Sex Determination and Differentiation and this necessitated a finer modification of the said
Dra. Gloria Marie Valerio experiment
o Several experiments were done over a period of 10 Sex Determination years and the conclusion was, if the Barr body is - Factors that make an individual a male or a female present in AT LEAST 20 out of 100 cells tested, the - Established at fertilization subject is a genetic female o If the Barr body is present in 0-4 cells out 100 cells Gametogenesis tested, the subject is a genetic male - Oogenesis - In blood smears, the Barr body is present in the neutrophil, o Development and maturation of the female sex cell attached to the nucleus, appearing as an appendage o Occurs in the ovaries o If the Barr body is present in 1-5 out of 300 o An oocyte has 44 autosomal chromosomes and 2 sex neutrophils, the test is positive and the subject is a chromosomes (XX) genetic female o An oocyte undergoes several mitotic divisions and 2 - Vaginal smears have the highest positivity among the three meiotic divisions, giving rise to a mature ovum with 22 smears simply because of the direct effects of estrogen on the autosomal chromosomes and 1 chromosome (always vaginal epithelium X) - Spermatogenesis Clinical Sex Sex No. of Total No. of o Development and maturation of the male sex cell Conditions Chromatin Chromosomes Autosomal Chromosomes o Spermatocytes has 44 autosomal chromosomes and 2 Test Chromosomes Normal (+) XX 44 46 sex chromosomes (XY) Female o Just like the oocyte, the spermatocytes will undergo Normal Male (-) XY 44 46 several mitotic divisions and 2 meiotic divisions, giving Super Female (+) XXX 44 47 rise to a mature sperm cell with 22 autosomal Klinefelter’s (+) XXY 44 47 chromosomes and 1 sex chromosome (either X or Y) Turner’s (-) XO 44 45 - If during fertilization, the sperm cell that carries the X- Mongoloids (-) male XX or XY 45 47 chromosome fertilizes the ovum, the product or zygote will have (+) female 44 autosomal chromosomes and 2 sex chromosomes (XX), producing a normal genetic female Abnormal Sexual Chromosomal Constitution nd - On the other hand, if a Y-chromosome bearing sperm cell fertilizes - In normal males, after the 2 meiotic division, a mature sperm the ovum, the product or zygote will have 44 autosomal cell should only have 1 sex chromosome (X or Y) nd chromosomes and 2 sex chromosomes (XY), producing a normal - Similarly in normal females, after the 2 meiotic division, a genetic male mature ovum should only have 1 sex chromosome (always X) - Sex or gender of an individual means “genetic sex” which is - Any abnormalities in the sex chromosomal constitution of an established during fertilization individual is a result of the non-disjunction of the sex nd - An individual can have only 1 of the 2 sexes: chromosomes after the 2 meiotic division nd o XX female - If after the 2 meiotic division, there will be non-disjunction of o XY male the sex chromosome (1 sperm cell still has 2 sex chromosomes - For instance, a newborn child is delivered, a doctor’s basis of and the other has none or 1 ovum still has XX and the other has declaring the sex of the baby is primarily the appearance of the none), this may lead to sex chromosomal aberrations shown external genitalia below: o If a baby presents with a non-fused labia with a small OVUM SPERM clitoris, that baby will be declared as a female X XX O o If a baby presents with a fused scrotal sac with a penis X XX XXX XO or phallus, that baby will be declared as a male NORMAL FEMALE TRIPLE X FEMALE TURNER’S SYNDROME o If a newborn presents with an ambiguous external Y XX XXY YO genitalia, sex determination is more difficult, as in NORMAL MALE KLINEFELTER’S NON-VIABLE pseudo-hermaphrodites SYNDROME In cases like this, the doctor should first sign XY XXY XXXY XY the birth certificate and then perform a sex KLINEFELTER’S TRIPLE X MALE NORMAL MALE test SYNDROME O XO XX OO Sex Chromatin Test TURNER’S NORMAL MALE NON-VIABLE SYNDROME - In 1949, 2 scientists: Barr & Bertram, discovered the presence of the sex chromatin mass/body in female somatic cells - The first experiment was done on cat neurons and was later Hair Root Test modified in human samples of buccal smear, blood smear and - Pluck 3-4 strands of normal scalp hair with intact roots vaginal smear - Tease the hairroots and stain with a fluorescent dye and under a - They found the presence of the sex chromatin mass/body in microscope, look for fluorescent “F body” or “Y body” female somatic cells and later on, this sex chromatin mass was - “Y body” is part of the Y-sex chromosome called the Barr body - Present and positive only in genetic males - They concluded that if the Barr body is present, the sex chromatin - Invariably absent in genetic females test is positive and this is usually true when the sex chromosomal constitution of the individual is XX: (+) Barr body, (+) sex Karyotyping or Chromosomal Analysis chromatin test normal genetic female - Most accurate but most expensive - In a buccal smear taken from the inner cheek, the Barr body can - A tissue culture is done where autosomal chromosomes are be found in the inner surface of the nuclear membrane paired with each other as well as the sex chromosomes o Problems may arise with false positives where a - This test cannot be done by a mere medical technologist or a chromatin mass is incorrectly identified as a Barr body, medical doctor, only by a scientist who specializes in Genetics - After all of the chromosomes have