a), maternal smoking or smoking during

pregnancy had no detectable association with respiratory symptoms. Maternal smoking or

smoking during pregnancy was not associated with impairment of basic lung function, as

assessed by spirometry and IOS. Nor did smoking during pregnancy correlate with the values

of diffusing capacity tests. In the antenatal DEX study, however, the preterm children with

smoking mothers had significantly lower DLCO values and a tendency towards lower KCO

values compared with the children of non-smoking mothers. Smoking in the family also

tended to associate with decreased diffusion capacity. (IV)

Our study suggests that daily exposure to tobacco smoke might present a major threat to the

already impaired lung function of children born very preterm. Therefore, it is of utmost

importance to prevent children born very prematurely from starting smoking themselves, as

already emphasized by Northway et al (1990b) and more recently by Doyle et al (2003).

8. Lung function

Prematurely born children with or without BPD

Our study children had significantly impaired basic lung function in spirometry, comparable

to the previous studies of children born nearly ten years earlier (Hakulinen et al 1996,

Pelkonen et al 1997). The response to bronchodilator was increased, in line with the previous

findings of increased bronchial responsiveness and lability in home monitoring of PEF

(Pelkonen et al 1997). The values of basic spirometry were strikingly more seriously impaired

in BPD than in non-BPD children, similarly to the other studies using the criterion of oxygen

at 36 weeks (Pelkonen et al 1997, Gross et al 1998, Jacob et al 1998, Kilbride et al 2003), but

see Hakulinen et al (1996). (I)

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combination of antenatal and postnatal glucocorticoids or their repetitive use may further

increase the hazards. On the other hand, low dosage may decrease side effects. The possible

beneficial and adverse long-term effects on lung growth remain to be studied further.

Lung function after postnatal dexamethasone treatment

Few follow-up studies of postnatal DEX trials are available (Jones et al 1995, Yeh et al 1998,

O´Shea et al 1999, Shinwell et al 2000), with no results of long-term lung function. In both

groups of the present study, mechanically ventilated infants showed signs of airway

obstruction and increased bronchodilator responsiveness at school age in spirometry and,

correspondingly, in IOS. The only significant difference between the randomized groups was

the lower FVC in the placebo group. Half of the children in the placebo group failed to

perform the diffusing capacity test, rendering their results invalid.

Thus, we did not find any negative effects of our �moderately early� postnatal DEX

treatment on pulmonary outcome. The common use of later DEX treatment in the placebo

group (50%) may bias the results. (III)

Lung function after antenatal glucocorticoids

Follow-up studies of lung function after randomized antenatal trials continued until school

age are scarce: no negative effects of glucocorticoid on lung function were found at 10-12

years after betamethasone (Smolders-de Haas et al 1990) or at six years after DEX treatment

(Wiebicke et al 1988). In the latter case, however, only 12% of the original participants were

studied. In a cohort study, no significant differences were seen in lung function between the

betamethasone and control groups at the age of 14 years (Doyle et al 2000b).

In the present study, both groups of children born very prematurely showed signs of

airway obstruction and increased bronchodilator responsiveness in flow-volume spirometry

compared with the controls. Increased resistance and lower reactance in IOS in VLBW

children indicated airway obstruction. The study groups had lower diffusing capacity than the

controls. The only significant difference in lung function tests between our DEX and placebo

groups was the higher bronchodilator responsiveness (∆FEV1) in the placebo group. Although

the lower mean reactance, which mainly reflects respiratory dynamic compliance, and KCO

would seem to suggest more pronounced parenchymal involvement in the placebo group than

in the DEX group, the size of the placebo group was too small to warrant definite conclusions.

In line with previous studies, our data imply that antenatal DEX treatment has no negative

effects on pulmonary outcome. (IV)

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9. Cardiovascular findings

In our study, only one girl with severe BPD had cardiac hypertrophy and increased pulmonary

arterial pressure still at school age (III and IV). This confirms the previous findings of no

cardiac hypertrophy (Farstad et al 1995, Korhonen et al 2004b) or significant pulmonary

hypertension at 2-7 years of age (Korhonen et al 2004b) in children born in the late 1980´s or

early 1990´s, contrary to the frequent finding of right ventricular hypertrophy and

cardiomegaly with cor pulmonale that accompanied the severe �old� BPD (Northway et al

1967, Markestad & Fitzhardinge 1981, Smyth et al 1981, Northway 1990a).

