Somjate Manipalviratn, MD
Bradley Trivax, MD
Andy Huang, MD
GENETIC DISORDERS
MENDELIAN LAWS OF INHERITANCE
1. Types of Inheritance
Autosomal Dominant
In autosomal dominant inheritance, it is assumed that a mutation has occurred in 1 gene of an allelic pair and that the presence of this new gene produces enough of the changed protein to give a different phenotypic effect. Environment must also be considered because the effect may vary under different environmental conditions. The following are characteristic of autosomal dominant inheritance:
1. The trait appears with equal frequency in both sexes.
2. For inheritance to take place, at least 1 parent must have the trait unless a new mutation has just occurred.
3. When a homozygous individual is mated to a normal individual, all offspring will carry the trait. When a heterozygous individual is mated to a normal individual, 50% of the offspring will show the trait.
4. If the trait is rare, most persons demonstrating it will be heterozygous (Table 3–1).
Table 3–1. Examples of autosomal dominant conditions and traits.
Autosomal Recessive
The mutant gene will not be capable of producing a new characteristic in the heterozygous state in this circumstance under customary environmental conditions—ie, with 50% of the genetic material producing the new protein, the phenotypic effect will not be different from that of the normal trait. When the environment is manipulated, the recessive trait occasionally becomes dominant. The characteristics of this form of inheritance are as follows:
1. The characteristic will occur with equal frequency in both sexes.
2. For the characteristic to be present, both parents must be carriers of the recessive trait.
3. If both parents are homozygous for the recessive trait, all offspring will have it.
4. If both parents are heterozygous for the recessive trait, 25% of the offspring will have it.
5. In pedigrees showing frequent occurrence of individuals with rare recessive characteristics, consanguinity is often present (Table 3–2).
Table 3–2. Examples of autosomal recessive conditions and traits.
X-Linked Recessive
This condition occurs when a gene on the X chromosome undergoes mutation and the new protein formed as a result of this mutation is incapable of producing a change in phenotype characteristic in the heterozygous state. Because the male has only 1 X chromosome, the presence of this mutant will allow for expression should it occur in the male. The following are characteristic of this form of inheritance:
1. The condition occurs more commonly in males than in females.
2. If both parents are normal and an affected male is produced, it must be assumed that the mother is a carrier of the trait.
3. If the father is affected and an affected male is produced, the mother must be at least heterozygous for the trait.
4. A female with the trait may be produced in 1 of 2 ways. (A) She may inherit a recessive gene from both her mother and her father; this suggests that the father is affected and the mother is heterozygous. (B) She may inherit a recessive gene from 1 of her parents and may express the recessive characteristic as a function of the Lyon hypothesis; this assumes that all females are mosaics for their functioning X chromosome. It is theorized that this occurs because at about the time of implantation, each cell in the developing female embryo selects 1 X chromosome as its functioning X and that all progeny cells thereafter use this X chromosome as their functioning X chromosome. The other X chromosome becomes inactive. Because this selection is done on a random basis, it is conceivable that some females will be produced who will be using primarily the X chromosome bearing the recessive gene. Thus, a genotypically heterozygous individual may demonstrate a recessive characteristic phenotypically on this basis (Table 3–3).
Table 3–3. Examples of X-linked recessive conditions and traits.
X-Linked Dominant
In this situation, the mutation will produce a protein that, when present in the heterozygous state, is sufficient to cause a change in characteristic. The following are characteristic of this type of inheritance:
1. The characteristic occurs with the same frequency in males and females.
2. An affected male mated to a normal female will produce the characteristic in 50% of the offspring.
3. An affected homozygous female mated to a normal male will produce the affected characteristic in all offspring.
4. A heterozygous female mated to a normal male will produce the characteristic in 50% of the offspring.
5. Occasional heterozygous females may not show the dominant trait on the basis of the Lyon hypothesis (Table 3–4).
Table 3–4. Examples of X-linked dominant conditions and traits.
2. Applications of Mendelian Laws
Identification of Carriers
When a recessive characteristic is present in a population, carriers may be identified in a variety of ways. If the gene is responsible for the production of a protein (eg, an enzyme), the carrier often possesses 50% of the amount of the substance present in homozygous normal persons. Such a circumstance is found in galactosemia, where the carriers will have approximately half as much galactose-1-phosphate uridyltransferase activity in red cells as do noncarrier normal individuals.
At times, the level of the affected enzyme may be only slightly below normal, and a challenge with the substance to be acted upon may be required before the carrier can be identified. An example is seen in carriers of phenylketonuria, in whom the deficiency in phenylalanine hydroxylase is in the liver cells, and serum levels may not be much lower than normal. Nonetheless, when the individual is given an oral loading dose of phenylalanine, plasma phenylalanine levels may remain high because the enzyme is not present in sufficient quantities to act upon this substance properly.
In still other situations where the 2 alleles produce different proteins that can be measured, a carrier state will have 50% of the normal protein and 50% of the other protein. Such a situation is seen in sickle cell trait, where 1 gene is producing hemoglobin A and the other hemoglobin S. Thus, the individual has half the amount of hemoglobin A as a normal person and half the hemoglobin S of a person with sickle cell anemia. An interesting but important problem involves the detection of carriers of cystic fibrosis. This is the most common autosomal recessive disease in Caucasian populations of European background, occurring in 1 in 2500 births in such populations but found in the carrier state in 1 in 25 Americans. By 1990, over 230 alleles of the single gene responsible have been discovered. The gene is known as the cystic fibrosis transmembrane conductance regulator (CFTR), and the most common mutation, delta F508, accounts for about 70% of all mutations, with 5 specific point mutations accounting for over 85% of cases. Because so many alleles are present, population screening poses logistical problems that have yet to be worked out. Most programs screen for the most common mutations using DNA replication and amplification studies.
3. Polygenic Inheritance
Polygenic inheritance is defined as the inheritance of a single phenotypic feature as a result of the effects of many genes. Most physical features in humans are determined by polygenic inheritance. Many common malformations are determined in this way also. For example, cleft palate with or without cleft lip, clubfoot, anencephaly, meningomyelocele, dislocation of the hip, and pyloric stenosis each occur with a frequency of 0.5–2 per 1000 in white populations. Altogether, these anomalies account for slightly less than half of single primary defects noted in early infancy. They are present in siblings of affected infants—when both parents are normal—at a rate of 2–5%. They are also found more commonly among relatives than in the general population. The increase in incidence is not environmentally induced because the frequency of such abnormalities in monozygotic twins is 4–8 times that of dizygotic twins and other siblings. The higher incidence in monozygotic twins is called concordance.
Sex also plays a role. Certain conditions appear to be transmitted by polygenic inheritance and are passed on more frequently by the mother who is affected than by the father who is affected. Cleft lip occurs in 6% of the offspring of women with cleft lip, as opposed to 2.8% of offspring of men with cleft lip.
Many racial variations in diseases are believed to be transmitted by polygenic inheritance, making racial background a determinant of how prone an individual will be to a particular defect. In addition, as a general rule, the more severe a defect, the more likely it is to occur in subsequent siblings. Thus, siblings of children with bilateral cleft lip are more likely to have the defect than are those of children with unilateral cleft lip.
Environment undoubtedly plays a role in polygenic inheritance, because seasonal variations alter some defects and their occurrence rate from country to country in similar populations.
EPIGENETIC
Epigenetic is the regulation of gene expression not encoded in the nucleotide sequence of the gene. Gene expression can either be turned on or off by DNA methylation or histone modification (methylation, acetylation, phosphorylation, ubiquitination, or ADP-ribosylation). Epigenetic can subsequently be inherited by its descendants.
Genomic Imprinting
Genomic imprinting is an epigenetic process by which the male and female genomes are differently expressed. The imprinting mark on genes is either by DNA methylation or histone modification. The imprinting patterns are different according to the parental origin of the genes. Genomic imprints are erased in primordial germ cells and reestablished again during gametogenesis. The imprinting process is completed by the time of round spermatids formation in males and at ovulation of metaphase-II oocytes in females. The imprinted genes survive the global waves of DNA demethylation and remethylation during early embryonic development. In normal children, 1 set of chromosomes is derived from the father and the other from the mother. If both sets of chromosomes are from only 1 parent, the imprinted gene expression will be unbalanced. Prader-Willi syndrome and Angelman syndrome are examples of imprinting disorders. In Prader-Willi syndrome, both 15q13 regions are from the father, whereas in Angelman syndrome, both 15q13 regions are from the mother.
CYTOGENETICS
1. Identification of Chromosomes
In 1960, 1963, 1965, and 1971, international meetings were held in Denver, London, Chicago, and Paris, respectively, for the purpose of standardizing the nomenclature of human chromosomes. These meetings resulted in a decision that all autosomal pairs should be numbered in order of decreasing size from 1 to 22. Autosomes are divided into groups based on their morphology, and these groups are labeled by the letters A–G. Thus, the A group is comprised of pairs 1–3; the B group, pairs 4 and 5; the C group, pairs 6–12; the D group, pairs 13–15; the E group, pairs 16–18; the F group, pairs 19 and 20; and the G group, pairs 21 and 22. The sex chromosomes are labeled X and Y, the X chromosome being similar in size and morphology to the number 7 pair and thus frequently included in the C group (C-X) and the Y chromosome being similar in morphology and size to the G group (G-Y) (Fig. 3–1).
Figure 3–1. Karyotype of a normal male demonstrating R banding.
The short arm of a chromosome is labeled p and the long arm q. If a translocation occurs in which the short arm of a chromosome is added to another chromosome, it is written p +. If the short arm is lost, it is p−. The same can be said for the long arm (q+ and q−).
It has been impossible to separate several chromosome pairs from one another on a strictly morphologic basis because the morphologic variations have been too slight. However, there are other means of identifying each chromosome pair in the karyotype. The first of these is the incorporation of 3H-thymidine, known as the autoradiographic technique. This procedure involves the incorporation of radioactive thymidine into growing cells in tissue culture just before they are harvested. Cells that are actively undergoing DNA replication will pick up the radioactive thymidine, and the chromosomes will demonstrate areas of activity. Each chromosome will incorporate thymidine in a different pattern, and several chromosomes can therefore be identified by their labeling pattern. Nonetheless, with this method it is not possible to identify each chromosome, although it is possible to identify chromosomes involved in pathologic conditions, eg, D1 trisomy and Down syndrome.
Innovative staining techniques have made it possible to identify individual chromosomes in the karyotype and to identify small anomalies that might have evaded the observer using older methods. These involve identification of chromosome banding by a variety of staining techniques, at times with predigestion with proteolytic agents. Some of the more commonly used techniques are the following:
Q banding: Fixed chromosome spreads are stained without any pretreatment using quinacrine mustard, quinacrine, or other fluorescent dyes and observed with a fluorescence microscope.
G banding: Preparations are incubated in a variety of saline solutions using any 1 of several pretreatments and stained with Giemsa’s stain.
R banding: Preparations are incubated in buffer solutions at high temperatures or at special pH and stained with Giemsa’s stain. This process yields the reverse bands of G banding (Fig. 3–1).
