In this chapter, we shall consider some topics of interest to the student of embryology that do not deal with the development of any particular organ as such.

GROWTH OF THE EMBRYO

After its formation, the embryonic disc undergoes folding. Th is folding leads to major changes in body form (during the fourth to eighth weeks of fetal life). Th e embryo acquires the external features of a human being. All organ systems are formed. Th e following description summarises the changes in external form of the embryo during the fourth to eight weeks. Th e main developmental events taking place are also mentioned.

Fourth Week

At the beginning of the fourth week, the embryonic disc undergoes folding, and it becomes C shaped. Folding of the embryo also leads to the formation of the foregut, the midgut and the hindgut. Th e pharyngeal arches start appearing at the end of the fourth week. Th e heart produces a large prominence on the ventral aspect of the embryo. Th e heart starts functioning by the end of the fourth week. At this stage the forebrain is the most cranial and most prominent structure of the embryo. Th e upper limbs appear as paddle shaped buds. Th e otic pits and the lens placodes become visible.

Fifth Week

Further development of the head and of the face occurs rapidly. Th e mesonephric kidney starts forming.

Sixth Week

Th e upper limbs show further diff erentiation so that the elbow and digits can be recognised. Th e lower limb buds and the external ear start forming. Further development is seen in the eyes, eyelids and ear.

Some General Considerations

Chapter CD 2

Human Embryology

Seventh Week

The hand shows formation of digits. Ossification of bones starts in the upper limb.

Eighth Week

The limbs and digits are fully formed. Movements start taking place in the limbs. Bone formation begins in the lower limbs. The neck appears between the head and the thorax. The external genitalia may start showing sex differences. The tail disappears. At the end of the embryonic period the embryo can be recognised as human even though its size (CR length) is only about 30 mm.

DETERMINING THE AGE OF AN EMBRYO

It is important to know the age of a developing embryo because this can affect clinical procedures like amniocentesis or chorionic villus sampling. The exact age of a living embryo can be found out only when the date of conception (fertilisation) is known. This is almost impossible because fertilisation is an internal event. Usually, the age has to be determined indirectly. Estimation of gestational age from menstrual history may be unreliable: 1. as the expectant mother may not know the exact date of the last normal menstrual period,

or 2. her menstrual cycles may be irregular. The somites begin to be seen in embryos about twenty-one days old. Embryos younger than this are called presomite embryos and their age is reckoned in days. Once the somites appear, the age is described in terms of the number of somites present, e.g. one-somite stage, four-somite stage, etc. When the embryo is about 30 days old, it is large enough to be measured. However, the measurement of the length of an aborted embryo is not as simple as it sounds,

as the embryo is bent on itself and cannot be straightened without fear of damage to it. Hence, instead of measuring its full length we measure what is called the crownrump (C.R.) length (Fig. CD-2.1). CR length is measured from the vertex of the skull to the midpoint between the apices of the buttocks. At the present time it is possible to measure the C.R. length of an embryo accurately, within the womb of the mother, with the help of an ultrasound machine. The C.R. length of a one-month-old embryo is about 5 mm, and that of a two-month-old embryo is about 30 mm (Fig. CD-2.2). At full term the C.R. length is about 300 mm. However, because of variations in the degree of curvature of fetuses, C.R. length is not a very accurate index of fetal Fig. CD-2.1: Measurement of

C.R. length.

Chapter CD 2 – Some General Considerations

age. Various other measurements are also used. For example (with the use of ultrasound) we can measure the dimensions of some parts of the fetus (e.g.,head, foot length).

Carnegie Embryonic Staging System

This system divides the development of an embryo into various stages. It is a numeric system for characterising developmental stages. This system is used internationally by embryologists and research workers. The stages range from 1 to 23 and cover 1 to 56 days of embryonic development.