Abnormal lung vascular growth may be important in disturbing alveolar septation and the

formation of lung alveoli (Burri & Hislop 1998, Jobe & Bancalari 2001, Abman 2001, Parker

& Abman 2003). Glucocorticoids regulate lung and vascular growth and maturation at

different levels. Pharmacological doses may impair alveolarization by suppressing the

proliferation of capillaries within the septa (Tschanz et al 1995, Burri & Hislop 1998,

Tschanz et al 2003). However, this process may be reversible and depend on the length and

dosage of glucocorticoid therapy. As there are large interspecies differences in the duration of

vascular growth and the structural maturation of the lung, the interpretation of the

experimental results remains uncertain. (Burri & Hislop 1998, Tschanz et al 2003)

Pulmonary hypertension with raised pulmonary vascular resistance has been demonstrated

by cardiac catheterization in children with severe BPD until five years of age, associated with

poor prognosis, especially in those not responding to oxygen (Abman et al 1985, Berman et al

1986, Bush et al 1990). The neonatal decrease in pulmonary arterial pressure, assessed non-

invasively by echocardiography, is delayed in infants with RDS and may remain persistently

elevated in infants who subsequently develop BPD and serve as a sensitive marker of lung

injury (Gill & Weindling 1993, Subhedar et al 1998); it was still elevated at the age of one

year in BPD children (Subhedar & Shaw 2000). Pulmonary vasodilators, particularly inhaled

NO, may decrease the risk of BPD by decreasing intrapulmonary and extrapulmonary

shunting (Jobe & Bancalari 2001) and promoting lung growth and alveolarization by serving

as a downstream agonist of VEGF (Tang et al 2004).

Thus, the cardiac findings following BPD are modified in accordance with the changing

disease and treatments.

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Cardiological findings after postnatal dexamethasone treatment

In our study of survivors, no hypertrophic cardiomyopathy was found (III).

After postnatal DEX treatment, hypertension was frequent (CDTG 1991, Ng 1993)

contributing to the development of myocardial hypertrophy or cardiomyopathy (Werner et al

1992, Ng 1993, Evans 1994, Yeh et al 1997), and fatal cardiomyopathy was reported (Riede

et al 2001). However, in randomized trials (Kothadia et al 1999, Romagnoli et al 1999),

DEX-associated ventricular septal hypertrophy and left ventricular outflow obstruction were

usually reversible, although some infants presented with clinical symptoms (Bensky et al

1996, Zecca et al 2001).

The fall in PAP seen after DEX treatment did not correlate with the respiratory

improvement. This did not support the theory of a DEX effect mediated through a reduction

in pulmonary vascular resistance. (Evans 1994) We found no differences in the Doppler-

mode-derived variables between the DEX and placebo groups. Two patients in the DEX

group tended to have a short pulmonary artery acceleration time, but with no evidence of

elevated PAP, in contrast to a previous study showing short acceleration times (<100 ms) to

be associated with increased pulmonary vascular resistance (Farstad et al 1995). In the DEX

group, three patients had a tricuspid regurgitation gradient of 30-40 mmHg, with no clinical or

X-ray findings of pulmonary hypertension. Clinical and echocardiographic manifestations of

cor pulmonale were evident in one placebo group girl with BPD. Our DEX and placebo

groups did not differ in PAP measured non-invasively. Mild tricuspid regurgitation was found

in three asymptomatic patients. One girl in the placebo group had ASD secundum with a

remarkable left-to-right shunt of 2.0. She had no BPD, but chest X-ray revealed an enlarged

heart and minor congestion in the lungs without evidence of chronic pulmonary changes. The

ASD was successfully closed with a device at the age of nine years. (III) However, in early

infancy, ASD may be a risk factor for BPD (Motz et al 2000).

Cardiological findings after antenatal glucocorticoids

In the present study, a trend towards less PDA after DEX treatment (33%) than after placebo

(100%) was seen, as suggested previously (Eronen et al 1993), with no significance in

difference (IV).