been paired up with each - Absence of testes (in females or in males with non-functional other, an individual’s chromosomal constitution can be testes) determined o Mullerian ducts will develop and Wolffian ducts will regress or involute Sex Differentiation o Ovaries – not needed for development of female - Gonads are the primary organs of reproduction internal genitalia - Testes in males and ovaries in females o Genital ducts have inherent tendency to feminize in - Genital ducts give rise to the internal genitalia the absence of functional testes o Wolffian ducts – male internal genitalia o Mullerian ducts – female internal genitalia - Structures that make up the external genitalia Male Differentiation - Genetic male – embryonic testes secretes: Gonadal Differentiation o MRF (Mullerian Regression Factor) inhibits - The testes and ovaries share a common anlage, the genital ridge development of Mullerian ducts no female internal (bilateral) genitalia (action is unilateral) th - From fertilization until the 7 week of embryonic life, the genital In the case of a non-functional testes, no ridge is indifferent, thus there is no anatomical evidence whether substance will be secreted to inhibit the it will develop into testes or ovaries development of the Mullerian ducts - The genital ridge is divided into 2 parts: inner medulla and outer o DOS (Duct Organizing Substance) and Testosterone cortex stimulate the development of the Wolffian ducts - So although the testes and ovaries will come from the same - In the presence of these three secretory products of the structure, each will come from a different part of that structure FUNCTIONAL embryonic testes, development of the Mullerian (medulla for testes and cortex for ovaries) ducts is inhibited and development and maturation of the - Also genetically determined Wolffian ducts is stimulated
Testicular Development Female Differentiation
th th - In the presence of the Y-chromosome during the 7 – 8 week of - Without testes No MRF No DOS No Testosterone life, the medulla develops into testes while the cortex regresses genital ducts will have inherent tendency to feminize o Present in the Y-chromosomes is a membrane protein - Presence of ovary is not a prerequisite for the development of the called H-Y antigen which is the Testes-Determining female internal genitalia Product - No gonad (e.g. Turner’s syndrome) No MRF, DOS, Testosterone still normal development of uterus and fallopian tubes because Ovarian Development again, in the absence of functional testes, the genital ducts have - In the absence of the Y-chromosome and in the absence of the H- an inherent tendency to feminize th th Y antigen between the 8 – 10 week of life, it is the cortex that o Rudimentary gonad only uterus and fallopian tubes will develop into the ovaries while the medulla regresses develop, regardless of chromosomal sex - Presence of XX is a prerequisite for normal ovarian development o XO – neither testicular nor ovarian differentiation Gonad will remain as a long pale streak of Differentiation of External Genitalia connective tissue Anlage Male Female - Hormonal treatment of the mother during pregnancy will have no Genital tubercle Penis Clitoris effect on gonadal development Thus, gonadal differentiation is Urethral folds Part of corpus spongiosum Labia minora genetically determined Laboscrotal swelling Fuse to form scrotal sac Labia majora Urogenital slit Closed, complete fusion Vagina Differentiation of Genital Ducts - Just like the gonads, both the Mullerian and Wolffian ducts are Development of External Genitalia present in their indifferent stage during the early weeks of - Males embryonic life o 44 autosomal chromosomes and 2 sex chromosomes o In male differentiation, Mullerian ducts regress and XY Wolffian ducts develop o Y-chromosome H-Y antigen testes will develop o Conversely, in female differentiation, Wolffian ducts o Functional testes MRF, DOS, Testosterone regress and Mullerian ducts develop Mullerian ducts regress, Wolffian ducts develop into - Mullerian ducts will give rise to the female internal genitalia male internal genitalia o Uterus o In the case of a functional testes, it will also secrete o A pair of fallopian tubes or oviducts dihydrotestosterone in addition to testosterone - Wolffian ducts will give rise to the male internal genitalia These 2 will masculinize the external o Epididymis genitalia o Vas deferens Because just like the internal genitalia, in o Seminal vesicles the absence of a functional testes, the o Ejaculatory duct external genitalia has an inherent tendency - Presence of functional testes to feminize o In males (XY), the Y-chromosome indicates the Without testosterone and presence of the H-Y antigen testes develop dihydrotestosterone female external o If the testes are functioning normally, it will stimulate genitalia the Wolffian ducts complete their development o In male adults, testosterone is also responsible for the (Mullerian structures involute and the male internal development of male secondary sexual characteristics genitalia will fully develop) that will include: growth of facial hair, body hair, pubic o The mere presence of the testes will not guarantee hair, development of a low-pitched voice and extra that it will be the Wolffian ducts that will develop. It protein deposition in the muscle (anabolic nature of should also be functioning normally testosterone so males are more muscular than females) - Females - High FSH and LH: more than 96 