Prenatal Cotinine Levels and ADHDAmong OffspringAndre Sourander, MD, PhD,a,b,c Minna Sucksdorff, MD,a,d Roshan Chudal, MBBS, MPH, PhD,a Heljä-Marja Surcel, PhD,e,f

Susanna Hinkka-Yli-Salomäki, PhLic,a David Gyllenberg, MD, PhD,a,g,h Keely Cheslack-Postava, PhD,c Alan S. Brown, MD, MPHc,i

abstractOBJECTIVES: An association between maternal smoking during pregnancy and offspringattention-deficit/hyperactivity disorder (ADHD) has been shown across several studies basedon self-reports. No previous studies have investigated the association of nicotine exposuremeasured by cotinine levels during pregnancy and offspring ADHD.

METHODS: In this population-based study, 1079 patients born between 1998 and 1999 anddiagnosed with ADHD according to the International Classification of Diseases and 1079matched controls were identified from Finnish nationwide registers. Maternal cotinine levelswere measured by using quantitative immunoassays from maternal serum specimenscollected during the first and second trimesters of pregnancy and archived in the nationalbiobank.

RESULTS: There was a significant association between increasing log-transformed maternalcotinine levels and offspring ADHD. The odds ratio was 1.09 (95% confidence interval [CI]1.06–1.12) when adjusting for maternal socioeconomic status, maternal age, maternalpsychopathology, paternal age, paternal psychopathology, and child’s birth weight forgestational age. In the categorical analyses with cotinine levels in 3 groups, heavy nicotineexposure (cotinine level .50 ng/mL) was associated with offspring ADHD, with an odds ratioof 2.21 (95% CI 1.63–2.99) in the adjusted analyses. Analyses by deciles of cotinine levelsrevealed that the adjusted odds for offspring ADHD in the highest decile was 3.34 (95% CI2.02–5.52).

CONCLUSIONS: The study reveals an association with and a dose-response relationship betweennicotine exposure during pregnancy and offspring ADHD. Future studies incorporatingmaternal smoking and environmental, genetic, and epigenetic factors are warranted.

WHAT’S KNOWN ON THIS SUBJECT: Exposure to maternalsmoking is associated with various adverse perinataloutcomes. Association between maternal smoking andoffspring attention-deficit/hyperactivity disorder has beenshown across studies. However, the causality of theassociation has been questioned to be mostly due tofamilial confounding.

WHAT THIS STUDY ADDS: In this first nationwide study,objectively measured nicotine exposure through maternalcotinine levels allows us to overcome underreporting ofsmoking during pregnancy. We report a strongassociation as well as a dose-dependent relationshipbetween prenatal nicotine exposure and offspringattention-deficit/hyperactivity disorder.

To cite: Sourander A, Sucksdorff M, Chudal R, et al.Prenatal Cotinine Levels and ADHD Among Offspring.Pediatrics. 2019;143(3):e20183144

aDepartment of Child Psychiatry, University of Turku, Turku, Finland; Departments of bChild Psychiatry anddPediatric and Adolescent Medicine, Turku University Hospital, Turku, Finland; cNew York State PsychiatricInstitute, Department of Psychiatry, Columbia University Medical Center, New York, New York; eFaculty of Medicine,Medical Research Center, University of Oulu, Oulu, Finland; fBiobank Borealis of Northern Finland, Oulu UniversityHospital, Oulu, Finland; gNational Institutes of Health and Welfare, Helsinki, Finland; hDepartment of AdolescentPsychiatry, Helsinki University Central Hospital, Helsinki, Finland; and iDepartment of Epidemiology, MailmanSchool of Public Health, Columbia University, New York, New York

Dr Sourander conceptualized the study, participated in the study design, drafted the initialmanuscript, and contributed to the interpretation of the data and the critical review and revision ofthe manuscript; Dr Sucksdorff participated in the study design, conducted the literature search,drafted portions of the initial manuscript, and contributed to the interpretation of the data and thecritical review and revision of the manuscript; Dr Chudal participated in the study design, draftedportions of the initial manuscript, and contributed to the interpretation of the data and the criticalreview and revision of the manuscript; Drs Surcell, Gyllenberg, Cheslack-Postava, and Brownconceptualized the study, participated in the study design, and contributed to the interpretation ofthe data and the critical review and revision of the manuscript; Ms Hinkka-Yli-Salomäki designedthe study, conducted the analyses, and contributed to the interpretation of the data and the criticalreview and revision of the manuscript; and all authors approved the final manuscript as submitted.

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Despite its proven negative effects onfetal development, smoking duringpregnancy remains a significantpublic health issue. Approximately7.2% of women who gave birth in theUnited States smoked cigarettesduring pregnancy in 2016.1 Theprevalence rate was similar inFinland, with ∼7% of all pregnantwomen continuing to smokethroughout their pregnancy.2

Exposure to maternal smoking hasbeen associated with various adverseoutcomes, such as obstetriccomplications, low birth weight,preterm birth, sudden infant deathsyndrome, and increased infections inchildhood.3 An association betweenmaternal smoking and offspringattention-deficit/hyperactivitydisorder (ADHD) has been shownacross numerous studies fromdifferent populations.3–7 However, thecausality of the association has beenquestioned to be mostly due tofamilial confounding.3,8–13 Moreover,to date, the information on maternalsmoking is typically based solely onmaternal self-report.