In randomized trials comparing fetal effects, both betamethasone and DEX induced

significant but reversible changes in heart rate acceleration and variability (Mulder et al 1997,

Rotmensch et al 1999, Subtil et al 2003). No constant effect has been confirmed on PDA

(Crowley 1995, Wright et al 1995). Antenatal DEX did not appear to affect cardiac

93

dimensions and blood flow velocity in neonatal preterm infants (Skelton et al 1998).

In a follow-up of randomized trials, more abnormalities, mostly heart murmur, were

reported at three years in the placebo group (CGAST 1984). Systolic blood pressure was

significantly lower in the betamethasone group, but no difference was seen at 20 years

(Dessens et al 2000). No echocardiographic studies exist on the long-term effects of antenatal

glucocorticoid on heart function or growth. In the present study, the cardiac outcome did not

differ significantly at school age, regardless of whether antenatal DEX or placebo was

administered. None of the infants required cardiovascular treatment. (IV)

To conclude, antenatal or postnatal DEX treatment was not associated with abnormal cardiac

function or growth. Nor was BPD frequently accompanied by cardiac findings. However,

cardiac evaluation is important in symptomatic children after DEX treatment, as some

individuals may be liable to serious side-effects, and in preterm born children with severe

lung disease, as the lung disease may lead to cardiac sequelae or mask the cardiac symptoms.

10. Neurosensory development

Before the era of antenatal glucocorticoid and surfactant in the 1970-1980´s, the live birth

prevalence of CP, especially spastic diplegia, increased in a population-based Swedish

follow-up, reflecting the improved survival of VLBW infants (Hagberg et al 1989). Later, no

further increase was seen, but there was a slight decrease in preterm CP. In 1991-1994, the

prevalence was 82 per 1000 for infants of 1000-1499 g birth weight (Hagberg et al 2001).

IVH and PVL were powerful predictors of CP (Hagberg et al 1996, Salokorpi et al 1999). The

neurosensory development of ELBW infants was recently reviewed by Tommiska (2003).

Previously, VLBW infants born in the 1970´s, especially those with BPD, were shown to

have more neurological abnormality than term controls at the age of 10-12 years (Vohr et al

1991).

Neurosensory development after glucocorticoid treatment

Glucocorticoids impair the growth and development of the immature brain, but they may

either enhance or reduce brain injury after hypoxia, depending on dosage or timing.

(Whitelaw & Thoresen 2000) After widespread use of postnatal DEX and initial reports of

adverse neurodevelopment in follow-up, concern also arose about the possible long-term

adverse effects of antenatal glucocorticoid, especially when used repetitively (Ng 1993).

94

Antenatal glucocorticoid treatment has been considered a �physiological� glucocorticoid

stress for the fetus, comparable to other maternal environmental stress factors. During a

sensitive time �window�, it may program the cardiopulmonary system, brain and other organs

for postnatal life. The effect may be modified by the immediate postnatal environment.

Apparently, similar events early in life may produce different responses, depending on their

intensity and timing. (Matthews 2000, Seckl 2001) Thus, by improving neonatal adaptation,

antenatal glucocorticoid treatment may improve the outcome.

Neurosensory development after postnatal dexamethasone treatment

After �delayed� DEX treatment, new periventricular abnormalities on brain ultrasound scans

were reported (Noble-Jamieson et al 1989). The following studies of �moderately early� and

�early� treatment did not reveal any significant effects on the incidence of severe IVH or

severe ROP (Halliday & Ehrenkranz 2000ab), but did reveal slower head growth during DEX

treatment in 2-week and 4-week groups (Papile et al 1998). One �early� trial was terminated

owing to the adverse outcome, particularly PVL, in the DEX group (Soll et al 1999, Vermont

Oxford Study 2001), but one study found more IVH of any grade in the placebo group

(Garland et al 1999). Head circumference was smaller at 36 gestational weeks in the DEX

group in a recent study of �early� low-dose treatment (Stark et al 2001).