C banding: Preparations are either heated in saline to temperatures just below boiling or treated with certain alkali solutions and then stained with Giemsa’s stain. This process causes prominent bands to develop in the region of the centromeres.
2. Cell Division
Each body cell goes through successive stages in its life cycle. As a landmark, cell division can be considered as the beginning of a cycle. Following this, the first phase, which is quite long but depends on how rapidly the particular cell is multiplying, is called the G1 stage. During this stage, the cell is primarily concerned with carrying out its function. Following this, the S stage, or period of DNA synthesis, takes place. Next there is a somewhat shorter stage, the G2 stage, during which time DNA synthesis is completed and chromosome replication begins. Following this comes the M stage, when cell division occurs.
Somatic cells undergo division by a process known as mitosis (Fig. 3–2). This is divided into 4 periods. The first is prophase, during which the chromosome filaments shorten, thicken, and become visible. At this time they can be seen to be composed of 2 long parallel spiral strands lying adjacent to one another and containing a small clear structure known as the centromere. As prophase continues, the strands continue to unwind and may be recognized as chromatids. At the end of prophase, the nuclear membrane disappears and metaphase begins. This stage is heralded by the formation of a spindle and the lining up of the chromosomes in pairs on the spindle. Following this, anaphase occurs, at which time the centromere divides and each daughter chromatid goes to 1 of the poles of the spindle. Telophase then ensues, at which time the spindle breaks and cell cytoplasm divides. A nuclear membrane now forms, and mitosis is complete. Each daughter cell has received chromosome material equal in amount and identical to that of the parent cell. Because each cell contains 2 chromosomes of each pair and a total of 46 chromosomes, a cell is considered to be diploid. Occasionally, an error takes place on the spindle, and instead of chromosomes dividing, with identical chromatids going to each daughter cell, an extra chromatid goes to 1 daughter cell and the other lacks that particular member. After completion of cell division, this leads to a trisomic state (an extra dose of that chromosome) in 1 daughter cell and a monosomic state (a missing dose of the chromosome) in the other daughter cell. Any chromosome in the karyotype may be involved in such a process, which is known as mitotic nondisjunction. If these cells thrive and produce their own progeny, a new cell line is established within the individual. The individual then has more than 1 cell line and is known as a mosaic. A variety of combinations and permutations have occurred in humans.
Figure 3–2. Mitosis of a somatic cell.
Germ cells undergo division for the production of eggs and sperm by a process known as meiosis. In the female it is known as oogenesis and in the male as spermatogenesis. The process that produces the egg and the sperm for fertilization essentially reduces the chromosome number from 46 to 23 and changes the normal diploid cell to an aneuploid cell, ie, a cell that has only 1 member of each chromosome pair. Following fertilization and the fusion of the 2 pronuclei, the diploid status is reestablished.
Meiosis can be divided into several stages (Fig. 3–3). The first is prophase I. Early prophase is known as the leptotene stage, during which chromatin condenses and becomes visible as a single elongated threadlike structure. This is followed by the zygotene stage, when the single threadlike chromosomes migrate toward the equatorial plate of the nucleus. At this stage, homologous chromosomes become arranged close to one another to form bivalents that exchange materials at several points known as synapses. In this way, genetic material located on 1 member of a pair is exchanged with similar material located on the other member of a pair. Next comes the pachytene stage in which the chromosomes contract to become shorter and thicker. During this stage, each chromosome splits longitudinally into 2 chromatids united at the centromere. Thus, the bivalent becomes a structure composed of 4 closely opposed chromatids known as a tetrad. The human cell in the pachytene stage demonstrates 23 tetrads. This stage is followed by the diplotene stage, in which the chromosomes of the bivalent are held together only at certain points called bridges or chiasms. It is at these points that crossover takes place. The sister chromatids are joined at the centromere so that crossover can only take place between chromatids of homologous chromosomes and not between identical sister chromatids. In the case of males, the X and Y chromosomes are not involved in crossover. This stage is followed by the last stage of prophase, known as diakinesis. Here the bivalents contract, and the chiasms move toward the end of the chromosome. The homologs pull apart, and the nuclear membrane disappears. This is the end of prophase I.
Figure 3–3. Meiosis in the human.
Metaphase I follows. At this time, the bivalents are now highly contracted and align themselves along the equatorial plate of the cell. Paternal and maternal chromosomes line up at random. This stage is then followed by anaphase I and telophase I, which are quite similar to the corresponding events in mitosis. However, the difference is that in meiosis the homologous chromosome of the bivalent pair separates and not the sister chromatids. The homologous bivalents pull apart, 1 going to each pole of the spindle, following which 2 daughter cells are formed at telophase I.
Metaphase, anaphase, and telophase of meiosis II take place next. A new spindle forms in metaphase, the chromosomes align along the equatorial plate, and, as anaphase occurs, the chromatids pull apart, 1 each going to a daughter cell. This represents a true division of the centromere. Telophase then supervenes, with reconstitution of the nuclear membrane and final cell division. At the end, a haploid number of chromosomes is present in each daughter cell (Fig. 3–3). In the case of spermatogenesis, both daughter cells are similar, forming 2 separate sperms. In the case of oogenesis, only 1 egg is produced, the nuclear material of the other daughter cell being present and intact but with very little cytoplasm, this being known as the polar body. A polar body is formed at the end of meiosis I and the end of meiosis II. Thus, each spermatogonium produces 4 sperms at the end of meiosis, whereas each oogonium produces 1 egg and 2 polar bodies.
Nondisjunction may also occur in meiosis. When it does, both members of the chromosome pair go to 1 daughter cell and none to the other. If the daughter cell that receives the entire pair is the egg, and fertilization ensues, a triple dose of the chromosome, or trisomy, will occur. If the daughter cell receiving no members of the pair is fertilized, a monosomic state will result. In the case of autosomes, this is lethal, and a very early abortion will follow. In the case of the sex chromosome, the condition may not be lethal, and examples of both trisomy and monosomy have been seen in humans. Any chromosome pair may be involved in trisomic or monosomic conditions.
3. Abnormalities in Chromosome Morphology & Number
As has been stated, nondisjunction may give rise to conditions of trisomy. In these cases, the morphology of the chromosome is not affected, but the chromosome number is. Be this as it may, breaks and rearrangements in chromosomes may have a variety of results. If 2 chromosomes undergo breaks and exchange chromatin material between them, the outcome is 2 morphologically new chromosomes known as translocations. If a break in a chromosome takes place and the fragment is lost, deletion has occurred. If the deletion is such that the cell cannot survive, the condition may be lethal. Nonetheless, several examples of deleted chromosomes in individuals who have survived have been identified. If a break takes place at either end of a chromosome and the chromosome heals by having the 2 ends fuse together, a ring chromosome is formed. Examples of these have been seen clinically in all of the chromosomes of the karyotype, and generally they exhibit a variety of phenotypic abnormalities.
At times a chromosome will divide by a horizontal rather than longitudinal split of the centromere. This leaves each daughter cell with a double dose of 1 of the arms of the chromosome. Thus, 1 daughter cell receives both long arms and the other both short arms of the chromosome. Such a chromosome is referred to as an isochromosome, the individual being essentially trisomic for 1 arm and monosomic for the other arm of the chromosome. Examples of this abnormality have been seen in humans.
Another anomaly that has been recognized is the occurrence of 2 breaks within the chromosome and rotation of the center fragment 180 degrees. Thus, the realignment allows for a change in morphology of the chromosome, although the original number of genes is preserved. This is called an inversion. At meiosis, however, the chromosome has difficulty in undergoing chiasm formation, and abnormal rearrangements of this chromosome, leading to partial duplications and partial losses of chromatin material, do take place. This situation may lead to several bizarre anomalies. If the centromere is involved in the inversion, the condition is called a pericentric inversion.
Breaks occasionally occur in 2 chromosomes, and a portion of 1 broken chromosome is inserted into the body of another, leading to a grossly abnormal chromosome. This is known as an insertion and generally leads to gross anomalies at meiosis.
4. Methods of Study
Sex Chromatin (X-Chromatin) Body (Barr Body)
The X-chromatin body was first seen in the nucleus of the nerve cell of a female cat in 1949 by Barr and Bertram. It has been found to be the constricted, nonfunctioning X chromosome. As a general rule, only 1 X chromosome functions in a cell at a given time. All other X chromosomes present in a cell may be seen as X-chromatin bodies in a resting nucleus. Thus, if one knows the number of X chromosomes, one can anticipate that the number of Barr bodies will be 1 less. If one counts the number of Barr bodies, the number of X chromosomes can be determined by adding 1.
Drumsticks on Polymorphonuclear Leukocytes
Small outpouchings of the lobes of nuclei in polymorphonuclear leukocytes of females have been demonstrated to be the X-chromatin body in this particular cell. Hence, leukocyte preparations may be used to detect X-chromatin bodies in much the same way as buccal cells are used.
Chromosome Count
In the karyotypic analysis of a patient, it is the usual practice to count 20–50 chromosome spreads for chromosome number. The purpose of this practice is to determine whether mosaicism exists because if a mosaic pattern does exist, there will be at least 2 cell lines of different counts. Photographs are made of representative spreads, and karyotypes are constructed so that the morphology of each chromosome can be studied.
Banding Techniques
As previously described, it is possible after appropriate pretreatment to stain metaphase spreads with special stains and construct a karyotype that demonstrates the banding patterns of each chromosome. In this way, it is possible to identify with certainty every chromosome in the karyotype. This is of value with problems such as translocations and trisomic conditions. Another use depends on the fact that most of the long arm of the Y chromosome is heterochromic and stains deeply with fluorescent stains. Thus, the Y chromosome can be identified at a glance, even in the resting nucleus.
APPLIED GENETICS & TERATOLOGY
1. Chromosomes & Spontaneous Abortion
An entirely new approach to reproductive biology problems became available with the advent of tissue culture and cytologic techniques that made it possible to culture cells from any tissue of the body and produce karyotypes that could be analyzed. In the early 1960s, investigators in a number of laboratories began to study chromosomes of spontaneous abortions and demonstrated that the earlier the spontaneous abortion occurred, the more likely it was due to a chromosomal abnormality. It is now known that in spontaneous abortions occurring in the first 8 weeks, the fetuses have about a 50% incidence of chromosome anomalies.
Of abortuses that are abnormal, approximately one-half are trisomic, suggesting an error of meiotic nondisjunction. One-third of abortuses with trisomy have trisomy 16. Although this abnormality does not occur in liveborn infants, it apparently is a frequent problem in abortuses. The karyo-type 45,X occurs in nearly one-fourth of chromosomally abnormal abortuses. This karyotype occurs about 24 times more frequently in abortuses than in liveborn infants, a fact that emphasizes its lethal nature. Over 15% of chromosomally abnormal abortuses have polyploidy (triploidy or tetraploidy). These lethal conditions are seen only in abortuses except in extremely rare circumstances and are due to a variety of accidents, including double fertilization and a number of meiotic errors. Finally, a small number of chromosomally abnormal abortuses have unbalanced translocations and other anomalies.
