Chpter 9 Normal and Abnormal Sexual Development

1. CLINICAL GYNECOLOIC ENDOCRINOLOGY AND INFERTILITY Chpter 9 Normal and Abnormal Sexual Development OBGY R1 Lee Eun Suk

2. Normal Sex Differentiation • Gender identity the result of the following determinants • Genetic sex • Gonadal sex • Internal & external genitalia • The secondary sexual characteristics that appear at puberty • The role assigned by society • Four major steps which constitute normal sexual differentiation • Fertilization and determination of genetic sex • Formation of organs common to both sexes • Gonadal differentiation • Differentiation of the internal ducts and external genitalia

3. Fertilization and Determination of Genetic Sex • Step 1 in sex differentiation:Determination of genetic sex Egg (23,X) + Sperm (23,X)=46,XX genetic female OR Egg (23,X) + Sperm (23, Y)=46, XY genetic male karyotype

4. Formation of Organs Common to Both Sexes • The fertilized egg multiplies to form a large number of cells Differentiation of the sex organs in this development ; At that stage, both 46,XX & 46,XY fetuses have similar sex organs, specifically • Gonadal ridges • Internal ducts • External genitalia

5. Internal reproductive organs

7. Gonadal Differentiation • The important event in gonadal differentiation is of the gonadal ridge to become either an ovary or a testis • In males, the gonadal ridge develops into testes as a result of a product from a gene located on the Y chromosome “Testis determining factor" (TDF) : Gonadal medullary resion -> Sertoli cell “Sex determining region of the Y chromosome" (SRY) • In females, the absence of SRY, due to the absence of a Y chromosome, permits the expression of other genes which will trigger the gonadal ridge to develop into ovaries

8. Gonadal Differentiation • Pseudoautosomal region • The distal ends of the short arms of the X and Y chromosomes • During meiosis the homologous distal short arms of the X and Y chromosomes pairs, and interchange of genetic material occurs in autosomes • Gene deletions in this area of the X chromosome (Xp22.3) are associated with various conditions short stature , mental retardation, X- linked ichthyosis, Kallmann’s sydrome

9. Gonadal Differentiation • Subsequent sexual differentiation requires direction by various genes with TDF • SRY: The Y chromosome sex determinants region • SOX9: An autosomal testis-determining gene • DAX1: A potential testis-suppressing gene on X-chromosome • SF1: The link between SRY and the male development pathway • WT1: necessary for normal renal and gonadal development • WNT4: A potential ovary-determining gene on an autosome

10. Gonadal Differentiation • SRY • Sex determining region of the Y chromosome • Locate on the short arm of the Y chromosome • Transcription factor contains HMG (high-mobility group) box - a DNA binding domain => conrol of gene transcrition • Investigations of the DNA-binding properties of the protein of SRY in the promoter P450 aromatase (conversion of testosterone to estradiol that is down-regulated in the embryo) & anti-mullerian hormone (responsible for regression of the mullerian ducts) • The expression of SRY in the tissue destined to become a gonad directs the cells of this gonadal primordium to differentiate as Sertoli cell

11. Gonadal Differentiation • SOX9 :An autosomal testis-determining gene (SRY-like box) genes are similar in sequence to SRY : an extra copy of SOX9 developes males, even if they have no SRY gene – XX mice made transgenic for SOX9 develop testes • SF1 ;Steroidogenic factor, necessary to make the bipotential gonad Collaboration with SOX9  elevate levels of AMH transcription • Wnt4 :Activate DAX1 expression Lack the Wnt4 gene  Ovary fail to form properly, express testis specific markers

12. Postulated cascades leading to the formation of the sexual phenotypes

13. Summary of key genetic events in early sex determination • Migration of primordial germ cells to the urogenital ridge • Differentiation of the bipotential gonadal tissue under the direction of WT-1 and SF-1 • SRY activation of male-specific genes, especially SOX9, to produce the testes by cell proliferation, differentiation, migration and vascularization • Ovarian differentiation by suppression of SOX9 through the activity of DAX1 and Wnt4