In previous studies, maternal self-report of smoking has been shown tounderestimate true smoking by 8%to 28%.14–16 Disclosure of smokingis lower among pregnant smokersthan women who smoke in general.Cotinine is the most appropriatebiomarker indicating nicotineexposure because of the short half-lifeof nicotine, which is primarilymetabolized to cotinine in the liver.17

Although previous studies are basedon self-reports of active smoking,cotinine measurements enablequantifying the amount of nicotineexposure and detecting nicotineexposure from other sources, such asnicotine replacement therapy orpassive smoking. However, authors ofno previous studies have investigatedthe association of prenatal cotininelevels and the risk of offspring ADHD.In 2 previous studies that haveexamined the association betweenmaternal cotinine levels in pregnancy

and childhood behavioral outcomeson the basis of parentalquestionnaires, the authors reportedno association.18,19

This is the first study to investigatethe association between maternalcotinine levels during pregnancyand ADHD diagnosis in offspring. Inthis population-based case-controlstudy, the use of objectivelymeasured nicotine exposure allowsus to overcome underreporting ofsmoking. On the basis of findingsfrom previous studies revealing anassociation between self-reportedmaternal smoking duringpregnancy and ADHD diagnosis inoffspring, we hypothesized thatthere is an association betweenmaternal cotinine levels andoffspring ADHD. In addition, ourstudy allows us to examine possibledose-response effects betweenmaternal cotinine levels andoffspring ADHD.

METHODS

The current study is based on theFinnish prenatal study of ADHD(Fig 1), a nested case-control studyderived from all singleton live birthsin Finland between January 1, 1998,and December 31, 1999, andfollowed-up for any ADHD diagnosisin the Finnish Hospital DischargeRegister (FHDR) by December31, 2011.

Finnish Maternity Cohort

All offspring in the Finnish prenatalstudy of ADHD were derived from theFinnish Maternity Cohort (FMC). TheFMC consists of 2 million serumsamples collected during the first andearly second trimester of pregnancy(fifth to 95th percentile: months 2–4of pregnancy) from .950000 womensince the beginning of 1983. Afterinformed consent, blood sampleswere collected at Finnish maternityclinics for the purpose of screening

FIGURE 1Flowchart of the study design. PRC, Population Register Center.

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for congenital infections (HIV,hepatitis B, and syphilis). Onematernal serum sample was obtainedfrom each pregnancy. The mediangestational age of serum collection forsubjects in this study was 10 weeks(interquartile range: 8–12 weeks).After the screening, ∼1 to 3 mL ofserum from each pregnancy arestored at 225°C in a protectedbiorepository at Biobank Borealis inOulu, Finland, and are available forscientific research.20 Linkage betweenFMC data and the Finnish healthregistries and other data sources ispossible by using the Finnishpersonal identity code (PIC), whichhas been assigned to all residents ofFinland since 1971.21

Nationwide Registers

The Population Information Systemwas established by the FinnishPopulation Register Center in 1969and subsequently computerized in1971. It is a computerized nationalregister containing basic informationabout Finnish citizens and foreignpermanent residents. The personaldata in the system include name,PIC, address, citizenship and nativelanguage, family relations, and date ofbirth and death (if applicable). TheFinnish Medical Birth Register(FMBR), established in 1987, includescomprehensive and standardized dataon the perinatal period for all livebirths and stillbirths of fetuses witha birth weight of at least 500 g ora gestational age of at least 22 weeks.

Patients and Controls

The patients with ADHD wereidentified by linking the informationfrom the FHDR with the FMC usingthe PIC. The FHDR contains allinpatient diagnoses since 1967 andoutpatient diagnoses from specializedservices since 1998. The FHDRcontains the patient’s PIC, hospitalidentification, and primary andsecondary diagnoses. The diagnosticclassification is based on theInternational Classification ofDiseases; the International

Classification of Diseases, 10thRevision (ICD-10) has been used since1996.22 From 1987 to 1995 thediagnoses were coded according tothe International Classification ofDiseases, Ninth Revision (ICD-9),23

and from 1969 to 1986 according tothe International Classification ofDiseases, Eighth Revision (ICD-8).24

Finland has a system of regularassessment of children’s physical andpsychological development by well-trained health professionals. Almostall children attend free health check-ups at least 15 times during their first6 years of life followed by annualcheck-ups at school. ADHD is typicallydiagnosed on the basis of theassessment of a specialist inpsychiatry or neurology in publicoutpatient services free of charge.A previous study, in which theauthors evaluated the validity of theADHD diagnosis in the FHDR,revealed that 88% of subjectsexamined met the Diagnostic andStatistical Manual of Mental Disorders,Fourth Edition diagnostic criteria forADHD.25