Recurrent Pregnancy Loss
Couples who experience habitual abortion constitute about 0.5% of the population. The condition is defined as 2 or more spontaneous abortions. Several investigators have studied groups of these couples using banding techniques and have found that 10–25% of them have a chromosome anomaly in either the male or female partner. Those seen are 47,XXX, 47,XYY, and a variety of balanced translocation carriers. Those with sex chromosome abnormalities frequently demonstrate other nondisjunctional events. Chromosome anomalies are thus a major cause of habitual abortion, and the incorporation of genetic evaluation into such a work-up is potentially fruitful.
Lippman-Hand and Bekemans reviewed the world literature and studied the incidence of balanced translocation carriers among 177 couples who had 2 or more spontaneous abortions. These studies suggest that in 2–3% of couples experiencing early fetal loss, 1 partner will have balanced translocations. This percentage is not markedly increased when more than 2 abortions occur. Females had a somewhat higher incidence of balanced translocations than did males.
2. Chromosomal Disorders
This section is devoted to a brief discussion of various autosomal abnormalities. Table 3–5 summarizes some of the autosomal abnormalities that have been diagnosed. They are represented as syndromes, together with some of the signs typical of these conditions. In general, autosomal monosomy is so lethal that total loss of a chromosome is rarely seen in an individual born alive. Only a few cases of monosomy 21–22 have been reported to date, which attests to the rarity of this disorder. Trisomy may occur with any chromosome. The 3 most common trisomic conditions seen in living individuals are trisomies 13, 18, and 21. Trisomy of various C group chromosomes has been reported sporadically. The most frequently reported is trisomy 8. Generally, trisomy of other chromosomes must be assumed to be lethal, because they occur only in abortuses, not in living individuals. To date, trisomy of every autosome except chromosome 1 has been seen in abortuses.
Table 3–5. Autosomal disorders.
Translocations can occur between any 2 chromosomes of the karyotype, and a variety of phenotypic expressions may be seen after mediocre arrangements. Three different translocation patterns have been identified in Down syndrome: 15/21, 21/21, and 21/22.
Deletions may also occur with respect to any chromosome in the karyotype and may be brought about by a translocation followed by a rearrangement in meiosis, which leads to the loss of chromatin material, or by a simple loss of the chromatin material following a chromosome break. Some of the more commonly seen deletion patterns are listed in Table 3–5.
The most frequent abnormality related to a chromosome abnormality is Down syndrome. Down syndrome serves as an interesting model for the discussion of autosomal diseases. The 21 trisomy type is the most common form and is responsible for approximately 95% of Down syndrome patients. There is a positive correlation between the frequency of Down syndrome and maternal age. Babies with Down syndrome are more often born to teenage mothers and even more frequently to mothers over 35. Although the reason for these findings is not entirely clear, it may be that, in older women at least, the egg has been present in prophase of the first meiotic division from the time of fetal life and that, as the egg ages, there is a greater tendency for nondisjunction to occur, leading to trisomy. A second theory is that coital habits are more erratic in both the very young and the older mothers, and this may lead to an increased incidence of fertilization of older eggs. This theory maintains that these eggs may be more likely to suffer nondisjunction or to accept abnormal sperm. Be this as it may, the incidence of Down syndrome in the general population is approximately 1 in 600 deliveries and at age 40 approximately 1 in 100 deliveries. At age 45, the incidence is approximately 1 in 40 deliveries (Table 3–6). The other 5% of Down syndrome patients are the result of translocations, the most common being the 15/21 translocation, but examples of 21/21 and 21/22 have been noted. In the case of 15/21, the chance of recurrence in a later pregnancy is theoretically 25%. In practice, a rate of 10% is observed if the mother is the carrier. When the father is the carrier, the odds are less because there may be a selection not favoring the sperm carrying both the 15/21 translocation and the normal 21 chromosome. In the case of 21/21 translocation, there is no chance for formation of a normal child because the carrier will contribute either both 21s or no 21 and, following fertilization, will produce either a monosomic 21 or trisomic 21. With regard to 21/22 translocation, the chance of producing a baby with Down syndrome is 1 in 2.
Table 3–6. Estimates of rates per thousand of chromosome abnormalities in live births by single-year interval.
In general, other trisomic states occur with greater frequency in older women, and the larger the chromosome involved, the more severe the syndrome. Because trisomy 21 involves the smallest of the chromosomes, the phenotypic problems of Down syndrome are the least severe, and a moderate life expectancy may be anticipated. Even these individuals will be grossly abnormal, however, because of mental retardation and defects in other organ systems. The average life expectancy of patients with Down syndrome is much lower than for the general population.
3. Prenatal Diagnosis
Currently the most common use for applied genetics in obstetrics and gynecology is in prenatal counseling, screening, and diagnosis. Prenatal diagnosis first came into use in 1977 with the discovery of the significance of serum α fetoprotein (AFP). The United Kingdom Collaboration Study found that elevated AFP in maternal serum drawn between 16 and 18 weeks of gestation correlated with an increased incidence of neural tube defects (NTDs). Since that time, much research effort has been aimed at perfecting the technique. We now can screen not only for NTDs but also for trisomy 21 and trisomy 18. In addition, cystic fibrosis, sickle cell disease, and Huntington’s disease, as well as many inborn errors of metabolism and other genetic disorders, can now be identified prenatally.
Neural Tube Disease
Most neural tube diseases, eg, anencephaly, spina bifida, and meningomyelocele, are associated with a multifactorial inheritance pattern. The frequency of their occurrence varies in different populations (eg, rates as high as 10 per 1000 births in Ireland and as low as 0.8 per 1000 births in the western United States). Ninety percent are index cases, ie, they occur spontaneously without previous occurrence in a family. In general, if a couple has a child with such an anomaly, the chance of producing another affected child is 2–5%. If they have had 2 such children, the risk can be as high as 10%. However, other diagnostic possibilities involving different modes of inheritance should be considered. Siblings also run greater risks of having affected children, with the highest risk being to female offspring of sisters and the lowest to male offspring of brothers. Maternal serum screening is now available to all mothers between 16 and 20 weeks of gestation. If an elevation of 2.5 or more standard deviations above the mean is noted, amniocentesis for AFP should be done along with a careful ultrasound study of the fetus for structural anomalies. Evidence for an NTD noted on ultrasound and suspected by amniotic fluid AFP elevation of 3.0 or more standard deviations indicates a diagnosis of an NTD and allows for appropriate counseling and decision making for the parents.
Maternal serum AFP screening detects about 85% of all open NTDs, thus allowing detection of 80% of all open NTDs and 90% of all anencephalic infants. Serum AFP screening does not detect skin-covered lesions or the closed form of NTDs. Thus, most encephaloceles may be missed.
Approximately 5–5.5% of women screened will have abnormally elevated values (≥ 2.5 times the mean). Most of these will be false-positive results (a repeat test should determine this) due to inaccurate dating of gestational age, multiple gestation, fetal demise or dying fetus, or a host of other structural abnormalities. In most cases, repeat AFP testing and ultrasound examination will identify the problem. If the serum AFP level remains elevated and ultrasound examination does not yield a specific diagnosis, amniotic fluid AFP levels should be measured as well as amniotic fluid acetylcholinesterase levels. Further testing and counseling may be necessary before a final diagnosis can be made. When the correct gestational age is used, the false-positive rate for second-trimester maternal screening is 3–4%.
Chromosomal Abnormalities
In 1984, maternal serum AFP levels were found to be lower in patients who delivered infants with Down syndrome. Using the AFP value with maternal age, 25–30% of fetuses with Down syndrome were detected prenatally. In 1988, 2 additional tests were added to the maternal AFP: human chorionic gonadotropin (hCG) and unconjugated estriol (uE3). Using the “triple screen,” a 60% detection rate for Down syndrome was accomplished. In addition, the use of uE3 allowed for detection of trisomy 18.
Fetuses with Down syndrome have low maternal AFP, low uE3, and high hCG. Fetuses with trisomy 18 have low values across all of the serum markers. The false-positive rate for women less than 35 years of age is 5%. Above this age cutoff, the false-positive rate is increased. The definitive diagnosis of a chromosomal abnormality must be confirmed with a fetal karyotype.
The risk of fetal trisomies increases with increasing maternal age. At age 35 the risk of a trisomy is approximately 1 in 200. At age 40 the risk is 1 in 20 (Table 3–6). Prior to the discovery of serum markers, advanced maternal age was used to guide which women received fetal karyotyping. Trisomies, however, are not the only abnormality increased in this population of women. Sex chromosome aneuploidies (47,XXY and 47,XXX) also occur at an increased rate in women 35 years of age and older. Despite the advances in serum screening, fetal karyotyping continues to be the gold standard for prenatal testing in this group of women. The use of maternal serum screening in this subset of women is hindered by a high false-positive rate, less than 100% detection rate for trisomy 18 and 21, and the lack of ability to screen for the sex chromosome aneuploidies.
Cystic Fibrosis
Cystic fibrosis affects 1 in 3300 individuals of European descent in the United States. The carrier frequency is 1 in 29 for North Americans of European descent and Ashkenazi Jewish descent and 1 in 60 for African Americans. A deletion of phenylalanine at position 508 of the CFTR gene on chromosome 7 leads to the disease. All individuals with a family history of cystic fibrosis or a high carrier frequency should be offered carrier testing. For couples who are both carriers of the defective allele, fetal testing may be provided.
Future Advances in Prenatal Screening
In the detection of certain trisomies, the triple-marker screen provides better sensitivity than any single marker alone. Nonetheless, the detection rate for trisomy 18 and trisomy 21 still remains quite low. According to the Serum Urine and Ultrasound Screening Study (SURUSS), integration of nuchal translucency measurement and pregnancy-associated plasma protein-A (PAPP-A) in the first trimester improves screening. This information in conjunction with early second-trimester measurement of AFP, uE3, free β-hCG (or total hCG), and inhibin-A with maternal age provides the most effective method for screening of Down syndrome, with an 85% detection rate and 0.9% false-positive rate. As the field of prenatal diagnostics continues to evolve, higher detection rates with lower false-positive rates can be expected. With continued research and advancing technology, prenatal screening may move into the first trimester. It may involve new markers (proform of eosinophil major basic protein [proMBP], nasal bone) and may even involve markers taken in both the first and second trimesters.
Fetal Karyotyping
A. Amniocentesis
Amniocentesis for prenatal diagnosis of genetic diseases is an extremely useful tool in the following circumstances or classes of patients:
1. Maternal age 35 years or above
2. Previous chromosomally abnormal child
3. Three or more spontaneous abortions
4. Patient or husband with chromosome anomaly
5. Family history of chromosome anomaly
6. Possible female carrier of X-linked disease
7. Metabolic disease risk (because of previous experience or family history)
8. NTD risk (because of previous experience or family history)
9. Positive second-trimester maternal serum screen
Currently, so many metabolic diseases can be diagnosed prenatally by amniocentesis that when the history elicits the possible presence of a metabolic disease, it is prudent to check with a major center to ascertain the availability of a diagnostic method.