In our study of �moderately early� DEX treatment, no differences emerged between the

groups in abnormal ultrasound findings; one girl in the DEX had IVH grade 3-4 and one boy

in the placebo group IVH grade 2-3, both diagnosed before study entry. In each group, one

child received cryotherapy for ROP. (III)

Few follow-up results of DEX trials are available. At first, the small trial of a 42-day

�moderately early� course showed improved neurodevelopmental outcome at 15 months of

age (Cummings et al 1989). Thereafter, in the 209 VLBW children of the CDTG trial (1991)

born before the era of antenatal glucocorticoid and surfactant, no difference was evident at

three years after �delayed� treatment in CP (DEX vs placebo, 20% vs 17%), severe disability,

severe or moderate disability, visual or hearing loss or expected need for special school or

extra help at school (Jones et al 1995). Later, a 42-day �delayed� course in surfactant-treated

infants (Kothadia et al 1999) increased the risk of CP (25% vs 7%) (O´Shea et al 1999).

Similarly, in two �early� trials, a 4-week course (Yeh et al 1997) increased neurological

abnormality at two years (40% vs 17%) (Yeh et al 1998), and a 3-day course in surfactant-

treated infants (Shinwell et al 1996) increased the incidence of CP (49% vs 15%) and

95

developmental delay at 2-6 years of age (55% vs 29%) (Shinwell et al 2000). In the latter

trial, the DEX-treated infants tended to have more PVL and less IVH than the placebo-treated

ones, with no significant difference; all infants with PVL, 38% of those with IVH grade 1-2

and 53% of those with IVH grade 3-4 developed CP. In these �early� studies, the children

were more mature at birth than ours. In a recent follow-up of an early trial, the 4-week course

associated with smaller head circumference, poorer motor skills, lower IQ and more

significant disabilities at eight years of age (Yeh et al 2004).

Five (31%) of our preterm study children had CP (DEX 38%, placebo 25%) and 13%

were severely and 44% severely or moderately disabled (DEX 38%, placebo 50%), one with

visual loss. All the children were able to walk without support, and 88% attended normal

school. All could perform at least some lung function tests. The hospital record data of the

four children lost from this follow-up did not change the results: no CP was found, all could

walk without support, all attended normal school, and one in each group had learning

difficulties. Our findings of CP and disability are in line with those of Jones et al (1995).

Thus, severe or moderate disability was common, as were minor neurological impairment and

learning disabilities. We found no significant association between postnatal DEX treatment

and neurological outcome at school age.

In the recent trials, DEX was used less often in the placebo group than earlier (Yeh et al

11%, O´Shea et al 0%, Shinwell et al 33%, our study 50%, Jones et al 40%), and the use of

antenatal glucocorticoid was more common (Yeh et al 32%, O´Shea et al 32%, Shinwell et al

26%, our study 25%, Jones et al 0%). In a review by Barrington (2001), the studies with less

than 30% contamination with open-label DEX showed a greater steroid effect and a high risk

of CP associated with postnatal DEX. As half of our placebo children received open-label

DEX, the long-term safety of DEX cannot be suggested based on the similar outcomes

between the DEX and placebo groups.

After the documented effects of DEX on developing brain, based on �early� and some

�delayed� trials, the American Academy of Pediatrics and the Canadian Paediatric Society

(2002) published guidelines for postnatal glucocorticoid treatment: the use was to be limited

to exceptional clinical circumstances. However, glucocorticoids continue to be used

frequently in the USA and Europe (Soll 2003, Truffert et al 2003), especially in ELBW

infants (Barrington 2001). The excess of adverse neurological outcomes after �early� DEX

treatment may be explained by the instability of the infant in the first few days after birth,

when its tolerance of adverse events may be low, and the risk of cerebral ischemia may be

96

increased. According to currently available evidence, neurodevelopmental risks supervene the

benefits to lung function. The cost-benefit ratio of DEX treatment may possibly shift from

negative to positive when treatment is started after the age of 7-10 days. (Halliday 2003) This

assumption is supported by the present results. This is constant dilemma in neonatological

practise. Antenatal glucocorticoids, especially repeated courses, may influence the effect of

DEX and complicate the conclusions of long-term sequelae. Further research is needed to

assess the dosage and duration of DEX or other glucocorticoid in the treatment of VLBW

infants predisposed to various antenatal insults, infections, mechanical ventilation and

postnatal interventions.