In the current study, the patients withADHD included singletons born inFinland between 1998 and 1999 andregistered in the FHDR with the ICD-10 code for hyperkinetic disorders(F90.0) until 2011. Controls weredefined as singleton offspring born inFinland without a diagnosis of ADHD,conduct disorder, or severe orprofound intellectual disability in theFHDR. Each participant with ADHDwas matched with 1 control on dateof birth (630 days), sex, and place ofbirth. The controls had to be alive andliving in Finland at the date of theADHD diagnosis of the matchedpatient. Patients and controls wereselected from these years because ofthe lack of earlier availability of datafrom the FMBR (established in 1987)and to ensure greater uniformity ofthe serum specimens with regard tothe time of sampling and storage. Inthe current study, of the 1320 totalpatients and controls identified,

sufficient serum was available in theFMC for 1079 patients and matchedcontrols (n = 1079). The study wasapproved by the ethical committeesof the Hospital District of SouthwestFinland and the National Institutes ofHealth and Welfare. Informed consentwas obtained before collection ofmaternal serum.

Cotinine Measurements

Cotinine measurements from the FMCsamples were conducted blind topatient/control status. Serumcotinine levels were measured byusing a commercially availablequantitative immunoassay kit(OraSure Technologies, Bethlehem,PA) (sensitivity = 96%–97%;specificity = 99%–100%). Intra-assayand interassay variation are 3.5% to6.2%, and 6.0% to 9.6%, respectively.The results are reported in nanogramper milliliter.

Covariates

The covariates included variables thathave been previously shown to beassociated with both maternalsmoking as well as offspring ADHD:number of previous births, maternalsocioeconomic status (SES), maternalpsychiatric history, maternal historyof ADHD diagnosis, maternal historyof substance use disorder, paternalpsychiatric history, paternal history ofADHD diagnosis, gestational age ofthe child, birth weight for gestationalage (BWGA; based on national sex-specific growth curves26), maternalage, paternal age, and gestationalweek of blood draw. Information onnumber of previous births, maternalSES, maternal age, and gestational agewas obtained from the FMBR.Maternal and paternal psychiatricdiagnoses were obtained from theFHDR. A parent was defined as havinga psychiatric history if he or she hadany psychiatric diagnoses registeredin the FHDR during his or herlifetime (ICD-10 codes F10–F99[corresponding to ICD-9 codes291–316 and ICD-8 codes 291–309]),excluding intellectual disability (ICD-

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10 codes F70–F79) or a diagnosis ofADHD (ICD-10 code F90.x or ICD-9code 314.x). For mothers, theassociation with disorders due toalcohol or substance abuse (ICD-10codes F10–F19; ICD-9 codes 291–292and 303–305; and ICD-8 codes 291,303, and 304) was tested separatelyfrom other psychiatric disorders.Paternal age was obtained from theFinnish Population Register Center,and gestational week of blood drawwas obtained from the FMC. Thecovariates were considered forinclusion in the models on the basisof associations with both cotinineexposure and ADHD at P # .1, inaccord with standard texts.27

Statistical Analysis

The analysis was based on a nestedcase-control design, in which thecontrols for each patient wereidentified from the population at riskand matched on selected factors (seePatients and Controls). Initially,cotinine was examined as a continuousmeasure. Because of the skeweddistribution of cotinine, the variablewas log-transformed before analysis.

To further facilitate datainterpretation, we examined maternal

cotinine as categorized into decilesand as a 3-class categorical variable:reference (,20 ng/mL); moderateexposure (20–50 ng/mL); and heavyexposure (.50 ng/mL). The decilesfor the patient and control groups inthe analyses were derived from thecut points of maternal cotinine levelsthat defined the deciles for thisbiomarker in the control group. Wehypothesized that a significantassociation would be observed formaternal cotinine classified in thehighest decile compared with thereference group, which was definedas the lowest decile. The cutoff pointsfor the 3-class categorical variablewere based on the manufacturer’srecommendation and have been usedin previous studies on the basis of theFMC serum bank.28,29 These levelsalso corresponded well with thefrequencies of self-reported maternalsmoking. Among mothers of patientswith ADHS, smoking was reported by56 of 702 (7.9%) mothers in thereference category of ,20 ng/mL, by62 of 80 (77%) mothers with cotininelevels of 20 to 50 ng/mL, and by 196of 214 (91.5%) mothers with cotininelevels .50 ng/mL (P , .001).Appropriate to the nested case-

control study design, point andinterval estimates of odds ratios(ORs) were obtained by fittingconditional logistic regression modelsfor matched pairs. Statisticalsignificance was based on P , .05. Allthe statistical analyses wereperformed with SAS software (SAS9.4; SAS Institute, Inc, Cary, NC).