Amniocentesis generally is carried out at 15 to 17 weeks of gestation but can be offered earlier (12–14 weeks). The underlying risk of amniocentesis when performed at 15 weeks of gestation and beyond is increased risk of miscarriage. This risk is estimated at 1 in 200 (0.5%), which is approximately the risk of Down syndrome in a 35-year-old woman. When amniocentesis is performed prior to 15 weeks, the miscarriage rate is slightly increased. Table 3–7 lists some of the conditions that now can be diagnosed prenatally by biochemical means.
Table 3–7. Examples of hereditary diseases diagnosable prenatally.
B. Chorionic Villus Sampling
Chorionic villus sampling (CVS) is a technique used in the first trimester to obtain villi for cytogenetic testing. Most commonly, it is performed transcervically; however, transabdominal routes may also be attempted. The value of CVS is that it can be performed earlier in the pregnancy, and thus the decision of pregnancy termination can be made earlier. The downfall of CVS, however, is a slightly higher miscarriage rate of 1–5% and an association with distal limb defects. These risks appear to be dependent on operator experience, and lower numbers have been reported when CVS is performed between 10 and 12 weeks of gestation.
Karyotyping & Fluorescence In Situ Hybridization Analysis
Once the fetal cells are obtained, they must be processed. Formal karyotyping should be performed on all specimens. This involves culturing the cells, replication, and eventually karyotyping. The entire process often takes 10–14 days until the final report becomes available. Fortunately, a quicker analysis can be obtained for some of the most common chromosomal anomalies.
The fluorescence in situ hybridization (FISH) study is a rapid assay for the detection of specific chromosomal aneuploidies using fluorescent-labeled DNA probes. Currently, probes exist for chromosomes 13, 18, 21, and 22, as well as the X and Y sex chromosomes among others. The average time to obtain a result is 24 hours. However, certain chromosomal probes may return as quickly as 4 hours. The more rapid turnaround time can be attained because the probes are mixed with uncultured amniocytes obtained from amniotic fluid or cells from CVS. If a patient is late in gestation or if the ultrasound is highly suggestive of a certain chromosomal composite, FISH analysis may be an appropriate study. With the development of multicolor FISH, all human chromosomes are painted in 24 different colors, allowing identification of chromosome rearrangement.
Single Gene Defects
If 1 parent is affected and the condition is caused by an autosomal dominant disorder, the chances are 1 in 2 that a child will be affected. If both parents are carriers of an autosomal recessive condition, the chances are 1 in 4 that the child will be affected and 1 in 2 that the child will be a carrier. Carrier status of both parents can be assumed if an affected child has been produced or if a carrier testing program is available and such testing determines that both parents are carriers. Tay-Sachs disease and sickle cell disease detection programs are examples of the latter possibility.
When carrier testing is available and the couple is at risk, as with Tay-Sachs disease in Jewish couples and sickle cell disease in blacks, the physician should order these carrier tests before pregnancy is undertaken, or immediately if the patient is already pregnant. When parents are carriers and pregnancy has been diagnosed, prenatal diagnostic testing is indicated if a test is available. If a physician does not know whether or not a test exists or how to obtain the test, the local genetic counseling program, local chapter of the National Foundation/March of Dimes, or state health department should be called for consultation. These sources may be able to inform the physician about new research that may have produced a prenatal test. A new test may be likely because this area of research is very dynamic. If genetic counseling services are readily available, patients with specific problems should be referred to those agencies for consultation. It is impossible for a physician to keep track of all of the current developments in the myriad conditions caused by single gene defects.
X-linked traits are frequently amenable to prenatal diagnostic testing. When such tests are not available, the couple has the option of testing for the sex of the fetus. If a fetus is noted to be a female, the odds are overwhelming that it will not be affected, although a carrier state may be present. If the fetus is a male, the chances are 1 in 2 that it will be affected. With this information, the couple can decide whether or not to continue the pregnancy in the case of a male fetus. Again, checking with genetic counseling agencies may reveal a prenatal diagnostic test that has only recently been described or information such as gene linkage studies that may apply in the individual case.
All options should be presented in a nonjudgmental fashion with no attempt to persuade, based on the best information available at the time. The couple should be encouraged to decide on a course of action that suits their particular needs. If the decision is appropriate, it should be supported by the physician and the genetic counselor. Very rarely, the patient will make a decision the physician regards as unwise or unrealistic. Such a decision may be based on superstition, religious or mystical beliefs, simple naiveté, or even personality disorder. The physician should make every attempt to clarify the issues for the patient. Rarely, other resources such as family members or spiritual leaders may be consulted in strict confidence. The physician and the genetic counselor must clearly set forth the circumstances of the problem in the record, in case the patient undertakes a course of action that ends in tragedy and perhaps attempts to blame the professional counselors for not preventing it.
Genetic Counseling
Genetic counseling involves interaction between the physician, the family, and the genetic counselor. It is the physician’s responsibility to utilize the services of the genetic consultant in the best interest of the patient. The genetic counselor will take a formal family history and construct a family tree (Fig. 3–4). The assessment of the underlying general population risk of a disease and the specific family risk should be provided. When a specific diagnosis is known in the proband and the relatives are dead or otherwise not available, the counselor may ask to see photographs, which may show characteristics of the suspected condition. In many cases, when the pedigree is constructed, the inheritance pattern can be determined. If this can be done, the relative risks that future progeny will be affected can be estimated. This pedigree information is also useful in discussing the case with a genetic counselor.
Figure 3–4. Pedigree showing unaffected offspring, carrier offspring, and affected offspring in a family with an autosomal recessive trait (sickle cell anemia).
GYNECOLOGIC CORRELATES
THE CHROMOSOMAL BASIS OF SEX DETERMINATION
Syngamy
The sex of the fetus normally is determined at fertilization. The cells of normal females contain 2 X chromosomes; those of normal males contain 1 X and 1 Y. During meiotic reduction, half of the male gametes receive a Y chromosome and the other half an X chromosome. Because the female has 2 X chromosomes, all female gametes contain an X chromosome. If a Y-bearing gamete fertilizes an ovum, the fetus is male; conversely, if an X-bearing gamete fertilizes an ovum, the fetus is female.
Arithmetically, the situation described previously should yield a male/female sex ratio of 100—the sex ratio being defined as 100 times the number of males divided by the number of females. However, for many years, the male/female sex ratio of the newborns in the white population has been approximately 105. Apparently the sex ratio at fertilization is even higher than at birth; most data on the sex of abortuses indicate a preponderance of males.
Abnormalities of Meiosis and Mitosis
The discussion in this section is limited to anomalies of meiosis and mitosis that result in some abnormality in the sex chromosome complement of the embryo.
Chromosome studies in connection with various clinical conditions suggest that errors in meiosis and mitosis do indeed occur. These errors result in any of the following principal effects: (1) an extra sex chromosome, (2) an absent sex chromosome, (3) 2 cell lines having different sex chromosomes and arising by mosaicism, (4) 2 cell lines having different sex chromosomes and arising by chimerism, (5) a structurally abnormal sex chromosome, and (6) a sex chromosome complement inconsistent with the phenotype.
By and large, an extra or a missing sex chromosome arises as the result of an error of disjunction in meiosis I or II in either the male or the female. In meiosis I, this means that instead of each of the paired homologous sex chromosomes going to the appropriate daughter cell, both go to 1 cell, leaving that cell with an extra sex chromosome and the daughter cell with none. Failure of disjunction in meiosis II simply means that the centromere fails to divide normally.
A variation of this process, known as anaphase lag, occurs when 1 of the chromosomes is delayed in arriving at the daughter cell and thus is lost. Theoretically, chromosomes may be lost by failure of association in prophase and by failure of replication, but these possibilities have not been demonstrated.
Persons who have been found to have 2 cell lines apparently have experienced problems in mitosis in the very early stage of embryogenesis. Thus, if there is nondisjunction or anaphase lag in an early (first, second, or immediately subsequent) cell division in the embryo, mosaicism may be said to exist. In this condition, there are 2 cell lines; 1 has a normal number of sex chromosomes, and the other is deficient in a sex chromosome or has an extra number of sex chromosomes. A similar situation exists in chimerism, except that there may be a difference in the sex chromosome: 1 may be an X and 1 may be a Y. This apparently arises by dispermy, by the fertilization of a double oocyte, or by the fusion, very early in embryogenesis, of 2 separately fertilized oocytes. Each of these conditions has been produced experimentally in animals.
Structural abnormalities of the sex chromosomes—deletion of the long or short arm or the formation of an isochromosome (2 short arms or 2 long arms)—result from injury to the chromosomes during meiosis. How such injuries occur is not known, but the results are noted more commonly in sex chromosomes than in autosomes—perhaps because serious injury to an autosome is much more likely to be lethal than injury to an X chromosome, and surviving injured X chromosomes would therefore be more common.
The situation in which there is a sex chromosome complement with an inappropriate genotype arises in special circumstances of true hermaphroditism and XX males (see later sections).
The X Chromosome in Humans
At about day 16 of embryonic life, there appears on the undersurface of the nuclear membrane of the somatic cells of human females a structure 1 μm in diameter known as the X-chromatin body. There is genetic as well as cytogenetic evidence that this is 1 of the X chromosomes (the only chromosome visible by ordinary light microscopy during interphase). In a sense, therefore, all females are hemizygous with respect to the X chromosome. However, there are genetic reasons for believing that the X chromosome is not entirely inactivated during the process of formation of the X-chromatin body. In normal females, inactivation of the X chromosome during interphase and its representation as the X-chromatin body are known as the Lyon phenomenon (for Mary Lyon, a British geneticist). This phenomenon may involve, at random, either the maternal or the paternal X chromosome. Furthermore, once the particular chromosome has been selected early in embryogenesis, it is always the same X chromosome that is inactivated in the progeny of that particular cell. Geneticists have found that the ratio of maternal to paternal X chromosomes inactivated is approximately 1:1.
The germ cells of an ovary are an exception to the X inactivation concept in that X inactivation does not characterize the meiotic process. Apparently, meiosis is impossible without 2 genetically active X chromosomes. Although random structural damage to 1 of the X chromosomes seems to cause meiotic arrest, oocyte loss, and therefore failure of ovarian development, an especially critical area necessary for oocyte development has been identified on the long arm of the X. This essential area involves almost all of the long arm and has been specifically located from Xq13 to Xq26. If this area is broken in 1 of the X chromosomes as in a deletion or translocation, oocyte development does not occur. However, a few exceptions to this rule have been described.
It is a curious biologic phenomenon that if 1 of the X chromosomes is abnormal, it is always this chromosome that is genetically inactivated and becomes the X-chromatin body, regardless of whether it is maternal or paternal in origin. Although this general rule seems to be an exception to the randomness of X inactivation, this is more apparent than real. Presumably, random inactivation does occur, but the disadvantaged cells—ie, those left with a damaged active X—do not survive. Consequently, the embryo develops only with cells with a normal active X chromosome (X-chromatin body) (Fig. 3–5).
Figure 3–5. Relation of X-chromatin body to the possible sex chromosome components.
