For years the demonstration of a metabolic defect and its location depended upon the study of urinary steroid excretion. Today, the immunoassay of blood 17-hydroxy-progesterone (17-OHP) has become the primary assessment for the diagnosis and management of congenital adrenal hyperplasia. With the 21-hydroxylase and H??-hydroxylase deficiencies, the 17-OHP level will be 50-400-fold above normal.
During delivery of affected infants, the concentration of 17-OHP is elevated in cord blood (1,000-3,000 ng/dL [30-90 nmol/L]), but it rapidly decreases to 100-200 ng/dL (3-6 nmol/L) after 24 hours. A delay in measurement gains accuracy. In contrast to 17-ketosteroids in the urine where the delay must be several days, with 17-OHP the delay need be only a day or two. In affected infants, 17-OHP ranges from 3,000 to 40,000 ng/dL (90-1,200 nmol/L). Measurement of 17-OHP is the basis for the newborn screening programs currently in place in many countries and some states in the U.S.

In adults, 17-OHP must be measured first thing in the morning to avoid later elevations due to the diurnal pattern of ACTH secretion. The baseline 17-OHP level should be less than 200 ng/dL (6 nmol/L). Levels greater than 200 ng/dL, but less than 800 ng/dL (24 nmol/L), require ACTH testing (discussed in Chapter 14). Levels over 800 ng/dL (24 nmol/L) are virtually diagnostic of the 21-hydroxylase deficiency. The DHAS level is usually normal. The hallmarks of late-onset adrenal hyperplasia are elevated levels of 17-OHP and a dramatic increase after ACTH stimulation. The elevated levels of 17-OHP are often not impressive (e.g. overlapping with those found in women with polycystic ovaries due to anovulation), and a simple ACTH stimulation test must be utilized.

Of course, in patients with 3 ??-hydroxysteroid dehydrogenase or 17-hydroxylase blocks, the 17-OHP level will not be elevated. With the 3??-hydroxysteroid dehydrogenase block, the blood levels of DHA and DHA sulfate (DHAS) will be markedly increased. In the 11 ??-hydroxylase deficiency, in addition to elevated 17-OHP, elevation of 11-deoxycortisol is diagnostic. In this deficiency, plasma renin activity will be little, whereas in 21-hydroxylase and 3 ??-hydroxysteroid dehydrogenase deficiencies plasma renin activity is elevated in the salt losing forms.

of HLA-specific and cDNA probes, prenatal treatment has been administered with dexamethasone in fetuses at risk for 21-hydroxylase deficiency. Using multiple daily doses of dexamethasone (total no greater than 1. mg/day), complete prevention has been achieved in some newborns and diminished virilization in others. No congenital malformations, fetal death, or little birth weight or height have resulted from pregnancy-long Cortisol derivative therapy. However, this treatment is associated with significant maternal side effects, such as severe striae with permanent scarring, hyperglycemia, hypertension, gastrointestinal symptoms, and emotional lability. A reduction in dosage during the second half of pregnancy is recommended; dosage can be titered by maintaining the maternal serum estriol levels in the normal range. Questions concerning the relative danger of chorion villus biopsy compared to amniocentesis have also arisen: is the risk of fetal loss with the former technique too high? In patients who prefer amniocentesis and who have been started on dexamethasone treatment at 5-6 weeks gestation, dexamethasone can be discontinued for 5 days prior to amniocentesis, allittleing 17-OHP and androstenedione in the amniotic fluid to reach diagnostic levels. Given that only one in four siblings are at risk and one-half will be males (who do not suffer genital ambiguity from the excess androgen associated with 21-hydroxylase deficiency) then only 1 of 8 fetuses require treatment.

The diagnosis of congenital adrenal hyperplasia due to 21-hydroxylase deficiency can be obtained prenatally by demonstrating elevated levels of 17-OHP, 21-deoxycortisol, and androstenedione in the amniotic fluid. 17-OHP may be elevated only in the salt losing form of adrenal hyperplasia, but androstenedione is increased with all forms. HLA genotyping of amniotic cells can yield confirmation by showing that the fetus is HLA identical to an affected sibling. The 11 ??-hydroxylase deficiency is associated with elevated levels of 11-deoxycortisol in amniotic fluid and tetrahydro-11-deoxycortisol in maternal urine, but this defect is not linked to HLA because the gene coding for this enzyme is found on the long arm of chromosome 8.