RESULTS

The mean age of the patients at thetime of the ADHD diagnosis was7.3 years (SD: 1.9; range: 2–13.7years). As shown in SupplementalTable 7, there was no difference incovariates between 1079 patientsincluded in the analysis and 241patients not included because ofinsufficient serum, except for paternalage. The mean paternal age amongpatients not included was 1.2 years(SD: 6.9) older than that of patientsincluded (P = .02) in the analysis. Themean cotinine level was 27.4 ng/mL(SD: 54.8; range: 0.0–427.7 ng/mL)among patients and 11.3 ng/mL (SD:34.5; range: 0.0–320.0 ng/mL) amongcontrols. Supplemental Table 4reveals the distribution of serumcotinine levels and corresponding

TABLE 1 Association Between Maternal Serum Cotinine (Log-Transformed and 3-Class Categorical Variables) and Offspring ADHD

Patient Control Association With Maternal Serum Cotinine

OR 95% CI P

Maternal cotinine levels (ng/mL), median 27.4 11.3 — — —

Log-transformed analysisUnadjusted — — 1.14 1.11–1.17 ,.001Model 1a — — 1.09 1.06–1.13 ,.001Model 2b — — 1.10 1.06–1.13 ,.001

Categorical analysisUnadjustedReference, n (%)c 777 (72.0) 936 (86.8) — — —

Moderated 84 (7.8%) 54 (5.0%) 1.92 1.33–2.77 ,.001Heavy exposuree 218 (20.2%) 89 (8.2%) 2.95 2.25–3.88 ,.001

Model 1a

Moderate exposured — — 1.27 0.84–1.92 .25Heavy exposuree — — 2.21 1.64–2.99 ,.001

Model 2b

Moderate exposured — — 1.31 0.87–1.96 .20Heavy exposuree — — 2.27 1.69–3.07 ,.001

—, not applicable.a Model 1: adjusted for BWGA, maternal SES, maternal age, maternal psychopathology, maternal substance abuse, paternal age, and paternal psychopathology.b Model 2: adjusted for BWGA, maternal SES, maternal age, maternal psychopathology, maternal substance abuse, and paternal age.c Reference: ,20 ng/mL.d Moderate exposure: 20–50 ng/mL.e Heavy exposure: .50 ng/mL.

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self-reported smoking status in thesample. As shown in the table, theprevalence of self-reported smokingstatus increased with increasingserum cotinine levels.

As shown in Supplemental Table 5,maternal SES, psychopathology,substance abuse, ADHD, age, andnumber of previous births; paternalpsychopathology, ADHD, and age;and the child’s gestational age andBWGA were associated (P , .1)with offspring ADHD (seeSupplemental Table 5). As shown inSupplemental Table 6, maternal SES,psychopathology, substance use, andage; paternal psychopathology; andthe child’s BWGA were associatedwith maternal serum cotinine levels.Gestational week of blood draw wasnot associated with these outcomes(see Supplemental Table 6). MaternalSES, psychopathology, substance use,and age; paternal psychopathology;and the child’s BWGA were associatedwith both offspring ADHD andmaternal cotinine levels and thereforefulfilled criteria for confounding.Furthermore, adjustment was madefor paternal age to address anypotential selection bias because of itsdifference among included andmissing patients with ADHD.

The main findings of the currentstudy are shown in Table 1. Theadjusted analyses were performed byusing 2 models. Model 1 containedthe child’s BWGA, maternal SES,maternal age, maternalpsychopathology, maternal substanceabuse, paternal age, and paternalpsychopathology as confounders.Model 2 contained the sameconfounders as in model 1 except forpaternal psychopathology tospecifically assess the effect ofmaternal factors. There wasa significant association betweenincreasing log-transformed maternalcotinine levels and offspring ADHDboth in unadjusted analyses (OR:1.14; 95% confidence interval [CI]:1.11–1.17) as well as adjustedanalyses. In model 1, the OR was 1.09

(95% CI: 1.06–1.12), and in model 2,the OR was 1.10 (95% CI: 1.06–1.13).

In the categorical analyses, cotininelevels were categorized into 3 groups:heavy ( .50 ng/mL), moderate(20–50 ng/mL), and no or lownicotine exposure (,20 ng/mL). Asshown in Table 1, heavy exposurewas associated with offspring ADHD inthe unadjusted analyses (OR: 2.95; 95%CI: 2.25–3.88) as well as in the adjustedanalyses both in model 1 (OR: 2.21;95% CI: 1.63–2.99) and in model 2 (OR:2.27; 95% CI: 1.68–3.07). Moderatecotinine levels were associated withoffspring ADHD in the unadjustedanalyses (OR: 1.92; 95% CI: 1.33–2.77)but did not remain significant in theadjusted models (model 1: OR: 1.27[95% CI: 0.84–1.92]; model 2: OR: 1.31[95% CI: 0.87–1.96]).

The distribution of cotinine exposurein deciles by case-control status ispresented in Table 2. As shown inFig 2, the strongest association was inthe highest decile (90%–100%). Theodds for offspring ADHD in thehighest decile in the unadjustedanalyses was 4.90 (95% CI: 3.10–7.76),whereas in the analyses adjusting forBWGA, maternal SES, maternal age,maternal psychopathology, maternalsubstance abuse, paternal age, andpaternal psychopathology, it was 3.34(95% CI: 2.02–5.52). For the secondhighest decile (80%–89%), theassociation in the unadjusted analysisrevealed an OR of 2.71 (95% CI:1.70–4.31) and in the adjusted

analyses, an OR of 1.91 (95% CI:1.15–3.18; Table 3). The differencebetween the deciles (80%–89% vs90%–100%) was significant, revealingthat the risk of ADHD was higher witha higher level of nicotine exposure.