FURTHER GROWTH OF THE FETUS

At the beginning of the fetal period (ninth week to third month) the embryo has developed into a recognisable human being and the primordia of all organ system have formed. During the fetal period, development is mainly directed towards the rapid growth in body size and towards differentiation of tissues, organs and organ systems. During this period the growth of the head is slow as compared to that of the rest of body. At the beginning of third month the head is half the CR length, while at birth it is about one fourth of CR length. Fetal weight gain is very rapid (Fig. CD-2.2) in the last month of pregnancy. A brief summary of growth in the fetal period is given below.

Third Month (9 to 12 weeks)

The eyes and ears are now in their definitive positions. As a result the fetus becomes more human. The process of ossification is seen in all long bones. Intestinal loops that had herniated out of the abdominal cavity now return into it. The male and female external genitalia can be visualised using ultrasound imaging. Urine formation begins.

Fourth Month (13 to 16 weeks)

The proportion of the size of the head relative to the rest of the body is less as compared to that in the third fetal month. The length of the fetus increases rapidly.

Fig. CD-2.2: Growth of fetus in length and weight.

Human Embryology

The lower limbs reach their final relative length as compared to the rest of the body. Movements in the limbs are not very strong but can be seen on ultrasound examination.

Fifth Month (17 to 20 weeks)

The length of fetus increases. Increase in weight is however slow. The mother can now feel the movements of the fetus (quickening). Hair on the the head, and eyebrows, can be seen. The skin becomes covered with sebaceous gland secretion (vernix caseosa).

Sixth Month (21 to 24 weeks)

The skin is wrinkled due to absence of subcutaneous tissue. Lung alveoli begin to secrete surfactant, which helps to maintain the patency of the alveoli of the lung. This is a sign of the maturity of the respiratory system. Rapid eye movements begin. The fetus now starts gaining body weight rapidly.

Seventh Month (25 to 29 weeks)

Blood formation now begins to shift from spleen to bone marrow. The central nervous system is now mature enough to be able to control respiration, if the fetus is born at this stage. The respiratory system is mature enough to perform gaseous exchanges between pulmonary vessels and lung alveoli. A fetus born prematurely in the seventh month can survive. Such fetuses are said to be viable.

Eighth Month (30 to 34 weeks)

The skin is smooth due to deposition of subcutaneous fat. It is pink due to increase in blood supply. Body weight increases rapidly. The pupillary light reflex can be elicited.

Ninth Month (35 to 38 weeks)

The circumference of the head is almost the same as that of the abdomen in the terminal weeks of pregnancy. At the end of the fetal period the skull has the largest circumference of all parts of body. The testes usually lie in the scrotum. The length of the foot is slightly more than the length of the femur. Most fetuses are born within few a days of expected date of delivery. From the date of fertilization the expected date of delivery is calculated as 266 days or 38 weeks (eight and three fourth calendar months or nine and a half lunar months).

Chapter CD 2 – Some General Considerations

DETERMINING THE AGE OF A LIVING FETUS

The ability to find out the age of a living fetus is very important clinically. The age of such a fetus can be determined by making measurements using ultrasound examination. 1. Between 7 to 14 weeks, the CR length can be measured. 2. Estimation of fetal age in the second and third trimesters of pregnancy is based on the

measurements of various body parts. These are: a. biparietal diameter, b. circumference of the head, c. circumference of the abdomen d. length of femur e. and foot length.

CONTROL OF FETAL GROWTH

Intrauterine growth of the fetus is influenced by maternal factors, placental factors and fetal factors.

Maternal Factors

Adequate availability of nutrition in maternal blood and its transfer across the placenta are essential for normal growth of the fetus. Malnutrition in the mother affects fetal growth and can possibly cause fetal malformations. As a rule, maternal hormones do not pass through the placenta and hence they cannot affect fetal growth. However, they can influence the fetus indirectly by controlling maternal metabolic processes.

Placental Factors

1. Hormones secreted by the placenta can influence the fetus indirectly by influencing maternal metabolism. For example, somatomammotropin (hCS) secreted by the placenta has an anti-insulin effect leading to increased plasma levels of glucose and amino acids in maternal blood. The availability of these to the fetus is, thereby, increased.