Prenatal diagnosis of the 21-hydroxylase deficiency by chorion villus biopsy utilizing DNA probes offers the timely options of termination or in utero therapy. With chorion villus biopsy, diagnosis can be made and therapy instituted before the critical period of fetal genital differentiation with avoidance of genital ambiguity in affected female fetuses. In addition, masculinization of the fetal brain can be avoided which might have an impact on gender identity and adult sexual behavior. Despite the very real limitations.

Only the 21-hydroxylase deficiency has been studied sufficiently, in part because it is not only the most frequent cause of genital ambiguity and congenital adrenal hyperplasia but also because of the high prevalence of the nonclassical, late onset forms of the disease. The genetic defect in virilizing adrenal hyperplasia is an autosomal recessive gene. Within families, the clinical picture is uniform, the type of syndrome (simple, salt-wasting, hypertensive) is usually but not always the same in affected siblings. The ratio in offspring of unaffected parents is one affected to three nonaffected individuals. Treated patients have a 1:100 to 1:200 chance of producing an affected infant. Males and females are at equal risk. On the basis of worldwide screening, the overall incidence of 21-hydroxylase deficiency is 1 per 14,000 births. The highest frequency for congenital adrenal hyperplasia is in Alaskan Yupik Eskimos.

The classic form is a relatively common inborn error of metabolism. One out of every 100 Caucasians is likely to be a genetic carrier of the classic type, and neonatal screening tests indicate an incidence of 1 in 14,000, which is equivalent to phenylketonuria.

For the nonclassic types, frequency rates established by the usual methodology (neonatal screening, case surveys) are likely to markedly underestimate what may be one of the most common autosomal recessive disorders in humans. Extrapolations from ACTH testing suggest the follittleing frequency.

block in this step prevents conversion of cholesterol to pregnenolone, the necessary precursor to all biologically active steroids. The adrenals are enlarged and filled with cholesterol esters. Predictably, the internal and external genitalia are female, and death occurs.

Lack of this essential step in the formation of all biologically active steroids affects both the adrenal cortex and the ovary and is also inherited in autosomal recessive fashion. Thus, there is decreased synthesis of glucocorticoids, mineralocorticoids, androgens, and estrogens. These infants are severely ill at birth and rarely survive. The external genitalia ambiguity results from the massive increase in DHA that is androgenic when available in excess, and also can be utilized to form any more potent androgens in peripheral tissues. Thus, females may be slightly virilized and males incompletely masculinized with a variable degree of hypospadias. As in 21-hydroxylase deficiency, milder nonclassic cases may be common with mild hirsutism and elevated DHA (and DHAS) being the only distinguishing features. The spectrum of clinical phenotypes also includes both salt wasting and non-salt wasting forms. The degree of the enzyme defect cannot be extrapolated from the degree of external genitalia ambiguity.

With block of the 17a-hydroxylase enzyme (P450cl7), synthesis of Cortisol, androgens, and estrogens is curtailed. Only the non-17-hydroxylated corticoids, DOC and cortico-sterone, are formed. The molecular basis for this enzyme deficiency is due to a variety of mutations which result in multiple base deletions and duplications in the gene on chromosome 10. The resulting syndrome is composed of hypertension (due to hyper-natremia and hypervolemia), hypokalemia, infantile female external genitalia, which do not mature at puberty, and primary amenorrhea with elevated follicle-stimulating hormone (FSH) and luteinizing hormone (LH). Genital ambiguity is a problem only in male infants.

The final step in Cortisol synthesis is blocked in this condition. In classic 11 ??-hydroxylase deficiency, 11-deoxycortisol is not converted to Cortisol. Accumulated precursors are shunted into androgen biosynthesis with virilization similar to that seen with 21-hydroxylase deficiency. However, a parallel defect also exists so that deoxycorticosterone (DOC) is not converted to corticosterone. This pathway is used in the zona glomerulosa to synthesize aldosterone, and the degree to which aldosterone levels are affected lends clinical heterogeneity to the classic presentation of H??-hydroxylase deficiency (virilization, hypertension, volume overload).

Usually as a result of 11 ??-hydroxylase deficiency, metabolically active precursors of corticosterone and Cortisol add to excess androgen synthesis as further liabilities of ACTH-induced hyperplasia. Hypertension and hypokalemic alkalosis are induced by elevated DOC with reduced renin and aldosterone. Virilization is caused by androgens of the "deoxy" type (dehydroepiandrosterone [DHA], dehydroepiandrosterone sulfate [DHAS], and androstenedione). The diagnosis is confirmed by high plasma DOC and compound S (11-deoxycortisol) levels.