DISCUSSION

With this study, we provide evidencefor the association between nicotineexposure during pregnancy andincreased risk of ADHD in theoffspring because previouspopulation-based studies have beenbased on self-report of smokingduring pregnancy. The associationbetween fetal exposure to nicotine,quantified as cotinine duringgestation, and ADHD were similarwhen cotinine was classified asa continuous variable and when itwas classified in 3 categories on thebasis of cotinine levels. Mostimportantly, the findings reveala dose-dependent relationshipbetween fetal exposure to nicotineand risk of ADHD. The findingspersisted in all analyses afteradjusting for potential confounders,including maternal SES, maternal andpaternal psychopathology, maternaland paternal age, and BWGA. To thebest of our knowledge, this is the firstbiomarker-based study to reveala relationship between fetal nicotineexposure and later offspring ADHD.

The findings add to the evidence ofa complex association between

TABLE 2 Maternal Serum Cotinine Levels by Deciles in Patients With ADHD and Matched Controls

Maternal Cotinine by Deciles,a % Range, ng/mL Patients(n = 1079)

Controls(n = 1079)

n % n %

,10 ,0.008 67 6.2 107 9.910–19 0.009–0.011 73 6.8 108 10.020–29 0.012–0.014 66 6.1 107 9.930–39 0.015–0.019 89 8.2 109 10.140–49 0.020–0.024 96 8.9 112 10.450–59 0.025–0.033 99 9.2 104 9.660–69 0.034–0.052 88 8.2 108 10.070–79 0.053–0.165 118 10.9 108 10.080–89 0.166–41.08 139 12.9 108 10.0$90 $41.08 244 22.6 108 10.0

a The cut points for deciles are based on maternal cotinine levels in the control group.

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exposure to nicotine duringpregnancy and later ADHD. Aneurobiological mechanism viaintrauterine effects seems possiblebecause nicotine crosses the placentaduring pregnancy.30 Animal studieshave demonstrated an associationbetween in utero nicotine exposure

and increased locomotor activity aswell as changes in theneurotransmitter systems in thebrain.31,32 Human studies haverevealed structural and functionalchanges in the brain associated withprenatal smoking exposure.33 Themechanisms behind these effects are

thought to develop through nicotinemodulating the maturation of thedeveloping central nervous system. Inaddition, other components ofsmoking, such as carbon monoxide,are thought to lead to fetal hypoxiaand affect fetal brain development.34

The association between smokingduring pregnancy and offspringADHD may be explained by genetic orsocial factors.9,35 On the basis ofseveral sibling and family studies, theassociation between fetal exposure tosmoking and ADHD has beensuggested to be mostly explained byfamilial confounding.8–13 Thesestudies, as well as a novel studydesign on children born with assistedconception,36 suggest that inheritedeffects and unmeasured household-level confounding seem more likelycontributors behind the associationthan direct intrauterine effects ofsmoking. There are also contradictorysibling-study findings, includingresults from a Finnish study with.150000 sibling pairs, revealing thatif the mother stopped smokingbetween the pregnancies, the secondsibling did not have an increased riskof externalizing diagnoses, includingADHD. Correspondingly, if the motherstarted smoking between thepregnancies, a significantly higherrisk was observed in the secondsibling.37 A Dutch family study foundthat maternal compared withpaternal smoking during pregnancywas associated with a greater effect,with externalizing problems amongoffspring. The same study revealed

FIGURE 2ORs and 95% CIs for maternal serum cotinine (deciles) and offspring ADHD.

TABLE 3 Association Between Maternal Serum Cotinine Categorized Into Deciles and Offspring ADHD

Maternal Cotinine by Deciles, % OR 95% CI P ORa 95% CI P

,10 Reference — — — Reference —

10–19 1.15 0.71–1.85 .56 1.18 0.71–1.96 .5320–29 1.19 0.72–1.95 .50 1.07 0.63–1.83 .8130–39 1.58 0.97–2.58 .07 1.67 0.98–2.84 .0640–49 1.79 1.09–2.93 .02 1.60 0.94–2.72 .0950–59 2.09 1.27–3.44 .004 2.01 1.18–3.44 .0160–69 1.92 1.15–3.2 .013 1.71 0.98–2.98 .0670–79 2.78 1.66–4.68 ,.001 2.38 1.35–4.21 .00380–89 2.71 1.70–4.31 ,.001 1.91 1.15–3.18 .01$90 4.90 3.10–7.76 ,.001 3.34 2.02–5.53 ,.001

—, not applicable.a Adjusted for BWGA, maternal SES, maternal age, maternal psychopathology, maternal substance abuse, paternal age, and paternal psychopathology.