2. Placental hormones also have a direct influence on fetal growth. Somatomammotropin increases fetal growth. Human chorionic gonadotropin (hCG) stimulates growth of the fetal testis.

Fetal Factors

Fetal growth is influenced by genetic factors. However, genetic factors that determine the height of the individual operate mainly in postnatal life (through the action of the growth hormone and of the thyroid hormone). Fetal endocrine glands start functioning near the middle of intrauterine life. The effects of hormones produced by them may be different from those seen in postnatal life, the modifications being necessary for requirements of the fetus.

Human Embryology

For example, the fetal adrenal gland starts producing cortisol in the 9th week. In an adult, cortisol has a catabolic effect. To prevent this, cortisol secreted by the fetus is converted to cortisone (which does not have this effect). Growth hormone (produced by the hypophysis cerebri) and thyroid hormones have very little effect on fetal growth. Infants in whom these hormones are deficient do not show growth retardation. However, as discussed earlier, sex hormones produced by developing gonads greatly influence differentiation of genital organs in both sexes.

Fetal Growth Retardation

When the growth of a fetus is less than that seen in 90 per cent of fetuses (i.e. it is below the 90th percentile) the phenomenon is described as intrauterine growth retardation (IUGR). Such infants are also described as small for gestational age. Such fetuses have an increased risk of congenital malformations. Apart from genetic factors like chromosomal abnormalities, growth retardation can also be caused by infections, poor nutrition, cigarette smoking, alcohol and use of harmful drugs by the mother.

CAUSATION OF CONGENITAL ANOMALIES (TERATOGENESIS)

The study of congenital malformations constitutes the science of teratology. Factors that cause anomalies are called teratogens. The development of the embryo is dependent primarily on genetic influences. However, environmental conditions can also exert an important effect. For example, a chick embryo can develop properly only if the egg is kept at a suitable temperature. It, therefore, follows that congenital anomalies may occur either as a result of genetic or environmental defects, or by a combination of both. Embryos with major abnormalities are aborted early in pregnancy, and this may occur even before the mother is aware of the pregnancy. According to some estimates the total number of abnormal embryos may be as high as 50 per cent and their spontaneous abortion may be nature’s way of reducing the birth of malformed babies. In spite of this fact, two to three per cent of infants born alive, show one or more congenital malformations. Some anomalies are not obvious at birth but are discovered later. The total incidence of malformations may, therefore, be as high as five per cent of live births. These figures are cited to highlight the great importance of the need to understand the causation of congenital abnormalities. With regard to the mode of action of teratogens, some general principles may be stated as follows: 1. The susceptibility to a teratogen, and the degree of damage it causes, depends upon the

stage of embryonic development at which the embryo or fetus is exposed to the teratogen. When a teratogen acts before differentiation of germ layers, the effects are drastic and often lead to death of the embryo.

The organ systems of the fetus are established between 3 and 8 weeks of pregnancy, and this is referred to as the embryonic period or period of organogenesis. Most anomalies are produced during this period. Unfortunately this is an early stage of pregnancy and

Chapter CD 2 – Some General Considerations

the mother may not even be aware of her pregnancy. Therefore she may not take the necessary precautions.She may keep on consuming harmful products like drugs, alcohol or cigarettes.

During the fetal period that follows, teratogenic influences become much less severe. 2. The type of malformation produced depends on the exact timing of the teratogenic

influence. Each organ seems to have a critical period during which it is most sensitive to teratogens.

3. The susceptibility to a teratogen is influenced by genetic factors. A fetus of one genotype can be much more susceptible to the same teratogen than a fetus of another genotype.

4. Teratogenic agents act by influencing metabolic processes. 5. The dose and duration of exposure to teratogen is also important. High concentration and

long period of exposure to a teratogen is relatively more harmful. About 80 per cent of all congenital malformations are produced by a combination of genetic and environmental factors. Of the remaining 20 per cent, about half are caused exclusively by genetic or chromosomal factors and the remaining half exclusively by environmental factors.