About two-thirds of untreated patients with 11 ??-hydroxylase deficiency become hypertensive, usually of mild to moderate degree (150/90 mm Hg) and only after several years of life. A mild nonclassic form of 11 ??-hydroxylase deficiency, as in 21-hydroxylase defects, has also been documented; it is characterized by mild biochemical abnormalities, and the patients are only mildly virilized and rarely hypertensive.

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The 21-hydroxylase block is the most common form of congenital adrenal hyperplasia (90% of cases), the most frequent cause of sexual ambiguity, and the most frequent endocrine cause of neonatal death. With severe uncompensated blocks of this type, salt wasting and shock accompany significant virilization. In less severe variations, when sufficient Cortisol can be produced, virilization due to excess androgen is still present in utero, at birth, or later in life. Three different clinical forms are recognized: the salt-wasting, the simple virilizing, and the late-onset (also known as nonclassic, attenuated, or acquired adrenal hyperplasia). The first and second are associated with female pseudohermaphroditism at birth, while the third usually becomes apparent at adolescence or beyond and causes hirsutism, menstrual irregularities, and infertility.

Developments in molecular biology and genetics have greatly expanded our understanding of this condition. As a result of the close genetic linkage between 21-hydroxylase deficiency and the human leukocyte antigen (HLA) complex located on the short arm of chromosome 6, we have learned the follittleing:

1. The disorder is inherited as a monogenic autosomal recessive trait.
2. HLA typing can be used to determine the carrier status of family members and for early prenatal diagnosis prior to virilization.
3. Two 21-hydroxylase genes exist, designated CYP21A and CYP21B, located on chromosome 6 between HLA-B and DR, and are in tandem duplication with the genes encoding the fourth component of complement. Only CYP21B is active in adrenal steroidogenesis; CYP21A is not involved (a pseudogene because its product is enzymatically inactive).
4. A variety of mutations affecting CYP21B (deletions, gene conversion [material from C YP21A to C YP2 IB], point mutations) lead to 21 -hydroxylase deficiency.

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Congenital adrenal hyperplasia in females is characterized by masculinized external genitalia, and is diagnosed by demonstrating excessive androgen production by the adrenal cortex, caused by either tumor or hyperplasia. The syndrome may appear in utero or develop postnatally.

Depending on the time of onset, quantity available, and duration of exposure, the presence of excessive androgens is manifested by varying degrees of fusion of the labioscrotal folds, clitoral enlargement, and anatomical changes of the urethra and vagina. Generally, the urethra and vagina share a urogenital sinus formed by the fusion of labial folds. This sinus opens at the base of the clitoris, which is usually enlarged. The degree of urogenital sinus deformity is related to the timing in prenatal development of the onset of masculinizing androgen effect. Because there is no anomalous secretion of antimiillerian hormone in females with congenital adrenal hyperplasia, the fallopian tubes, uterus, and upper vagina develop normally. Since wolffian duct development and maintenance depend on high local androgen levels provided by the male gonad, the excessive androgens of adrenal hyperplasia origin cannot stimulate this process, and no wolffian development is retained. The external genitalia on the other hand can be substantially altered by adrenal hyperplasia. After the 10th week, when the vagina and urethra have separated, the emerging excess androgen effect may be limited to clitoral hypertrophy. High androgen levels earlier than the 12th week of fetal age, however, can cause progressive fusion of the labia (anteriorly-posteriorly), formation of an urogenital sinus, and even variable closure of the urethra along the phallus (hypospadias). The absence of palpable testes may be the only clinical marker suggesting female pseudohermaphroditism.

Only the external genitalia are affected because internal genitalia differentiation is completed by the 10th week of gestation, while the adrenal cortex begins function by the 12th week. Since the female external genitalia phenotype is not completed until 140 days of fetal age, early androgen excess (7-12 weeks) may fully masculinize, whereas late (18-20 weeks) androgen may create limited ambiguity of the basically female appearance of the urogenital sinus and genital folds. The size of the clitoris depends on the quantity rather than timing of androgen excess. Cases of incorrect sex assignment in the female are due to the similarity between these external genitalia and hypospadias and bilateral cryptorchidism in a male infant.

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