mouse units / 24 hours o XX no Y ovaries develop no MRF, DOS, - Negative sex-chromatin pattern Testosterone feminine external genitalia o Main secretory product of ovary: estrogen Female Pseudohermaphroditism o Estrogen is responsible for development of female - Large phallus, labia majora resembling a scrotal sac, incomplete secondary sexual characteristics that will include: closure of the urogenital sinus development of a high-pitched voice, extra fat - Short stature, with body hair and facial hair, masculine deposition into the subcutaneous tissues of the distribution of pubic hair, small breasts, muscular breasts, buttocks and thighs - XX presence of ovaries No MRF, DOS and Testosterone - Thus, no amount of surgical or hormonal modification in an female internal genitalia ambiguity is confined exclusively to individual born male or female will ever change his or her genetic the external genitalia makeup - External genitalia of newborn female pseudohermaphrodite with congenital adrenal hyperplasia Components of an Individual’s Sexual Identity o If adrenal glands are hyperplastic, androgen secretion Male Female is elevated and this will masculinize the external Chromosomal/Genetic Sex XY XX genitalia Nuclear Sex (-) Barr body (+) Barr body o If the pregnant mother is taking exogenous androgens Gonadal Sex Testes Ovary th before the 12 week of gestation, it will have an effect Hormonal Sex Mainly androgen Mainly estrogen on the external genitalia of the developing fetus Internal Genitalia Wolffian ducts Mullerian ducts ambiguous genitalia o Measurement of androgen levels is now a part of Sex Chromosomal Constitutions in Ambiguous External Genitalia newborn screening procedures - Normal 46 XX female - Ovaries, Mullerian derivatives normally developed - Normal 46 XY male - Anatomical ambisexuality – limited to external genitalia - Abnormal sex chromosome constitutions - Masculinzation – due to extragonadal androgens - Even with severe masculinisation of external genitalia – uterus Differential Diagnosis of Ambiguous External Genitalia and fallopian tubes remain normal - Perform a Sex Chromatin Test o (+) Result may suggest: True Hermaphroditism Female pseudo-hermaphroditism - Not only caused by non-disjunction of sex chromosomes, a True hermaphroditism mutation of the sex chromosomes is also implicated in the Chromatin positive variants of gonadal etiology of true hermaphroditism dysgenesis (e.g. Klinefelter’s XXY so even if - Presence of both ovarian and testicular tissues the subject is a male, he could test positive - An ovotestis may be observed on histological examination for the sex chromatin test) - External genitalia – may resemble either male or female o (-) Result may suggest: o Often ambiguous Male pseudo-hermaphroditism o ¾ of patients reared as males because of size of phallus True hermaphroditism - In virtually all cases there is a uterus Chromatin negative variants of gonadal - 2/3 of patients have appreciable breast development dysgenesis (e.g. Turner’s syndrome XO) - 70% - are chromatin positive o (+) menstruate Male Pseudohermaphroditism o (-) will not menstruate - XY sex chromosomes Gonads: exclusively testes but the mere - Note well that true hermaphrodites can never impregnate presence of testes is not an actual determinant of one’s sex and themselves thus it should also be functional - One cause of male pseudohermaphroditism is testicular failure Case testes is not functioning normally or if the testes is functioning - 20 y/o – reared as female normally but there are inadequate receptors in the target cells for - Physical examination (Dept. Of Ob-Gyn FEUH) the hormone secreted by the testes o Ht: 5’6 and ½“ - Wt: 138 lbs. o Because there is an inherent tendency of the internal o Breasts: not developed and external genitalia to feminize in the presence of a - External genitalia is ambiguous: 2 scrotal-like labia each containing a 2 x 3 x non-functional testes, the inherent tendency to 2 cm rounded, slightly firm, movable mass - A longitudinal groove in the middle of the 2 labia feminize will not be overcome resulting to both o Upper end – enlarged clitoral-like structure ambiguous external and internal genitalia o Lower end – opening that enters into a vagina-like cavity – 10 - Genital ducts or external genitalia cm long admits only a probe o Lacks masculinisation - Internal genitalia - no grossly palpable uterus, tubes or ovaries (by rectal o Some degree of feminization exam) - Failure of fetal testes to overcome inherent tendency to feminize - Sex chromatin test (Clinical Laboratory, Department of Pathology, FEU- o Secretory failure of testes NRMF) Specimen: o Buccal smear and blood smear o Failure of target tissues to respond normally o Hair root test is (+) - Results Turner’s Syndrome o Barr chromatin bodies - Short stature, prominent webbing of neck, lack of breast Negative / 600 cells development, scant pubic hair, small phallus, child-like Control: 2 normal female average count: 69/100 appearance cells (+) Barr bodies - XO no gonads normal uterus and fallopian tube external o Drumstick – 0/500 PMN o Hair root – 2 slides prepared genitalia is female due to inherent tendency to feminize female 15/100 cells and 18/100 cells internal genitalia will remain infantile due to absence of ovaries With “Y-bodies” under fluorescent and estrogen no maturation of internal and external genitalia microscope - No estrogen no androgen no negative-feedback high FSH and high LH Paolo Warren (Med-IA)