Hereditary Causes

Anomalies may be caused by defects in a specific chromosome or in a specific gene. Chromosomal defects owe their effects to the absence of certain genes, or presence of extraneous ones on them. Hence, all hereditary defects are ultimately caused by failure of the cells to synthesise the right proteins (specially enzymes) at the right time. In producing an anomaly, the genetic defect may directly affect the organ, or may have an indirect effect. For example, a genetic defect that leads to agenesis of the testis, may indirectly influence the developing external genitalia by interfering with the production of hormones necessary for their development. Similarly, an anomaly of a blood vessel may interfere with the blood supply of an organ and hence adversely affect its development.

Environmental Causes (Teratogens)

1. Infections

Some disease-producing organisms (e.g. viruses) or harmful substances produced by them (toxins) can pass through the placental barrier and reach the fetus. Some of the diseases that can reach the fetus in this way are syphilis, chickenpox, HIV, measles and toxoplasmosis. There is a well-known correlation between a disease known as German measles and congenital anomalies. When the mother suffers from this disease in the early months of pregnancy, the offspring often has cataract (opaque lens of eye), anomalies of the heart, or deafness.

2. Malnutrition

The developing fetus requires all elements of nutrition in adequate quantity for normal development. In experimental animals, deficiencies of vitamins, minerals (like calcium or

Human Embryology

phosphorus), certain trace elements, and of some amino acids have been shown to cause anomalies. It is believed that iodine deficiency causes endemic cretinism. However, the extent to which nutritional deficiencies are responsible for anomalies in humans is controversial.

3. Antigenic Reactions

The body of every animal contains a large number of proteins. The proteins differ not only from species to species, but even amongst individuals of the same species. The body has the ability to recognise any protein that is foreign to it. A foreign protein is often called an antigen. Whenever such a protein enters the body, substances called antibodies are produced and their function is to destroy the antigen. Remember that a protein normally present in one person may act as an antigen when introduced into another person, whose body does not contain it. One such protein present in the blood of most persons is called the Rh-antigen. Persons having it are Rh-positive and those without it are Rh-negative. It is sometimes possible for an Rh-negative mother to have an Rh-positive fetus. Some Rh-antigen from the fetus can enter the mother’s blood. If this happens, the mother’s body produces antibodies against this antigen. These antibodies pass back into the fetal blood where they destroy the blood cells containing the antigen. This breaking up of blood cells is called haemolysis and the disease is called haemolytic disease of the newborn.

4. Drugs and Chemicals

Administration of certain drugs to an expectant mother during the early months of pregnancy is known to cause congenital malformations. The best known of these drugs is thalidomide, which produces varying degrees of agenesis of one or more limbs. Some other drugs known to have significant teratogenic effects are aminopterin (a folic acid antagonist); diphenylhydantoin and trimethadione (used for epilepsy); phenothiazine, lithium, meprobamate, chlordiazepoxide and diazepam (which are used as tranquillizers). Even aspirin in large doses, or cocaine can produce anomalies. Alcohol in fetal blood produces the fetal alcohol syndrome. Every new drug is now tested against such teratogenic effects, and it is recognised that no drug should be given to a pregnant woman unless it is absolutely necessary.

5. Hormones

Administration of synthetic oestrogens or progestins can cause malformations. Progestins ethisterone and norethisterone can cause masculinization of female genitalia. Fetuses exposed to diethylstilbestrol (a synthetic oestrogen) in intrauterine life, show increased incidence of carcinoma of the vagina and cervix in later life. Maternal diabetes can also cause congenital malformations.