If there are more than 2 X chromosomes, all X chromosomes except 1 are genetically inactivated and become X-chromatin bodies; thus, in this case, the number of X-chromatin bodies will be equal to the number of X chromosomes minus 1. This type of inactivation applies to X chromosomes even when a Y chromosome is present, eg, in Klinefelter’s syndrome.
Although the X chromosomes are primarily concerned with the determination of femininity, there is abundant genetic evidence that loci having to do with traits other than sex determination are present on the X chromosome. Thus, in the catalog of genetic disorders given in the 10th edition of Mendelian Inheritance in Man, 320 traits are listed as more or less definitely X-linked. Substantial evidence for X linkage has been found for about 160 of these traits; the rest are only suspected of having this relationship. Hemophilia, color blindness, childhood muscular dystrophy (Duchenne’s dystrophy), Lesch-Nyhan syndrome, and glucose-6-phosphate dehydrogenase deficiency are among the better known conditions controlled by loci on the X chromosome. These entities probably arise from the expression of a recessive gene due to its hemizygous situation in males.
X-linked dominant traits are infrequent in humans. Vitamin D-resistant rickets is an example.
At least 1 disorder can be classified somewhere between a structural anomaly of the X chromosome and a single gene mutation. X-linked mental retardation in males is associated with a fragile site at q26, but a special culture medium is required for its demonstration. Furthermore, it has been shown that heterozygote female carriers for this fragile site have low IQ test scores.
The Y Chromosome in Humans
Just as the X chromosome is the only chromosome visible by ordinary light microscopy during interphase, the Y chromosome is the only chromosome visible in interphase, after exposure to quinacrine compounds, by fluorescence microscopy. This is a very useful diagnostic method.
In contrast to the X chromosome, few traits have been traced to the Y chromosome except those having to do with testicular formation and those at the very tip of the short arm, homologous with those at the tip of the short arm of the X. Possession of the Y chromosome alone, ie, without an X chromosome, apparently is lethal, because such a case has never been described.
Present on the Y chromosome is an area that produces a factor that allows for testicular development. This factor is termed testis-determining factor (TDF). Without the presence of TDF, normal female anatomy will develop. When TDF is present, testicular development occurs with subsequent differentiation of Sertoli cells. The Sertoli cells in turn produce a second factor central to male differentiation, müllerian-inhibiting factor (MIF), also termed antimüllerian factor (AMF). The presence of MIF causes the regression of the müllerian ducts and thereby allows for the development of normal internal male anatomy.
Y-Chromosome Microdeletion
In addition to sex determination function, human Y chromosome also has a role in spermatogenesis controlled by multiple genes along proximal Yq. The locus for spermatogenesis is on the euchromatic part of Yq (Yq11) called azoospermic factor (AZF). The AZF region is divided into three nonoverlapping regions AZFa, AZFb, and AZFc. The term “microdeletion” means that the size of the deleted segment is not visualized on karyotyping but must be discerned through molecular biology technique. There is no specific phenotype–genotype correlation between the degree of spermatogenic failure and type of Yq microdeletion. Complete deletion of AZFa and AZFb regions is associated with Sertoli cell-only syndrome and spermatogenic arrest, respectively. However, partial deletions of AZFa or AZFb or complete/partial deletions of AZFc are associated with a variable degree of spermatogenic failure ranging from oligozoospermia to Sertoli cell-only syndrome. There are reports of progressive impairment of spermatogenesis over time in patient with AZFc deletion. The fourth AZFd region, which was earlier proposed, does not exist based on the Y chromosome sequencing. There are many candidate genes within the deleted regions that are responsible for impaired spermatogenesis. The extensively studied genes are DAZ on AZFc region, RBMY1A1 on AZFb region, and USP9Y, DBY, and UTY on AZFa region. Because the deleted genes are expressed mainly in testes, men carrying the deletions have no abnormalities other than spermatogenic failure.
The incidence of Yq microdeletions in infertile men varies from 1–55% depending on study design. The most frequently deleted region is AZFc (~60%), whereas the deletion of the AZFa region is extremely rare (5%). The identification of Yq microdeletion has a prognostic value for the chance of successful testicular sperm retrieval. Men with complete deletion of AZFa and AZFb regions have almost no chance of having sperm recovered from surgical testicular sperm retrieval procedure, and no treatment is presently available for their fertility problem besides the use of donor sperm.
In the past, the majority of cases of Yq microdeletions have been de novo in infertile men during embryogenesis or from meiotic error in the germline of the fertile father. However, with the advent of assisted reproductive technologies, these infertile men can conceive genetic offspring with intracytoplasmic sperm injection (ICSI) technique, so Yq microdeletion can pass from generation to generation. A few studies show that when a Yq microdeletion is present in infertile men, ICSI-derived sons will inherit the same deletion. In view of genetic counseling, although Yq microdeletion is transmitted to the male offspring, the phenotype of male offspring regarding the degree of spermatogenesis is unpredictable due to the influence of the presence or absence of environmental factors that could affect spermatogenesis and the period of lifetime when spermatogenesis is assessed.
ABNORMAL DEVELOPMENT
1. Ovarian Agenesis–Dysgenesis
In 1938, Turner described 7 girls 15–23 years of age with sexual infantilism, webbing of the neck, cubitus valgus, and retardation of growth. A survey of the literature indicates that “Turner’s syndrome” means different things to different writers. After the later discovery that ovarian streaks are characteristically associated with the clinical entity described by Turner, “ovarian agenesis” became a synonym for Turner’s syndrome. After discovery of the absence of the X-chromatin body in such patients, the term ovarian agenesis gave way to “gonadal dysgenesis,” “gonadal agenesis,” or “gonadal aplasia.”
Meanwhile, some patients with the genital characteristics mentioned previously were shown to have a normally positive X-chromatin count. Furthermore, a variety of sex chromosome complements have been found in connection with streak gonads. As if these contradictions were not perplexing enough, it has been noted that streaks are by no means confined to patients with Turner’s original tetrad of infantilism, webbing of the neck, cubitus valgus, and retardation of growth but may be present in girls with sexual infantilism only. Since Turner’s original description, a host of additional somatic anomalies (varying in frequency) have been associated with his original clinical picture; these include shield chest, overweight, high palate, micrognathia, epicanthal folds, low-set ears, hypoplasia of nails, osteoporosis, pigmented moles, hypertension, lymphedema, cutis laxa, keloids, coarctation of the aorta, mental retardation, intestinal telangiectasia, and deafness.
For our purposes, the eponym Turner’s syndrome will be used to indicate sexual infantilism with ovarian streaks, short stature, and 2 or more of the somatic anomalies mentioned earlier. In this context, terms such as ovarian agenesis, gonadal agenesis, and gonadal dysgenesis lose their clinical significance and become merely descriptions of the gonadal development of the person. At least 21 sex chromosome complements have been associated with streak gonads (Fig. 3–6), but only about 9 sex chromosome complements have been associated with Turner’s syndrome. However, approximately two-thirds of patients with Turner’s syndrome have a 45,X chromosome complement, whereas only one-fourth of patients without Turner’s syndrome but with streak ovaries have a 45,X chromosome complement.
Figure 3–6. The 21 sex chromosome complements that have been found in patients with streak gonads.
Karyotype–phenotype correlations in the syndromes associated with ovarian agenesis are not completely satisfactory. Nonetheless, if gonadal development is considered as 1 problem and if the somatic difficulties associated with these syndromes are considered as a separate problem, one can make certain correlations.
With respect to failure of gonadal development, it is important to recall that diploid germ cells require 2 normal active X chromosomes. This is in contrast to the somatic cells, where only 1 sex chromosome is thought to be genetically active, at least after day 16 of embryonic life in the human, when the X-chromatin body first appears in the somatic cells. It is also important to recall that in 45,X persons no oocytes persist, and streak gonads are the rule. From these facts, it can be inferred that failure of gonadal development is not the result of a specific sex chromosome defect but rather of the absence of 2 X chromosomes with the necessary critical zones.
Karyotype–phenotype correlations with respect to somatic abnormalities are even sketchier than the correlations with regard to gonadal development. However, good evidence shows that monosomy for the short arm of the X chromosome is related to somatic difficulties, although some patients with long-arm deletions have somatic abnormalities.
History of Gonadal Agenesis
The histologic findings in these abnormal ovaries in patients with gonadal streaks are essentially the same regardless of the patient’s cytogenetic background (Fig. 3–7).
Figure 3–7. Gonadal streaks in a patient with the phenotype of Turner’s syndrome. (Redrawn and reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
Fibrous tissue is the major component of the streak. It is indistinguishable microscopically from that of the normal ovarian stroma. The so-called germinal epithelium, on the surface of the structure, is a layer of low cuboid cells; this layer appears to be completely inactive.
Tubules of the ovarian rete are invariably found in sections taken from about the midportion of the streak.
In all patients who have reached the age of normal puberty, hilar cells are also demonstrated. The number of hilar cells varies among patients. In those with some enlargement of the clitoris, hilar cells are present in large numbers. These developments may be causally related. Nevertheless, hilar cells are also found in many normal ovaries. The origin of hilar cells is not precisely known, but they are associated with development of the medullary portion of the gonad. Their presence lends further support to the concept that in ovarian agenesis the gonad develops along normal lines until just before the expected appearance of early oocytes. In all cases in which sections of the broad ligament have been available for study, it has been possible to identify the mesonephric duct and tubules—broad ligament structures found in normal females.
Clinical Findings
A. Symptoms & Signs
1. In newborn infants—The newborn with streak ovaries often shows edema of the hands and feet. Histologically, this edema is associated with large dilated vascular spaces. With such findings, it is obviously desirable to obtain a karyotype. However, some children with streak ovaries—particularly those who have few or no somatic abnormalities—cannot be recognized at birth.
2. In adolescents—The arresting and characteristic clinical finding in many of these patients is their short stature. Typical patients seldom attain a height of 1.5 m (5 ft) (Fig. 3–8). In addition, sexual infantilism is a striking finding. As mentioned earlier, a variety of somatic abnormalities may be present; by definition, if 2 or more of these are noted, the patient may be considered to have Turner’s syndrome. Most of these patients have only 1 normal X chromosome, and two-thirds of them have no other sex chromosome. Patients of normal height without somatic abnormalities may also have gonadal streaks. Under these circumstances, there is likely to be a cell line with 2 normal sex chromosomes but often a second line with a single X. The internal findings are exactly the same as in patients with classic Turner’s syndrome, however.
Figure 3–8. Patient with Turner’s syndrome. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
B. Laboratory Findings
An important finding in patients of any age—but especially after expected puberty, ie, about 12 years—is elevation of total gonadotropin production. From a practical point of view, ovarian failure in patients over age 15 cannot be considered a diagnostic possibility unless the serum follicle-stimulating hormone level is more than 50 mIU/mL and luteinizing hormone level is more than 90 mIU/mL.
Nongonadal endocrine functions are normal. Urinary excretion of estrogens is low, and the maturation index and other vaginal smear indices are shifted well to the left.
Treatment
Substitution therapy with estrogen is necessary for development of secondary characteristics.
Therapy with growth hormone will increase height. Whether ultimate height will be greater than it otherwise would be is uncertain, but current evidence suggests that it will be.