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that quitting smoking was associatedwith less externalizing problemsamong offspring.38 Furthermore, anAmerican sibling study revealed thatfamilial confounding explainedinattentive but not hyperactive and/or impulsive ADHD behaviors,suggesting that the association mayvary by phenotype.39

Despite several strengths, siblingcomparison studies should also beinterpreted with caution given theirlimitations. It has been suggested thatwithin-sibling estimates will be biasedtoward the null by measurement errorand that they may be either more orless biased than between-familyestimates depending on the extent towhich siblings share confoundersversus the exposure.40 In addition,mothers who vary in their smokinghabits during different pregnanciescan perhaps not be generalized to allsmoking populations.40

It is possible that the associationbetween nicotine exposure duringpregnancy and ADHD might partiallybe explained also by gene-environment interaction.Accumulating data provide evidencethat prenatal smoking may actthrough epigenetic changes viaaltered DNA methylation andmicroRNA expression.41,42 Theexposure to nicotine may increase therisk of ADHD particularly amongchildren with genetic vulnerability forADHD. It is also possible thatmaternal smoking during pregnancyis a proxy risk factor leading toADHD by independent mechanisms.Smoking during pregnancy isassociated with poorer parentingskills, which are associated with childbehavioral problems.43

The current study has severalstrengths, including being based ona large nationwide sample, assessingnicotine exposure with objectivebiological measurement andincluding several potentialconfounders. However, wheninterpreting the findings, several

limitations should be considered.A key question is whether smokingduring pregnancy is causallyassociated with ADHD or is a proxy ofanother risk factor (eg, familialconfounding). Even if most of theeffect is due to familial or geneticconfounding, the current studyreveals that the association betweensmoking and ADHD has a dose-response effect. The limitation ofobservational data is that we cannotexamine causal processes. We did nothave access to biomarkers of siblingpregnancies that would have shedmore light on a possible causal linkbetween nicotine exposure duringpregnancy and offspring ADHD.However, in the current study, wewere able to adjust for severalconfounders, including BWGA,maternal SES, age, psychopathology,and substance use disorder as well aspaternal psychopathology. Informationabout possible substance use duringpregnancy was restricted to register-based substance use diagnoses. In thecurrent study, 5.3% of mothers ofchildren with ADHD had a diagnosis ofsubstance use disorder, which wassimilar to the estimated prevalence inFinland of 6.4% for the exposure tomaternal alcohol and drugdependence.44 The number of parentsdiagnosed with ADHD in this samplewas low, which is a limitation of thestudy (Supplemental Tables 5 and 6).The underdiagnoses among parentscould be primarily because ADHD wasnot a widely used diagnosis in theparental generation. In addition, it isunlikely that ADHD among parentswas treated in inpatient care. Becausethe FHDR did not cover outpatientdiagnoses before 1998, the diagnosisof ADHD among parents is likelyunderestimated. Finally, the subjectswith ADHD included in this studywere only those who were referred tospecialized services and likelyrepresent the more severe cases ofADHD. However, authors of a previousstudy reported an 88% validity of theADHD diagnoses in the FHDR

examined against Diagnostic andStatistical Manual of Mental Disorders,Fourth Edition criteria for ADHD.7

CONCLUSIONS

According to the World HealthOrganization, smoking is considered 1of the main public health concernsworldwide.45 The current studyreveals a strong association betweenprenatal nicotine exposure anda dose-dependent relationship withoffspring ADHD. This study adds 2important aspects; first, the use ofcotinine as a documented measure ofnicotine exposure during pregnancyand second, the finding of a dose-response effect in the association.Given the high prevalence of bothsmoking during pregnancy and ADHDamong children, these findingswarrant future studies on theinterplay between maternal smokingand environmental, genetic, andepigenetic factors.

ACKNOWLEDGMENTS

We thank Mr Jesse Fomin, Ms JarnaLindroos, and Mr Lauri Sillanmäki fortheir technical support.

ABBREVIATIONS

ADHD: attention-deficit/hyperactivity disorder

BWGA: birth weight forgestational age

CI: confidence intervalFHDR: Finnish Hospital Discharge

RegisterFMBR: Finnish Medical Birth

RegisterFMC: Finnish Maternity CohortICD-8: International Classification

of Diseases, Eighth RevisionICD-9: International Classification

of Diseases, Ninth RevisionICD-10: International Classification

of Diseases, 10th RevisionOR: odds ratioPIC: personal identity codeSES: socioeconomic status

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DOI: https://doi.org/10.1542/peds.2018-3144

Accepted for publication Nov 29, 2018

Address correspondence to Andre Sourander, MD, PhD, Department of Child Psychiatry, Research Centre for Child Psychiatry, University of Turku and Turku

University Hospital, Lemminkäisenkatu 3/Teutori (Third floor), 20014 Turku, Finland. E-mail: andsou@utu.fi

PEDIATRICS (ISSN Numbers: Print, 0031-4005; Online, 1098-4275).

Copyright © 2019 by the American Academy of Pediatrics

FINANCIAL DISCLOSURE: The authors have indicated they have no financial relationships relevant to this article to disclose.