Neurosensory development after antenatal glucocorticoids

Antenatal glucocorticoids prevent IVH (Crowley 1995, NIH 1995, Wright et al 1995).

However, in a retrospective study of 883 preterm infants, antenatal DEX, but not

betamethasone, predisposed infants to cystic PVL (Baud et al 1999), while others found no

increase in abnormal brain ultrasound findings, including PVL, after DEX in ELBW infants

(LeFlore et al 2002). Contrary to this, our study showed abnormal brain ultrasound findings to

be less common in the DEX group than in the placebo group. In a prospective cohort study,

repeated doses of glucocorticoids associated with a decrease in head circumference at birth,

representing a reduction in cranial volume of nearly 11% (French et al 1999). Recently,

betamethasone was preferred to DEX, and no more than two courses were recommended

(Baud 2004).

Few long-term follow-up results of randomized antenatal trials continued until school age or

adulthood are available. In a Dutch study of 81 adults at 20 years of age, no significant

differences were found in CP or cognitive function between the betamethasone and placebo

groups (Dessens et al 2000); these adults had been more mature at birth than our children,

ranging from 27 to 40 gestational weeks. Previous follow-up studies after antenatal

betamethasone (MacArthur et al 1982) and DEX trials (CGAST 1984) revealed no negative

neurological outcomes at six years (MacArthur et al 1982) or three years (CGAST 1984) of

age: 82% and 55% of the survivors were studied, respectively. A cohort study showed

teenagers exposed to antenatal betamethasone to have better cognitive functioning at 14 years

compared with VLBW controls (Doyle et al 2000b).

In our study, seven (20%) children had CP (DEX 11%, placebo 50%), 6% needed a

wheelchair or a walking aid, 9% had visual loss, and 6% needed a hearing aid. 17% were

97

severely and 34% severely or moderately disabled (DEX 22%, placebo 75%). Minor

neurological impairment and learning problems were common in both groups. 89% of the

children in the DEX group and 38% in the placebo group attended normal school; speech

therapy, physiotherapy or extra help at school was frequently needed. In the DEX group, the

normal school attendance rate was higher, and there were less severe to moderate disability

and a tendency to less CP than in the placebo group, in line with the previous findings of no

negative outcome at school age after antenatal betamethasone (MacArthur et al 1982, Dessens

et al 2000). In contrast to previous studies, we presented the data of non-participants, i.e. 98%

of the survivors of the original trial, with no change in the results. (IV)

Thus, in our study, antenatal DEX associated with better neurological outcome at school age.

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CONCLUSIONS

1. Obstructive and restrictive pulmonary dysfunction, bronchial hyperreactivity

and decreased gas exchange were evident in children born very prematurely,

especially in those with BPD. The impulse oscillometric method yielded lung

function information concordant with conventional methods. Being easier to

perform than spirometry, IOS served well in studying school-aged children, even

ones with neurological handicap or severe lung disease, and it may be useful in

preschool children as well.

2. Children with BPD rarely suffered from atopy. In non-atopic children born

very prematurely, exhaled nitric oxide values did not correlate with impaired lung

function, indicating that eNO was not an inflammatory marker in established BPD.

However, since atopy associated with increased eNO levels, we suggest that eNO

measurements may be valuable when studying atopic asthma even in preterm

children.

3. Neonatal dexamethasone therapy beginning ten days after birth in severely ill

ventilator-dependent infants associated with no detectable adverse effect on

cardiopulmonary function, somatic growth or neurosensory development at school

age. Severe or moderate disability was common. As half of the placebo children

received DEX after the trial, long-term safety cannot be suggested. More research is

needed for future conclusions.

4. Antenatal DEX treatment was not associated with any detectable adverse effect

on cardiopulmonary function or somatic growth. Besides initially helping more

infants to survive, it also caused a tendency for the children to have an improved

neurological outcome, as they had a higher rate of normal school attendance and

less severe or moderate disability. Survival without CP increased. In future studies

of different treatment strategies, the golden standard will be antenatal

betamethasone rather than placebo.