14. Summary of the genetics of gonadal dysgenesis • Gonadal streaks without germ cells in XX or XY (female phenotype) : • Deficiencies inWT-1 or SF-1 • Lack of testicular development in XY individuals pure gonadal dysgenesis (female phenotype) : • Deficiencies in SRY or SOX9 • Male phenotype in a 46,XX individual : • Presence of SRY • Mixed gonadal dysgenesis in mosaics (varying phenotype) : • Excess DAX1

15. Internal reproductive organs - Embryonic development • Urinary and genital tract • Closely related, anatomically and embryologically • Embryologic urinary system -> important inductive influence on developing genital system • Anomalies in one system are often mirrored by anomalies in another system • Urinary system, internal reproductive organs & external genitalia • Develop synchronously at an early embryologic age

16. Kidney, renal collecting system & ureters from nephrogenic cord

17. Mesonephric (Wolffian) duct • Singular importance for the following reasons • Grows caudally in developing embryo to open an excretory channel into the primitive cloaca and outside world • Serves as starting point for development of the metanephros which becomes definitive kidney • Differentiates into the sexual duct system in male • Although regressing in female fetuses, inductive role in development of the paramesonephric or mullerian duct

18. Mullerian duct • Paramesonephric or mullerian ducts - development • Form lateral to mesonephric ducts • Grow caudally and then medially to fuse in midline • Contact urogenital sinus in region of the post. urethra at slight thickening known as sinusal tubercle

19. Duct System Differentiation - Male • TDF • Results in degeneration of gonadal cortex and differentiation of the medullary region of the gonad into Sertoli cells • Sertoli cells • Secrete glycoprotein known as anti-mullerian hormone(AMH) • Regression of paramesonephric duct system in male embryo • Signal for differentiation of Leydig cells from the surrounding mesenchyme

20. Duct System Differentiation - Female fetus • In the absence of TDF, medulla regresses and cortical sex cords break up into isolated cell clusters (-> primordial follicles) • In the absence of AMH & testosterone • Mesonephric duct system degenerates • Then, paramesonephric duct system develops • Inf. fused portion • Uterovaginal canal -> uterus and upper vagina • Cranial unfused portions • Open into celomic cavity (future peritoneal cavity) • Fallopian tubes

21. Anti-Müllerian Hormone • A member of the transforming growth factor-ß family • Regression of Müllerian duct system in male embryo • AMH has an inhibitory effect on oocyte meiosis • Plays a role in the descent of the testes • Inhibits surfactant accumulation in the lungs • Proteolytic cleavage of AMH produces fragments that have the ability to inhibit growth of various tumor ( a potential therapeutic application)

22. Duct System Differentiation • Leydig cells • Produce testosterone & dihydrotestosterone with 5a-reductase • Testosterone • Responsible for evolution of mesonephric duct system into vas deferens, epididymis, ejaculatory duct & seminal vesicle • At puberty, leads to spermatogenesis & changes in primary and secondary sex characteristics • DHT(dihydrotestosterone) • Results in development of the male external genitalia , prostate and bulbourethral glands

23. External Genitalia Differentiation • In the female, absence of androgens permits the external genitalia to remain feminine • The genital tubercle becomes the clitoris • The labioscrotal swellings → the labia majora • The urogenital folds → the labia minora • In the male, fetal androgens from the testes masculinize the external genitalia • The genital tubercle grows to become the penis • The labioscrotal swellings fuse to form the scrotum

24. External Genitalia

25. Abnormal Sexual Development

26. CLASSIFICATION OF INTERSEXUALITY Disorders of fetal Endocrinology

27. CLASSIFICATION OF INTERSEXUALITY Primary gonadal defect – Swyer syndrome

28. How many children are born with intersex conditions? • A conservative estimate is that 1 in 2000 children born will be affected by an intersex condition • 98 % of affected babies are due to congenital adrenal hyperplasia