FUNDING: Supported by the following funding sources: Academy of Finland (decision 308552), National Institutes of Health, grants 1RO1ES028125-01 and

1(GG012038-01), the Brain and Behavior Research Foundation (Dr Gyllenberg), the Finnish Medical Foundation (Drs Sucksdorff and Gyllenberg), and the University of

Turku Graduate School (Drs Sucksdorff and Chudal). This study belongs to the Finnish Psychiatric Birth Cohort Consortium, which is funded by Academy of Finland.

The funding sources had no role in the design and conduct of the study; collection, management, analysis, or interpretation of the data; preparation, review, or

approval of the manuscript; and decision to submit the manuscript for publication. Funded by the National Institutes of Health (NIH).

POTENTIAL CONFLICT OF INTEREST: The authors have indicated they have no potential conflicts of interest to disclose.

REFERENCES

1. Drake P, Driscoll AK, Mathews TJ.Cigarette smoking during pregnancy:United States, 2016. NCHS Data Brief.2018;(305):1–8

2. Heino A, Vuori E, Gissler M. Perinatalstatistics - parturients, delivers andnewborns 2016 [in Finnish]. Availableat: http://urn.fi/URN:NBN:Fi-fe2017103150386. Accessed August 15,2018

3. Huang L, Wang Y, Zhang L, et al.Maternal smoking and attention-deficit/hyperactivity disorder in offspring:a meta-analysis. Pediatrics. 2018;141(1):e20172465

4. Tiesler CM, Heinrich J. Prenatal nicotineexposure and child behaviouralproblems. Eur Child Adolesc Psychiatry.2014;23(10):913–929

5. Obel C, Linnet KM, Henriksen TB, et al.Smoking during pregnancy andhyperactivity-inattention in theoffspring–comparing results fromthree Nordic cohorts. Int J Epidemiol.2009;38(3):698–705

6. Langley K, Rice F, van den Bree MB,Thapar A. Maternal smoking duringpregnancy as an environmental riskfactor for attention deficit hyperactivitydisorder behaviour. A review. MinervaPediatr. 2005;57(6):359–371

7. Joelsson P, Chudal R, Talati A, SuominenA, Brown AS, Sourander A. Prenatalsmoking exposure andneuropsychiatric comorbidity of ADHD:a finnish nationwide population-based

cohort study. BMC Psychiatry. 2016;16:306

8. Obel C, Zhu JL, Olsen J, et al. The risk ofattention deficit hyperactivity disorderin children exposed to maternalsmoking during pregnancy - a re-examination using a sibling design.J Child Psychol Psychiatry. 2016;57(4):532–537

9. Skoglund C, Chen Q, D’Onofrio BM,Lichtenstein P, Larsson H. Familialconfounding of the association betweenmaternal smoking during pregnancyand ADHD in offspring. J Child PsycholPsychiatry. 2014;55(1):61–68

10. Gustavson K, Ystrom E, Stoltenberg C,et al. Smoking in pregnancy and childADHD. Pediatrics. 2017;139(2):e20162509

11. Langley K, Heron J, Smith GD, Thapar A.Maternal and paternal smoking duringpregnancy and risk of ADHD symptomsin offspring: testing for intrauterineeffects. Am J Epidemiol. 2012;176(3):261–268

12. D’Onofrio BM, Van Hulle CA, Waldman ID,et al. Smoking during pregnancy andoffspring externalizing problems: anexploration of genetic andenvironmental confounds. DevPsychopathol. 2008;20(1):139–164

13. Lindblad F, Hjern A. ADHD after fetalexposure to maternal smoking. NicotineTob Res. 2010;12(4):408–415

14. Shipton D, Tappin DM, Vadiveloo T,Crossley JA, Aitken DA, Chalmers J.

Reliability of self reported smokingstatus by pregnant women forestimating smoking prevalence:a retrospective, cross sectional study.BMJ. 2009;339:b4347

15. Dietz PM, Homa D, England LJ, et al.Estimates of nondisclosure of cigarettesmoking among pregnant andnonpregnant women of reproductiveage in the United States. AmJ Epidemiol. 2011;173(3):355–359

16. Tong VT, Althabe F, Alemán A, et al;Prenatal Tobacco CessationIntervention Collaborative. Accuracy ofself-reported smoking cessation duringpregnancy. Acta Obstet Gynecol Scand.2015;94(1):106–111

17. Connor Gorber S, Schofield-Hurwitz S,Hardt J, Levasseur G, Tremblay M. Theaccuracy of self-reported smoking:a systematic review of the relationshipbetween self-reported and cotinine-assessed smoking status. Nicotine TobRes. 2009;11(1):12–24

18. Dürr DW, Høyer BB, Christensen LH,et al. Tobacco smoking duringpregnancy and risk of adversebehaviour in offspring: a follow-upstudy. Reprod Toxicol. 2015;58:65–72

19. Eskenazi B, Trupin LS. Passive andactive maternal smoking duringpregnancy, as measured by serumcotinine, and postnatal smokeexposure. II. Effects onneurodevelopment at age 5 years. AmJ Epidemiol. 1995;142(suppl 9):S19–S29

8 SOURANDER et al by guest on March 15, 2019www.aappublications.org/newsDownloaded from

20. University of Oulu. Biobank Borealis ofNorthern Finland – from bench tobedside. Available at: www.oulu.fi/university/node/38474. Accessed March14, 2018