The incidence of malignant degeneration is increased in the gonadal streaks of patients with a Y chromosome, as compared with normal males. Surgical removal of streaks from all patients with a Y chromosome is recommended.
2. True Hermaphroditism
By classic definition, true hermaphroditism exists when both ovarian and testicular tissue can be demonstrated in 1 patient. In humans, the Y chromosome carries genetic material that normally is responsible for testicular development; this material is active even when multiple X chromosomes are present. Thus, in Klinefelter’s syndrome, a testis develops with up to 4 Xs and only 1 Y. Conversely (with rare exceptions), a testis has not been observed to develop in the absence of the Y chromosome. The exceptions are found in true hermaphrodites and XX males, in whom testicular tissue has developed in association with an XX sex chromosome complement.
Clinical Findings
A. Symptoms & Signs
No exclusive features clinically distinguish true hermaphroditism from other forms of intersexuality. Hence, the diagnosis must be entertained in an infant with any form of intersexuality, except only those with a continuing virilizing influence, eg, congenital adrenal hyperplasia. Firm diagnosis is possible after the onset of puberty, when certain clinical features become evident, but the diagnosis can and should be made in infancy.
In the past, most true hermaphrodites have been reared as males because they have rather masculine-appearing external genitalia (Fig. 3–9). Nevertheless, with early diagnosis, most should be reared as females.
Figure 3–9. External genitalia of a patient with true hermaphroditism. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
Almost all true hermaphrodites develop female-type breasts. This helps to distinguish male hermaphroditism from true hermaphroditism, because few male hermaphrodites other than those with familial feminizing hermaphroditism develop large breasts.
Many true hermaphrodites menstruate. The presence or absence of menstruation is partially determined by the development of the uterus; many true hermaphrodites have rudimentary or no development of the müllerian ducts (Fig. 3–10).
Figure 3–10. Internal genitalia of a patient with true hermaphroditism. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
A few patients who had a uterus and menstruated after removal of testicular tissue have become pregnant and delivered normal children.
B. Sex Chromosome Complements
Most true hermaphrodites have X-chromatin bodies and karyotypes that are indistinguishable from those of normal females. In contrast to these, a few patients who cannot be distinguished clinically from other true hermaphrodites have been reported to have a variety of other karyotypes—eg, several chimeric persons with karyotypes of 46,XX/46,XY have been identified.
In true hermaphrodites, the testis is competent in its müllerian-suppressive functions, but an ovotestis may behave as an ovary insofar as its müllerian-suppressive function is concerned. The true hermaphroditic testis or ovotestis is as competent to masculinize the external genitalia as is the testis of a patient with the virilizing type of male hermaphroditism. This is unrelated to karyotype.
Deletion mapping by DNA hybridization has shown that most (but not all) XX true hermaphrodites have Y-specific sequences. Abnormal crossover of a portion of the Y chromosome to the X in meiosis may explain some cases. This latter statement is further supported by the finding of a positive H-Y antigen assay in some patients with 46,XX true hermaphroditism.
In general, the clinical picture of true hermaphroditism is not compatible with the clinical picture in other kinds of gross chromosomal anomalies. For example, very few true hermaphrodites have associated somatic anomalies, and mental retardation almost never occurs.
Treatment
The principles of treatment of true hermaphroditism do not differ from those of the treatment of hermaphroditism in general. Therapy can be summarized by stating that surgical removal of contradictory organs is indicated, and the external genitalia should be reconstructed in keeping with the sex of rearing. The special problem in this group is how to establish with certainty the character of the gonad. This is particularly difficult in the presence of an ovotestis, because its recognition by gross characteristics is notoriously inaccurate, and one must not remove too much of the gonad for study. In some instances, the gonadal tissue of 1 sex is completely embedded within a gonadal structure primarily of the opposite sex.
3. Klinefelter’s Syndrome
This condition, first described in 1942 by Klinefelter, Reifenstein, and Albright, occurs only in apparent males. As originally described, it is characterized by small testes, azoospermia, gynecomastia, relatively normal external genitalia, and otherwise average somatic development. High levels of gonadotropin in urine or serum are characteristic.
Clinical Findings
A. Symptoms & Signs
By definition, this syndrome applies only to persons reared as males. The disease is not recognizable before puberty except by routine screening of newborn infants. Most patients come under observation at 16–40 years of age.
Somatic development during infancy and childhood may be normal. Growth and muscular development may also be within normal limits. Most patients have a normal general appearance and no complaints referable to this abnormality, which is often discovered during the course of a routine physical examination or an infertility study.
In the original publication by Klinefelter and coworkers, gynecomastia was considered an essential part of the syndrome. Since then, however, cases without gynecomastia have been reported.
The external genitalia are perfectly formed and in most patients are quite well developed. Erection and intercourse usually are satisfactory.
There is no history of delayed descent of the testes in typical cases, and the testes are in the scrotum. Neither is there any history of testicular trauma or disease. Although a history of mumps orchitis is occasionally elicited, this disease has not been correlated with the syndrome. However, the testes are often very small in contrast to the rest of the genitalia (about 1.5 × 1.5 cm).
Psychological symptoms are often present. Most studies of this syndrome have been performed in psychiatric institutions. The seriousness of the psychological disturbance seems to be partly related to the number of extra X chromosomes—eg, it is estimated that about one-fourth of XXY patients have some degree of mental retardation.
B. Laboratory Findings
One of the extremely important clinical features of Klinefelter’s syndrome is the excessive amount of pituitary gonadotropin found in either urine or serum assay.
The urinary excretion of neutral 17-ketosteroids varies from relatively normal to definitely subnormal levels. There is a rough correlation between the degree of hypoleydigism as judged clinically and a low 17-ketosteroid excretion rate.
C. Histologic & Cytogenetic Findings
Klinefelter’s syndrome may be regarded as a form of primary testicular failure.
Several authors have classified a variety of forms of testicular atrophy as subtypes of Klinefelter’s syndrome. Be this as it may, Klinefelter believed that only those patients who have a chromosomal abnormality could be said to have this syndrome. Microscopic examination of the adult testis shows that the seminiferous tubules lack epithelium and are shrunken and hyalinized. They contain large amounts of elastic fibers, and Leydig cells are present in large numbers.
Males with positive X-chromatin bodies are likely to have Klinefelter’s syndrome. The nuclear sex anomaly reflects a basic genetic abnormality in sex chromosome constitution. All cases studied have had at least 2 X chromosomes and 1 Y chromosome. The most common abnormality in the sex chromosome constitution is XXY, but the literature also records XXXY, XXYY, XXXXY, and XXXYY, and mosaics of XX/XXY, XY/XXY, XY/XXXY, and XXXY/XXXXY. In all examples except the XX/XXY mosaic, a Y chromosome is present in all cells. From these patterns, it is obvious that the Y chromosome has a very strong testis-forming impulse, which can operate in spite of the presence of as many as 4 X chromosomes.
Thus, patients with Klinefelter’s syndrome will have not only a positive X-chromatin body but also a positive Y-chromatin body.
The abnormal sex chromosome constitution causes differentiation of an abnormal testis, leading to testicular failure in adulthood. At birth or before puberty, such testes show a marked deficiency or absence of germinal cells.
By means of nursery screening, the frequency of males with positive X-chromatin bodies has been estimated to be 2.65 per 1000 live male births.
Treatment
There is no treatment for the 2 principal complaints of these patients: infertility and gynecomastia. No pituitary preparation has been effective in the regeneration of the hyalinized tubular epithelium or the stimulation of gametogenesis. Furthermore, no hormone regimen is effective in treating the breast hypertrophy. When the breasts are a formidable psychological problem, surgical removal may be a satisfactory procedure. In patients who have clinical symptoms of hypoleydigism, substitution therapy with testosterone is an important physiologic and psychological aid. Donor sperm may be offered for treatment of infertility.
4. Double-X Males
A few cases of adult males with a slightly hypoplastic penis and very small testes but no other indication of abnormal sexual development have been reported. These males are sterile. Unlike those with Klinefelter’s syndrome, they do not have abnormal breast development. They are clinically very similar to patients with Del Castillo’s syndrome (testicular dysgenesis). Nevertheless, the XX males have a positive sex chromatin and a normal female karyotype. These may be extreme examples of the sex reversal that usually is partial in true hermaphroditism.
5. Multiple-X Syndromes
The finding of more than 1 X-chromatin body in a cell indicates the presence of more than 2 X chromosomes in that particular cell. In many patients, such a finding is associated with mosaicism, and the clinical picture is controlled by this fact—eg, if 1 of the strains of the mosaicism is 45,X, gonadal agenesis is likely to occur. There also are persons who do not seem to have mosaicism but do have an abnormal number of X chromosomes in all cells. In such persons, the most common complement is XXX (triplo-X syndrome), but XXXX (tetra-X syndrome) and XXXXX (penta-X syndrome) have been reported.
An additional X chromosome does not seem to have a consistent effect on sexual differentiation. The body proportions of these persons are normal, and the external genitalia are normally female. A number of such persons have been examined at laparotomy, and no consistent abnormality of the ovary has been found. In a few cases, the number of follicles appeared to be reduced, and in at least 1 case the ovaries were very small and the ovarian stroma poorly differentiated. About 20% of postpubertal patients with the triplo-X syndrome report various degrees of amenorrhea or some irregularity in menstruation. For the most part, however, these patients have a normal menstrual history and are of proved fertility.
Almost all patients known to have multiple-X syndromes have some degree of mental retardation. A few have mongoloid features. (The mothers of these patients tended to be older than the mothers of normal children, as is true with Down syndrome.) Perhaps these findings are in part circumstantial, as most of these patients were discovered during surveys in mental institutions. The important clinical point is that mentally retarded infants should have chromosomal study.
Uniformly, the offspring of triplo-X mothers have been normal. This is surprising, because theoretically in such cases meiosis should produce equal numbers of ova containing 1 or 2 X chromosomes, and fertilization of the abnormal XX ova should give rise to XXX and XXY individuals. Nevertheless, the triplo-X condition seems selective for normal ova and zygotes.
The diagnosis of this syndrome is made by identifying a high percentage of cells with double X-chromatin bodies in the buccal smear and by finding 47 chromosomes with a karyotype showing an extra X chromosome in all cells cultured from the peripheral blood. It should be noted that in the examination of the buccal smear, some cells have a single X-chromatin body. Hence, based on the chromatin examination, one might suspect XX/XXX mosaicism. Actually, in triplo-X patients, only a single type of cell can be demonstrated in cultures of cells from the peripheral blood. The absence of the second X-chromatin body in some of the somatic cells may result from the time of examination of the cell (during interphase) and from the spatial orientation, which could have prevented visualization of the 2 X-chromatin bodies (adjacent to the nuclear membrane). In this syndrome, the number of cells containing either 1 or 2 X-chromatin bodies is very high—at least 60–80%, as compared with an upper limit of about 40% in normal females.
6. Female Hermaphroditism due to Congenital Adrenal Hyperplasia
ESSENTIALS OF DIAGNOSIS
Female pseudohermaphroditism, ambiguous genitalia with clitoral hypertrophy, and, occasionally, persistent urogenital sinus.