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5. Children born very prematurely have an increased risk of abnormal lung

function and frequent respiratory symptoms at school age. Even though those with

the most severe initial lung disease were non-atopic, some children may

additionally develop atopic asthma. With immature survivors and differences in the

etiology of preterm birth, the clinical course of chronic lung disease may be

modified. We propose that lung function studies are important in the follow-up of

children born very preterm or with VLBW, with or without BPD. Asthma

medication should be tried if the clinical picture is reminiscent of asthma. Early

exposure to tobacco smoke associated with impaired gas exchange. These and other

very high-risk children should be strongly advised against smoking.

6. Antenatal or postnatal DEX treatment was not associated with abnormal

cardiac function or growth. We propose that cardiac evaluation should be included

in the follow-up of preterm children with severe lung disease, as the lung disease

may lead to cardiac sequelae or mask the symptoms of a cardiac defect.

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ACKNOWLEDGEMENTS

This follow-up study was carried out at the Hospital for Children and Adolescents and the Skin and Allergy Hospital, in collaboration with the Division of Clinical Physiology and the Division of Pediatric Cardiology, during the years 1998-1999. I express my sincere gratitude to Professor Jaakko Perheentupa, former Head of the Hospital for Children and Adolescents, and his successors Professor Christer Holmberg, Professor Mikael Knip, Professor Martti Siimes and Docent Veli Ylitalo, Administrative Head of the Hospital for Children and Adolescents, and Docent Sture Andersson, Head of the Neonatal Unit, for creating and maintaining the excellent research facilities of the hospital. I am indebted to Docent Kaisu Juntunen-Backman, former Head of the Pediatric Department, and to Professor Tari Haahtela, Head of the Division of Allergy, for promoting my work and providing me with excellent research facilities.

I warmly thank my first teachers in neonatology: Professor Kari Raivio, Professor of Perinatology, aroused in me an interest in poorly breathing infants during my medical studies already with his unforgettable enthusiasm, curiosity and optimism. Docent Anna-Liisa Järvenpää taught me systematically the basic facts of neonatology in the Neonatal Intensive Care Unit and later in the Helsinki Maternity Hospital. I owe my deepest thanks to her and Docent Leena Lope, in memoriam, former Chief of the Neonatal Ward at Jorvi Hospital, for teaching and sharing the challenging work of neonatal care and follow-up after intensive care, as well as to all the skilful colleagues and staff in the neonatal units.

I am most grateful to my supervisors, Docent Markku Turpeinen at the Skin and Allergy Hospital and Professor Mikko Hallman, Head of the Department of Pediatrics, University of Oulu, for their encouragement and guidance throughout this process. I warmly thank Docent Turpeinen for the wide range of conversations on asthma and premature lung during these years. I admire his enthusiasm, open mind and creativeness in scientific thinking as well as his wide experience in pediatric pulmonology. I am very grateful to Professor Hallman whose constructive advice, thorough answers and never-failing support and interest were of essential importance, especially at the state of finishing the final manuscript. I had the honour to benefit from his profound knowledge in perinatal pulmonology already when adjusting ventilators during night shifts at the time when our study children were treated in the Neonatal Intensive Care Unit.

Docent Kirsti Heinonen and Kirsi Timonen, M.D., Ph.D., are gratefully acknowledged for their thorough interest and careful review of the final manuscript as well as for the enjoyable conversations and valuable comments.

I warmly thank Anneli Kari, M.D., for her consistent collaboration and support during these years. Her initial studies during 1989-1991 were the basis of this follow-up study. Her help in reaching the families and her interest in this study and valuable advise in writing the neonatological parts of the original manuscripts were essential.

I deeply thank Docent Pekka Malmberg, Chief of the Lung Function Laboratory of the Division of Allergy, for placing his expert knowledge at my disposal and guiding me into the world of pediatric lung function measurements. He supervised all the lung function tests at the Skin and Allergy Hospital and patiently helped in writing the original papers. His encouragement at the time of my first oral presentation in Florence is to be underlined.

I owe my thanks to Anna Pelkonen, M.D., for arousing my interest in this study. Her previous studies of premature infants have shown the importance of pulmonary follow-up of these children and facilitated further research. She also collaborated in studying the control children.

102

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