Physical Factors

Physical environmental conditions are less likely to influence mammalian embryos that grow within the uterus, as compared to those that grow in eggs, or in water. However, the mammalian embryo is not completely immune to these influences. The greatest danger lies

Chapter CD 2 – Some General Considerations

from radiations of various kinds, including X-rays and radioactivity. These are capable of producing permanent changes (mutations) in the nature of genes, specially in the germ cells, and these in turn can lead to the production of congenital anomalies. Some physical factors that can produce abnormalities are as follows. 1. In experimental animals, hypoxia has been shown to produce anomalies. 2. Abnormal intrauterine environment due to an abnormal site of implantation, due to the

presence of twins, because of an abnormal position of the fetus within the uterus, because of too much amniotic fluid (hydramnios) or because of too little fluid (oligamnios).

3. Insufficient or excessive availability of oxygen. Too much oxygen leads to a condition called retrolental fibroplasia.

4. Hyperthermia or increased body temperature is teratogenic. Increase in temperature may be due to fever secondary to infection, or due to bathing with hot water for long duration. Hyperthermia leads to mental retardation, cleft lip and cleft palate, limb deficiency, spina bifida and anencephaly.

PRENATAL DIAGNOSIS OF FETAL DISEASES AND MALFORMATIONS

A physician can now detect congenital malformation or disease of the fetus even during early pregnancy. Early detection of severe abnormality helps the patient to decide the desirability of early termination of pregnancy. Many procedures are now available by which we can assess the fetal status within the womb of mother. These procedures are described briefly below.

Ultrasonography

Ultrasonography is widely used to assess the condition of the developing fetus in utero because the investigation is cheap, readily available, and has no harmful effects. One can determine fetal age and size, position of placenta, condition of fetal membranes and the presence of multiple births. Birth defects can be detected at an early stage of pregnancy.

Alpha-fetoprotein (AFP) Assay

AFP is normally produced in the liver and in the gut of a fetus in the second trimester of pregnancy. It reaches amniotic fluid and maternal serum through the placenta. Its concentration increases when the fetus is suffering from congenital malformations. However, its concentration decreases in a few chromosomal anomalies like Down’s syndrome, and trisomy 18.

Amniocentesis

In this procedure a needle is passed through the abdominal wall of the mother and made to enter the amniotic cavity of the fetus. About 20 ml of amniotic fluid is withdrawn. This procedure is usually performed between the 15 and 18 weeks of gestation. Amniotic fluid obtained is used for chemical analysis (alpha fetal protein and acetylcholinesterase). Amniotic fluid also

Human Embryology

contains epithelial cells from the skin of the fetus. These can be used for karyotyping. These epithelial cells can also be used for determining the sex of the fetus by the simple procedure of detecting sex chromatin. Such tests are however illegal in India as they often lead to female feticide.

Chorionic Villus Sampling

In this procedure a biopsy of chorionic villi is taken by entering the uterus either by an abdominal route or by a cervical route. The procedure is performed between 10 to 12 weeks of gestation. The biopsies are used for detecting chromosomal abnormalities, inborn errors of metabolism or X-linked disorders.

Fetoscopy

The fetal body may be directly observed for congenital anomalies by using a fiber-optic lighting instrument. This procedure is usually performed during 17 to 20 weeks of gestation.

MR Imaging

Magnetic resonance imaging of a fetus can be performed to get further information about conditions that have been detected in ultrasonographic images. MRI is safe and provides high soft tissue contrast and resolution.

Percutaneous Umbilical Cord Blood Sampling (PUBS)

In this procedure fetal blood is drawn from the umbilical vein for diagnosis of many conditions. The same procedure is also used for blood transfusion into the fetus or for injection of drugs.

FETAL TREATMENT

In selected cases, treatment of a third trimester fetus is possible with limited success. Such treatment has been used for fetal anemias, hemolytic disease, fetal cardiac arrhythmia, thyroid dysfunction and surgical corrections.

Fetal Transfusion

Fetal transfusion is given directly into the umbilical vein by the PUBS procedure.

Fetal Surgery

Surgical correction of some birth defects in the fetus has become possible. In this procedure the uterus is opened by cesarean section and the fetus is operated upon directly. After repair of the defect the fetus is placed back into the uterus. Most of the surgeries in utero are performed after 28th weeks of pregnancy.