Early appearance of sexual hair; hirsutism, dwarfism.
Urinary 17-ketosteroids elevated; pregnanetriol may be increased.
Elevated serum 17-hydroxyprogesterone level.
Occasionally associated with water and electrolyte imbalance—particularly in the neonatal period.
General Considerations
Female hermaphroditism due to congenital adrenal hyperplasia is a clearly delineated clinical syndrome. The syndrome has been better understood since the discovery that cortisone may successfully arrest virilization. The problem usually is due to a deficiency of a gene required for 21-hydroxylation in the biosynthesis of cortisol.
If the diagnosis is not made in infancy, an unfortunate series of events ensues. Because the adrenals secrete an abnormally large amount of virilizing steroid even during embryonic life, these infants are born with abnormal genitalia (Fig. 3–11). In extreme cases, there is fusion of the scrotolabial folds and, in rare instances, even formation of a penile urethra. The clitoris is greatly enlarged so that it may be mistaken for a penis (Fig. 3–12). No gonads are palpable within the fused scrotolabial folds, and their absence has sometimes given rise to the mistaken impression of male cryptorchidism. Usually, there is a single urinary meatus at the base of the phallus, and the vagina enters the persistent urogenital sinus as noted in Figure 3–13.
Figure 3–11. External genitalia of a female patient with congenital virilizing adrenal hyperplasia. Compare with Figure 3–12. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
Figure 3–12. External genitalia of a female patient with congenital virilizing adrenal hyperplasia. This is a more severe deformity than that shown in Figure 3–11.
Figure 3–13. Sagittal view of genital deformities of increasing severity (A–E) in congenital virilizing adrenal hyperplasia. (Redrawn and reproduced, with permission, from Verkauf BS, Jones HW Jr. Masculinization of the female genitalia in congenital adrenal hyperplasia. South Med J 1970;63:634–638.)
During infancy, provided there are no serious electrolyte disturbances, these children grow more rapidly than normal. For a time, they greatly exceed the average in both height and weight. Unfortunately, epiphyseal closure occurs by about age 10, and as a result, these people are much shorter than normal as adults (Fig. 3–14).
Figure 3–14. Untreated adult with virilizing adrenal hyperplasia. Note the short stature and the relative shortness of the limbs. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
The process of virilization begins at an early age. Pubic hair may appear as early as age 2 years but usually somewhat later. This is followed by growth of axillary hair and finally by the appearance of body hair and a beard, which may be so thick as to require daily shaving. Acne may develop early. Puberty never ensues. There is no breast development. Menstruation does not occur. During the entire process, serum adrenal androgens and 17-hydroxyprogesterone levels are abnormally high.
Although our principal concern here is with this abnormality in females, it must be mentioned that adrenal hyperplasia of the adrenogenital type may also occur in males, in whom it is called macrogenitosomia precox. Sexual development progresses rapidly, and the sex organs attain adult size at an early age. Just as in the female, sexual hair and acne develop unusually early, and the voice becomes deep. The testes are usually in the scrotum; however, in early childhood they remain small and immature, although the genitalia are of adult dimensions. In adulthood, the testes usually enlarge and spermatogenesis occurs, allowing impregnation rates similar to those of a control population. Somatic development in the male corresponds to that of the female; as a child, the male exceeds the average in height and strength, but (if untreated) as an adult he is stocky, muscular, and well below average height.
Both the male and the female with this disorder—but especially the male—may have the complicating problem of electrolyte imbalance. In infancy, it is manifested by vomiting, progressive weight loss, and dehydration and may be fatal unless recognized promptly. The characteristic findings are an exceedingly low serum sodium level, low CO2-combining power level, and high potassium level. The condition is sometimes misdiagnosed as congenital pyloric stenosis.
A few of these patients have a deficiency in 11-hydroxylation that is associated with hypertension in addition to virilization.
Adrenal Histology
The adrenal changes center on a reticular hyperplasia, which becomes more marked as the patient grows older. In some instances, the glomerulosa may participate in the hyperplasia, but the fasciculata is greatly diminished in amount or entirely absent. Lipid studies show absence of fascicular and glomerular lipid but an abnormally strong lipid reaction in the reticularis (Fig. 3–15).
Figure 3–15. Normal adrenal architecture and adrenal histology in congenital virilizing adrenal hyperplasia. Note the great relative increase in the zona reticularis.
Ovarian Histology
The ovarian changes can be summarized by stating that in infants, children, and teenagers, there is normal follicular development to the antrum stage but no evidence of ovulation. With increasing age, less and less follicular activity occurs, and primordial follicles disappear. This disappearance must not be complete, however, because cortisone therapy, even in adults, usually results in ovulatory menstruation after 4–6 months of treatment.
Developmental Anomalies of the Genital Tubercle & Urogenital Sinus Derivatives
The phallus is composed of 2 lateral corpora cavernosa, but the corpus spongiosum is normally absent. The external urinary meatus is most often located at the base of the phallus (Fig. 3–11). An occasional case may be seen in which the urethra does extend to the end of the clitoris (Fig. 3–12). The glans penis and the prepuce are present and indistinguishable from these structures in the male. The scrotolabial folds are characteristically fused in the midline, giving a scrotum-like appearance with a median perineal raphe; however, they seldom enlarge to normal scrotal size. No gonads are palpable within the scrotolabial folds. When the anomaly is not severe (eg, in patients with postnatal virilization), fusion of the scrotolabial folds is not complete, and by gentle retraction it is often possible to locate not only the normally located external urinary meatus but also the orifice of the vagina.
An occasional patient has no communication between the urogenital sinus and the vagina. In no case does the vagina communicate with that portion of the urogenital sinus that gives rise to the female urethra or the prostatic urethra. Instead, the vaginal communication is via caudal urogenital sinus derivatives; thus, fortunately, the sphincter mechanism is not involved, and the anomalous communication is with that portion of the sinus that develops as the vaginal vestibule in the female and the membranous urethra in the male. From the gynecologist’s point of view, it is much more meaningful to say that the vagina and (female) urethra enter a persistent urogenital sinus than to say that the vagina enters the (membranous [male]) urethra. This conclusion casts some doubt on the embryologic significance of the prostatic utricle, which is commonly said to represent the homologue of the vagina in the normal male.
Hormone Changes
Important and specific endocrine changes occur in congenital adrenal hyperplasia of the adrenogenital type. The ultimate diagnosis depends on demonstration of these abnormalities.
A. Urinary Estrogens
The progressive virilization of female hermaphrodites caused by adrenal hyperplasia would suggest that estrogen secretion in these patients is low, and this hypothesis is further supported by the atrophic condition of both the ovarian follicular apparatus and the estrogen target organs. Actually, the determination of urinary estrogens, both fluorometrically and biologically, indicates that it is elevated.
B. Serum Steroids
The development of satisfactory radioimmunoassay techniques for measuring steroids in blood serum has resulted in an increased tendency to measure serum steroids rather than urinary metabolites in diagnosing the condition and monitoring therapy. Serum steroid profiles of many patients with this disorder show that numerous defects in the biosynthesis of cortisol may occur. The most common defect is at the 21-hydroxylase step. Less frequent defects are at the 11-hydroxylase step and the 3β-ol-dehydrogenase step. Rarely, the defect is at the 17-hydroxylase step. In the most common form of the disorder—21-hydroxylase deficiency—the serum 17-hydroxyprogesterone level and, to a lesser extent, the serum progesterone level are elevated. This is easily understandable when it is recalled that 17-hydroxyprogesterone is the substrate for the 21-hydroxylation step (Fig. 3–16). Likewise, in the other enzyme defects, the levels of serum steroid substrates are greatly elevated.
Figure 3–16. Enzymatic steps in cortisol synthesis. Localization of defects in congenital adrenal hyperplasia.
Pathogenesis of Virilizing Adrenal Hyperplasia
The basic defects in congenital virilizing adrenal hyperplasia are 1 or more enzyme deficiencies in the biosynthesis of cortisol (Fig. 3–16). With the reduced production of cortisol, normal feedback to the hypothalamus fails, with the result that increased amounts of adrenocorticotropic hormone (ACTH) are produced. This excess production of ACTH stimulates the deficient adrenal gland to produce relatively normal amounts of cortisol—but also stimulates production of abnormally large amounts of estrogen and androgens by the zona reticularis. In this overproduction, a biologic preponderance of androgens causes virilization. These abnormal sex steroids suppress the gonadotropins so that untreated patients never reach puberty and do not menstruate.
Therefore, the treatment of this disorder consists in part of the administration of sufficient exogenous cortisol to suppress ACTH production to normal levels. This in turn should reduce overstimulation of the adrenal so that the adrenal will cease to produce abnormally large amounts of estrogen and androgen. The gonadotropins generally return to normal levels, with consequent feminization of the patient and achievement of menstruation.
The pathogenesis of the salt-losing type of adrenal hyperplasia involves a deficiency in aldosterone production.
Diagnosis
Hermaphroditism due to congenital adrenal hyperplasia must be suspected in any infant born with ambiguous or abnormal external genitalia. It is exceedingly important that the diagnosis be made at a very early age if undesirable disturbances of metabolism are to be prevented.
All patients with ambiguous external genitalia should have an appraisal of their chromosomal characteristics. In all instances of female pseudohermaphroditism due to congenital hyperplasia, the chromosomal composition is that of a normal female. A pelvic ultrasound in the newborn to determine the presence of a uterus is very helpful and, if positive, strongly suggests a female infant.
The critical determinations are those of the urinary 17-ketosteroid and serum 17-hydroxyprogesterone levels. If these are elevated, the diagnosis must be either congenital adrenal hyperplasia or tumor. In the newborn, the latter is very rare, but in older children and adults with elevated 17-ketosteroids, the possibility of tumor must be considered. One of the most satisfactory methods of making this different diagnosis is to attempt to suppress the excess androgens by administration of dexamethasone. In an adult or an older child, a suitable test dose of dexamethasone is 1.25 mg/45 kg (100 lb) body weight, given orally for 7 consecutive days. In congenital adrenal hyperplasia, there should be suppression of the urinary 17-ketosteroids on the seventh day of the test to less than 1 mg/24 h; in the presence of tumor, either there will be no effect or the 17-ketosteroid levels will rise.
Determination of urinary dehydroepiandrosterone (DHEA) or serum dehydroepiandrosterone sulfate (DHEAS) levels can also be helpful in differentiating congenital adrenal hyperplasia from an adrenal tumor. Levels in patients with congenital adrenal hyperplasia may be up to double the normal amount, whereas an adrenal tumor is usually associated with levels that are much higher than double the normal level.
Determination of the serum sodium and potassium levels and CO2-combining power is also important to ascertain whether electrolyte balance is seriously disturbed.
Treatment
The treatment of female hermaphroditism due to congenital adrenal hyperplasia is partly medical and partly surgical. Originally, cortisone was administered; today, it is known that various cortisone derivatives are at least as effective. It is most satisfactory to begin treatment with relatively large doses of hydrocortisone divided in 3 doses orally for 7–10 days to obtain rapid suppression of adrenal activity. In young infants, the initial dose is about 25 mg/d; in older patients, 100 mg/d. After the output of 17-ketosteroids has decreased to a lower level, the dose should be reduced to the minimum amount required to maintain adequate suppression. This requires repeated measurements of plasma 17α-hydroxyprogesterone in order to individualize the dose.