29. EXCESS FETAL ANDROGENS Congenital adrenal hyperplasia 21 -hydrxylase deficiency 11-hydroxylase deficiency 3ß-hydroxysteroid dehydrogenase deficiency EXCESS MATERNAL ADROGEN Maternal androgen secreting tumours (ovary, adrenal) Maternal ingestion of androgenic drugs FEMALE PSEUDOHERMAPHRODITISM

30. 21-hydrxylase deficiencycongenital adrenal hyperplasia Cholesterol Pituitary Pregnenolone Progesterone ACTH 17-OH progesterone Adrenal cortex 21-hydroxylase  Androgens Cortisol Cortisol Androgens

31. Masculinized females CONGENITAL ADRENAL HYPERPLASIA • There are several different forms of CAH, each related to one of the enzymes necessary to transform cholesterol to cortisol (hydrocortisone) • StAR / 20,22-hydroxylase, 3 -hydroxysteroid-dehydrogenase / 17-hydroxylase / 21-hydroxylase and 11 -hydroxylase • When one of these enzymes is deficient, this leads to a hyperfunction and increased size (hyperplasia) of the adrenals • Among the various forms of CAH, the 21-hydroxylase deficiency is by far the most frequent, representing more than 95% of all cases • Defect in cortisol biosynthesis, with or without aldosterone def, + androgen excess

32. Masculinized females CONGENITAL ADRENAL HYPERPLASIA - Biochemistry • Steroid 21 hyrdoxylase is a cytochrome p-450 enzyme located in ER • Catalyzed the conversion of 17-hydoxyprogesterone to 11-hydroxycortisol :precursor of cortisol • Conversion of progesterone to deoxycortisterone :precursor of aldosterone • This enzyme deficiency -> adrenal cortex is stimulated & over production of cortisol precursor

33. Pathways of steroid biosynthesis in the adrenal cortex

34. Hypothalamic pituitary adrenal axis Renin-angiotensin-aldosterone axis

35. Clinical menifestation • Salt wasting type : severe form with a concurrent defect in aldosterone synthesis • Simple virilizing type : normal aldosterone biosynthesis • Both are together termed classic 21-hydroxylase deficiency • There is also a mild,nonclassic form : may be asymptomatic • Classic 21-hydroxylase deficiency : 1 in 16,000 births

36. CONGENITAL ADRENAL HYPERPLASIA - salt wasting • 75 percent of patients with classic 21-hydroxylase deficiency  severely impaired 21-hydroxylation of progesterone  cannot synthesis of aldosterone • Elevated of 21-hydroxylase precursor: aldosterone antagonist • Aldosterone deficiency  hypovolemia and hypereninemia, and hyperkalemia (esp. in infant) • Cortisol deficiency poor cardiac contractility, poor vascular resp. to catecholamine, and GFR ↓ , antidiuretic H ↑

37. CONGENITAL ADRENAL HYPERPLASIA - salt wasting • So together : hyponatremic dehydration and shock • Adrenal medulla : depending on the glucocorticoid in part  so salt wasting 21hyroxylase deficiency  catecholamine deficiency  exacerbating shock Identify : e, aldosterone, plasma renin (hyperkalemia and low aldosterone and hyperreninemia)

38. Ambiguous genitalia • Girls with classic 21-hydroxylase deficiency : exposed to high level of adrenal androgen level (GA 7 wks) • Girls with ambiguous genitalia: -a large clitoris -rugated and partially fused labia majora -uterus,fallopian tubes, and ovaries : normal • Boys : -no overt signs of the disease except variable and subtle hyperpigmentation and penile en-largement

39. Postnatal virilization • Exposed to the high levels of sex steroids • Rapid growth (usually androgen effect) • Advanced bone age  premature epiphyseal closure (androgen’s extragonatal aromatization of estrogen) • Pubic and axillary hair: early develop • Girl : clitorial growth • Young Boys : penile growth • Long term stimulation : central precocious puberty