21. Gissler M, Surcel H. Combining healthregister data and biobank data. StatJ IAOS. 2012;28(1–2):53–58

22. World Health Organization.International Classification of Diseases,10th Revision. Geneva, Switzerland:World Health Organization; 1992

23. World Health Organization.International Classification of Diseases,Ninth Revision. Geneva, Switzerland:World Health Organization; 1977

24. World Health Organization.International Classification of Diseases,Eighth Revision. Geneva, Switzerland:World Health Organization; 1967

25. Joelsson P, Chudal R, Gyllenberg D, et al.Demographic characteristics andpsychiatric comorbidity of children andadolescents diagnosed with ADHD inspecialized healthcare. Child PsychiatryHum Dev. 2016;47(4):574–582

26. Sankilampi U, Hannila ML, Saari A,Gissler M, Dunkel L. New population-based references for birth weight,length, and head circumference insingletons and twins from 23 to 43gestation weeks. Ann Med. 2013;45(5–6):446–454

27. Rothman KJ, Greenland S. ModernEpidemiology. 2nd ed. Philadelphia, PA:Lippincott-Raven; 1998

28. Niemelä S, Sourander A, Surcel HM,et al. Prenatal nicotine exposure andrisk of schizophrenia among offspringin a national birth cohort. AmJ Psychiatry. 2016;173(8):799–806

29. Kapeu AS, Luostarinen T, Jellum E, et al.Is smoking an independent risk factor

for invasive cervical cancer? A nestedcase-control study within Nordicbiobanks. Am J Epidemiol. 2009;169(4):480–488

30. Jauniaux E, Gulbis B, Acharya G, Thiry P,Rodeck C. Maternal tobacco exposureand cotinine levels in fetal fluids in thefirst half of pregnancy. Obstet Gynecol.1999;93(1):25–29

31. Wickström R. Effects of nicotine duringpregnancy: human and experimentalevidence. Curr Neuropharmacol. 2007;5(3):213–222

32. Bublitz MH, Stroud LR. Maternalsmoking during pregnancy andoffspring brain structure and function:review and agenda for future research.Nicotine Tob Res. 2012;14(4):388–397

33. Ekblad M, Korkeila J, Parkkola R,Lapinleimu H, Haataja L, Lehtonen L;PIPARI Study Group. Maternal smokingduring pregnancy and regional brainvolumes in preterm infants. J Pediatr.2010;156(2):185–190.e1

34. Ekblad M, Korkeila J, Lehtonen L.Smoking during pregnancy affectsfoetal brain development. Acta Paediatr.2015;104(1):12–18

35. Obel C, Olsen J, Henriksen TB, et al. Ismaternal smoking during pregnancya risk factor for hyperkinetic disorder?–Findings from a sibling design. IntJ Epidemiol. 2011;40(2):338–345

36. Thapar A, Rice F, Hay D, et al. Prenatalsmoking might not cause attention-deficit/hyperactivity disorder: evidencefrom a novel design. Biol Psychiatry.2009;66(8):722–727

37. Ekblad M, Lehtonen L, Korkeila J, GisslerM. Maternal smoking during pregnancyand the risk of psychiatric morbidity insingleton sibling pairs. Nicotine TobRes. 2017;19(5):597–604

38. Dolan CV, Geels L, Vink JM, et al. Testingcausal effects of maternal smokingduring pregnancy on offspring’sexternalizing and internalizing behavior.Behav Genet. 2016;46(3):378–388

39. Marceau K, Cinnamon Bidwell L, KarolyHC, et al. Within-family effects ofsmoking during pregnancy on ADHD:the importance of phenotype. J AbnormChild Psychol. 2018;46(4):685–699

40. Frisell T, Öberg S, Kuja-Halkola R,Sjölander A. Sibling comparisondesigns: bias from non-sharedconfounders and measurement error.Epidemiology. 2012;23(5):713–720

41. Rzehak P, Saffery R, Reischl E, et al;European Childhood Obesity Trial Studygroup. Maternal smoking duringpregnancy and DNA-methylation inchildren at age 5.5 years: epigenome-wide-analysis in the EuropeanChildhood Obesity Project (CHOP)-study.PLoS One. 2016;11(5):e0155554

42. Knopik VS, Maccani MA, Francazio S,McGeary JE. The epigenetics ofmaternal cigarette smoking duringpregnancy and effects on childdevelopment. Dev Psychopathol. 2012;24(4):1377–1390

43. Tandon M, Si X, Belden A, Spitznagel E,Wakschlag LS, Luby J. Parentingpractices in pregnancy smokerscompared to non smokers. J Clin MedRes. 2013;5(2):84–91

44. Pajulo M, Savonlahti E, Sourander A,Helenius H, Piha J. Antenataldepression, substance dependency andsocial support. J Affect Disord. 2001;65(1):9–17

45. World Health Organization. MPOWER: APolicy Package to Reverse the TobaccoEpidemic. Geneva, Switzerland: WorldHealth Organization; 2008

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