It has been found that even with suppression of the urinary 17-ketosteroids to normal levels, the more sensitive serum 17-hydroxyprogesterone level may still be elevated. It seems difficult and perhaps undesirable to suppress the serum 17-hydroxyprogesterone values to normal because to do so may require doses of hydrocortisone that tend to cause cushingoid symptoms.
In the treatment of newborns with congenital adrenal hyperplasia who have a defect of electrolyte regulation, it is usually necessary to administer sodium chloride in amounts of 4–6 g/d, either orally or parenterally, in addition to cortisone. Furthermore, fludrocortisone acetate usually is required initially. The dose is entirely dependent on the levels of the serum electrolytes, which must be followed serially, but it is generally 0.05–0.1 mg/d.
In addition to the hormone treatment of this disorder, surgical correction of the external genitalia is usually necessary.
During acute illness or other stress, as well as during and after an operation, additional hydrocortisone is indicated to avoid the adrenal insufficiency of stress. Doubling the maintenance dose is usually adequate in such circumstances.
7. Female Hermaphroditism without Progressive Masculinization
Females with no adrenal abnormality may have fetal masculinization of the external genitalia with the same anatomic findings as in patients with congenital virilizing adrenal hyperplasia. Unlike patients with adrenogenital syndrome, patients without adrenal abnormality do not have elevated levels of serum steroids or urinary 17-ketosteroids, nor do they show precocious sexual development or the metabolic difficulties associated with adrenal hyperplasia as they grow older. At onset of puberty, normal feminization with menstruation and ovulation may be expected.
The diagnosis of female hermaphroditism not due to adrenal abnormality depends on the demonstration of a 46,XX karyotype and the finding of normal levels of serum steroids or normal levels of 17-ketosteroids in the urine. If fusion of the scrotolabial folds is complete, it is necessary to determine the exact relationship of the urogenital sinus to the urethra and vagina and to demonstrate the presence of a uterus by rectal examination or ultrasonography or endoscopic observation of the cervix. When there is a high degree of masculinization, the differential diagnosis between this condition and true hermaphroditism may be very difficult; an exploratory laparotomy may be required in some cases.
Classification
Patients with this problem may be seen because of a variety of conditions.
1. Exogenous androgen:
a. Maternal ingestion of androgen
b. Maternal androgenic tumor
c. Luteoma of pregnancy
d. Adrenal androgenic tumor
2. Idiopathic: No identifiable cause.
3. Special or nonspecific: The same as condition 2 except that it is associated with various somatic anomalies and with mental retardation.
4. Familial: A very rare anomaly.
8. Male Hermaphroditism
Persons with abnormal or ectopic testes may have external genitalia so ambiguous at birth that the true sex is not identifiable (Fig. 3–17). At puberty, these persons tend to become masculinized or feminized depending on factors to be discussed. Thus, the adult habitus of these persons may be typically male, ie, without breasts, or typically female, with good breast development. In some instances, the external genitalia may be indistinguishable from those of a normal female; in others, the clitoris may be enlarged; and in still other instances, there may be fusion of the labia in the midline, resulting in what seems to be a hypospadiac male. A deep or shallow vagina may be present. A cervix, a uterus, and uterine tubes may be developed to varying degrees; however, müllerian structures are often absent. Mesonephric structures may be grossly or microscopically visible. Body hair may be either typically feminine in its distribution and quantity or masculine in distribution and of sufficient quantity as to require plucking or shaving if the person is reared as a female. In a special group, axillary and pubic hair is congenitally absent. Although there is a well-developed uterus in some instances, all patients so far reported have been amenorrheic—in spite of the interesting theoretic possibility of uterine bleeding from endometrium stimulated by estrogen of testicular origin. There is no evidence of adrenal malfunction. In the feminized group, and less frequently in the nonfeminized group, there is a strong familial history of the disorder. Male hermaphrodites reared as females may marry and be well adjusted to their sex role. Others, especially when there has been equivocation regarding sex of rearing in infancy, may be less than attractive as women because of indecisive therapy. Psychiatric studies indicate that the best emotional adjustment comes from directing endocrine, surgical, and psychiatric measures toward improving the person’s basic characteristics. Fortunately, this is consonant with the surgical and endocrine possibilities for those reared as females, because current operative techniques can produce more satisfactory feminine than masculine external genitalia. Furthermore, the testes of male hermaphrodites are nonfunctional as far as spermatogenesis is concerned. Only about one-third of male hermaphrodites are suitable for rearing as males.
Figure 3–17. External genitalia in male hermaphroditism. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
Classification
Since about 1970, considerable progress has been made in identifying specific metabolic defects that are etiologically important for the various forms of male hermaphroditism. Details are beyond the scope of this text. Nevertheless, it is important to point out that all cases of male hermaphroditism have a defect in either the biologic action of testosterone or the MIF of the testis. Furthermore, it now seems apparent that nearly all—if not all—of these defects have a genetic or cytogenetic background. The causes and pathogenetic mechanisms of these defects may vary, but the final common pathway is 1 of the 2 problems just mentioned; in the adult a study of the serum gonadotropins and serum steroids, including the intermediate metabolites of testosterone, can often pinpoint a defect in the biosynthesis of testosterone. In other cases, the end-organ action of testosterone may be defective. In children, the defect is sometimes more difficult to determine before gonadotropin levels rise at puberty, but one may suspect a problem by observing abnormally high levels of steroids that act as substrates in the metabolism of testosterone. A working classification of male hermaphroditism is as follows:
I. Male hermaphroditism due to a central nervous system defect
A. Abnormal pituitary gonadotropin secretion
B. No gonadotropin secretion
II. Male hermaphroditism due to a primary gonadal defect
A. Identifiable defect in biosynthesis of testosterone
1. Pregnenolone synthesis defect (lipoid adrenal hyperplasia)
2. 3β-Hydroxysteroid dehydrogenase deficiency
3. 17α-Hydroxylase deficiency
4. 17,20-Desmolase deficiency
5. 17β-Ketosteroid reductase deficiency
B. Unidentified defect in androgen effect
C. Defect in duct regression (Figs. 3–18 and 3–19)
Figure 3–18. External genitalia in male hermaphroditism. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
Figure 3–19. Internal genitalia of the patient whose external genitalia are shown in Figure 3–18.
D. Familial gonadal destruction
E. Leydig cell agenesis
F. Bilateral testicular dysgenesis
III. Male hermaphroditism due to peripheral end-organ defect
A. Androgen insensitivity syndrome (Fig. 3–20)
Figure 3–20. Androgen insensitivity syndrome.
1. Androgen-binding protein deficiency
2. Unknown deficiency
B. 5α-Reductase deficiency
C. Unidentified abnormality of peripheral androgen effect
IV. Male hermaphroditism due to Y chromosome defect
A. Y chromosome mosaicism (asymmetric gonadal differentiation) (Fig. 3–21)
Figure 3–21. Internal genitalia in asymmetric gonadal differentiation. (Reproduced, with permission, from Jones HW Jr, Scott WW. Hermaphroditism, Genital Anomalies and Related Endocrine Disorders. 2nd ed. Philadelphia, PA: Williams & Wilkins; 1971.)
B. Structurally abnormal Y chromosome
C. No identifiable Y chromosome
9. Differential Diagnosis in Infants with Ambiguous Genitalia
Accurate differential diagnosis is possible in most patients with ambiguous genitalia (Table 3–8). This requires a complex history of the mother’s medication use, a complex sex chromosome study, rectal examination for the presence or absence of a uterus, measurement of serum steroid levels, pelvic ultrasonography, and information about other congenital anomalies. The following disorders, however, do not yield to differentiation by the parameters given in Table 3–8: (1) idiopathic masculinization, (2) the “special” forms of female hermaphroditism, (3) 46,XX true hermaphroditism, and, occasionally, (4) the precise type of male hermaphroditism. For these differentiations, laparotomy may be necessary for diagnosis and for therapy.
Table 3–8. Differential diagnosis of ambiguous external genitalia.
10. Treatment of Hermaphroditism
The sex of rearing is much more important than the obvious morphologic signs (external genitalia, hormone dominance, gonadal structure) in forming the gender role. Furthermore, serious psychological consequences may result from changing the sex of rearing after infancy. Therefore, it is seldom proper to advise a change of sex after infancy to conform to the gonadal structure of the external genitalia. Instead, the physician should exert efforts to complete the adjustment of the person to the sex role already assigned. Fortunately, most aberrations of sexual development are discovered in the newborn period or in infancy, when reassignment of sex causes few problems.
Regardless of the time of treatment (and the earlier the better), the surgeon should reconstruct the external genitalia to correspond to the sex of rearing. Any contradictory sex structures that may function to the patient’s disadvantage in the future should be eradicated. Specifically, testes should always be removed from male hermaphrodites reared as females, regardless of hormone production. In cases of testicular feminization, orchiectomy is warranted because a variety of tumors may develop in these abnormal testes if they are retained, but the orchiectomy may be delayed until after puberty in this variety of hermaphroditism.
In virilized female hermaphroditism due to adrenal hyperplasia, suppression of adrenal androgen production by the use of cortisone from an early age will result in completely female development. It is no longer necessary to explore the abdomen and the internal genitalia in this well-delineated syndrome. The surgical effort should be confined to reconstruction of the external genitalia along female lines.
Patients with streak gonads or Turner’s syndrome, who are invariably reared as females, should be given exogenous estrogen when puberty is expected. Those hermaphrodites reared as females who will not become feminized also require estrogen to promote the development of the female habitus, including the breasts. In patients with a well-developed system, cyclic uterine withdrawal bleeding can be produced even though reproduction is impossible. Estrogen should be started at about age 12 and may be given as conjugated estrogens, 1.5 mg/d orally (or its equivalent). In some patients, after a period of time, this dosage may have to be increased for additional breast development. In patients without ovaries who have uteri and in male hermaphrodites in the same condition, cyclic uterine bleeding can often be induced by the administration of estrogen for 3 weeks of each month. In other instances, this may be inadequate to produce a convincing “menstrual” period; if so, the 3 weeks of estrogen can be followed by 3–4 days of progestin (eg, medroxyprogesterone acetate) orally or a single injection of progesterone. Prolonged estrogen therapy increases the risk of subsequent development of adenocarcinoma of the corpus, so periodic endometrial sampling is mandatory in such patients.
Reconstruction of Female External Genitalia
The details of the operative reconstruction of abnormal external genitalia are beyond the scope of this chapter. However, it should be emphasized that the procedure should be carried out at the earliest age possible so as to enhance the desired psychological, social, and sexual orientation of the patient and to facilitate adjustment by the parents. Sometimes the reconstruction can be done during the neonatal period. In any case, operation should not be delayed beyond the first several months of life. From a technical point of view, early operation is possible in all but the most exceptional circumstances.
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