40. Linear growth • A meta-analysis of data from 18 centers showed : 1.4 SD below the population mean • Both undertreatment and overtreatment : risk for short stature • Undertreatment : causing premature epiphyseal closure induced by high levels of sex steroids • Overtreatment : glucocorticoid-induced inhibition of the growth

41. Reproductive function • Girls : problem at the reproductive system -oligomenorrhea, amenorrhea -prenatal androgen exposure: effect to sex-role behavior • Boys : fewer problems

42. CONGENITAL ADRENAL HYPERPLASIA - Diagnosis

43. CONGENITAL ADRENAL HYPERPLASIA • Normal infant :100ng/dl (17 hydroxyprogesterone) • Affected infants:10000ng/dl ↑ • New born -> screening test • 10% (severe affected infant) : 17 hydroxyprogesterone level ↓ • Preterm or sick infants -> 17 hydroxyprogesterone level ↑↑ • The severity of hormonal abnormalities: depends on the type of 21-hydroxylase def • Salt wasting : 17-hydroxyprogesterone: 100000ng/dl

44. CONGENITAL ADRENAL HYPERPLASIA • Random 17-hydroxyprogesterone(17-Ohpro) >80 ug/L or 242 nmol/L (nl : <2.95 ug/L or 9 nmol/L) • Salt losing > nonsalt loser -> corticotropin stimulation test : unneeded • Genetic analysis -> prenatal testing • Nonclassic CAH : random 17-OHpro : nl -> Screening : early morning basal 17-OHpro level >1.5 ug/L (4.5 nmol/L) in children & >2.0 ug/L(6nmol/L) in women during follicular phase of menstrual cycle -> Corticotropin ST(250ug of tetracosactide(cosyntropin) ) : any time during the day => positive : 17-OHpro >15.0 ug/L(45nmol/L)

45. CONGENITAL ADRENAL HYPERPLASIA • Heterozygote carrier : mild elevation, 17-OHpro <10 ug/L(30nmol/L) following CST • Carrier : symptoms or signs of the disease • Premature adrenarche 1/3 : heterozygote carriers of 21-OH def (recent) • 17-OHpro :10-15 ug/L -> DDx of heterozygote carrier or homozygote affected Pt -> Genetic analysis

46. CONGENITAL ADRENAL HYPERPLASIA – Management • According to the clinical course & hormonal level • Purpose : normal growth, B.Wt, pubertal development, optimal adult height • Growth velocity, body Wt velocity, bone age maturation F/U • Classic 21-OH def -> glucocorticoid : adrenal androgen secretion ↓ -> mineralocorticoid : electrolytes & plasma renin activity normalization

47. Treatment Problems • Hypercortisolism (iatrogenic Cushing’s syndrome) - Sn & Sx : obesity, growth failure, adult short stature, striae, osteoporosis, hyperlipidemia • Symptoms of hyperandrogenism : virilism, infertility of female, precocious virilisation of male, early puberty, adult short stature

48. Glucocorticoid and mineralocorticoid therapy • Hydrocortisone of physiologic dose -> corticotropin & androgen production suppression (Х): 6 mg/m2/day • Hydrocortisone 12-15mg/m2/day -> sufficient androgen suppression • 20 mg/m2/day (neonatal period) • 25 mg/m2/day -> do not use • Dose variability factor • Indivudual variation in the metabolism and sensitivity • Previous degree of hypothalamic-pituitary-adrenal axis suppression • High dose glucocorticoid short course therapy

49. Glucocorticoid and mineralocorticoid therapy • Mineralocorticoid (fludrocortisone) : plasma renin activity ->mid normal range • Dose :100-200 ug/day • Nonsalt losing :elevated plasam renin activity • Infant with salt losing : NaCl supply ->1-2 g/day • 17mEq Na/NaCl 1g

50. CONGENITAL ADRENAL HYPERPLASIA – Monitoring therapy • Commonly, serum 17-hydroxyprogesterone & androstenedione level • Testosterone level in female and prepubertal male • Test time : early morning(8:00 AM) • Target level of 17-OHpro : 4-12 ug/L (12-36 nmol/L)