The embryology of the female genital tract is complex but generally well defined and is described in detail in Chapter 4. Briefly summarized, it involves both the medial migration and midline fusion of the müllerian (paramesonephric) ducts to form the uterus, cervix, and upper vagina, and the vertical fusion of that developing ductal system with the invaginating urogenital sinus that forms the lower vagina and the introitus. Outflow tract abnormalities that result from failure of müllerian duct development include vaginal/müllerian agenesis and AIS; the presence or absence of pubic hair distinguishes the two. Abnormalities caused by failure of vertical fusion include imperforate hymen and transverse vaginal septum or cervical atresia. Although all are uncommon and the clinician may have only limited or no previous experience with any of the four, each has unique and distinguishing features that, in most cases, point to the correct diagnosis at time of the initial visit. Only limited additional evaluation, described in a later section of this chapter, is required to firmly establish the diagnosis and to plan treatment.

In those with primary or secondary amenorrhea having a patent vagina and visible cervix, the likelihood of a genital outflow tract abnormality is very small. The only possibilities that need be considered are cervical stenosis and intrauterine adhesions (Asherman syndrome) or other endometrial damage that may result from surgical trauma or infection. Since these are acquired conditions, they result in secondary amenorrhea with an onset that typically correlates closely with the time of the previous insult. Both receive separate and thorough discussion in the later section of this chapter devoted to specific disorders of the genital outflow tract and uterus.

A serum estradiol measurement is easy to obtain, relatively inexpensive, and objective. Reasonably, one would expect to find normal estrogen levels in women with normal ovaries whose amenorrhea results simply from mild dysregulation and chronic anovulation, as in obese women and those with PCOS, and to find low estrogen levels in women with ovarian failure, pituitary disease, or more severe hypothalamic dysfunction. Unfortunately, serum estradiol concentrations can fluctuate erratically in all conditions, normal or low on any given day, and therefore can be misleading. A random estradiol concentration greater than approximately 40 pg/mL clearly suggests the presence of functional ovarian follicles but also is common during a premature or normal perimenopause and occurs sporadically in women with hypothalamic amenorrhea. A low random estradiol concentration may suggest ovarian failure, but also is typical of women with hypothalamic amenorrhea and may be observed in those with less severe forms of chronic anovulation, as in normal women during the early follicular phase.

The observation of “estrogenic” cervical mucus—clear, watery, and relatively abundant— suggests a normal level of ovarian estrogen production, but its absence cannot be interpreted confidently because many normal women exhibit such mucus only during the late follicular phase of the cycle when estrogen levels are relatively high, or not at all.

The progestin challenge test is based on the premise that progestin treatment (e.g., medroxyprogesterone acetate 10 mg daily for 5-7 days, or progesterone in oil 200 mg i.m.) will induce menses only in those having normal circulating estrogen concentrations. A pure progestational agent must be used because endogenous estrogen status cannot be inferred from the response to an OCP that contains both estrogen and progestin. The more potent synthetic progestins such as medroxyprogesterone acetate are a better choice than oral micronized progesterone, which must be administered in relatively high doses (e.g., 300 mg daily) to achieve a response.¹ A positive test—bleeding within 2-7 days after completion of progestin treatment—implies normal estrogen production and ovarian function, and a negative test—no withdrawal menses—suggests hypogonadism. Scant withdrawal bleeding or spotting suggests marginal levels of endogenous estrogen production. However, the overall correlation between withdrawal bleeding and estrogen status is far from perfect; both false positive (withdrawal bleeding despite generally low levels of estrogen production) and false negative results (absent bleeding despite significant estrogen production) are relatively common. Up to 40-50% of women whose amenorrhea relates to stress, exercise, weight loss, hyperprolactinemia, or ovarian failure, in whom estrogen levels generally are low, exhibit withdrawal bleeding.^2,3 Up to 20% of amenorrheic women
with significant estrogen production have no withdrawal bleeding,⁴ in some because the endometrium is decidualized by high circulating androgen levels.

The endometrial thickness, determined by transvaginal ultrasonography (the maximum 2-layer thickness in the mid-sagittal plane), is a measure of endometrial proliferation, which reflects the level of estrogen production. Endometrial thickness correlates with both the serum estradiol concentration and with the response to a progestin challenge in women with amenorrhea. In one study involving 44 women with secondary amenorrhea, endometrial thickness was significantly greater in 32 women who had withdrawal bleeding (10.3±4.1 mm) than in 12 who did not (5.0±1.3 mm); the serum estradiol level also was significantly greater (45.3±19.4 vs. 18.6±8.0 pg/mL), and an endometrial thickness measuring 6.0 mm or greater predicted withdrawal bleeding with 95% accuracy.² An added potential benefit of endometrial thickness as a measure of ovarian estrogen production is that it can help to identify individuals with chronic anovulation at low risk for having associated pathology such as hyperplasia or cancer.

The serum FSH concentration is another obvious and useful, but indirect, measure of ovarian function. A normal or low serum FSH level indicates the presence of functional ovarian follicles and may be observed in a variety of conditions associated with amenorrhea, including chronic anovulation (e.g., PCOS), pituitary disease, and hypothalamic dysfunction. A high serum FSH concentration is a reliable indication of ovarian follicular depletion or failure. Exceptions are rare and include inactivating mutations involving the FSH or LH receptor, enzyme deficiencies (17a-hydroxylase, aromatase), and functional pituitary and ectopic FSH-secreting tumors. Because the clinical implications of an elevated FSH level are serious, one or more repeated measurements are warranted to confirm the finding.

Although certainly not inappropriate, generally it is not necessary or helpful to also measure the serum LH concentration because levels of the two gonadotropins typically move in parallel. The one notable and highly relevant exception in women with amenorrhea—the “monotropic” rise in FSH that signals a more advanced stage of follicular depletion—can be detected by measuring FSH alone. A moderately increased serum LH concentration frequently is observed in women with PCOS, but is not a diagnostic criterion and has no other clinical relevance. A “reversed” LH/FSH ratio (LH lower than FSH), like that seen in prepubertal girls, suggests but does not prove hypothalamic dysfunction. During the midcycle gonadotropin surge in ovulatory cycles, LH levels increase more than those of FSH, but that has little relevance in women with amenorrhea. Other conditions in which levels of the two gonadotropins diverge significantly are truly rare and include ectopic gonadotropin secretion by tumors outside the reproductive tract, single gonadotropin deficiencies resulting from mutations in genes encoding the b-subunit of LH or FSH, and the very rare functional gonadotroph adenoma that secretes clinically important amounts of one gonadotropin (FSH) but not the other.

Measurement of the serum FSH level traditionally has been recommended only for those having demonstrable evidence of hypogonadism (e.g., a negative progestin challenge), as the means to differentiate patients with gonadal failure from those with hypothalamic or pituitary causes of amenorrhea. However, routine measurement of serum FSH in the evaluation of amenorrhea is not difficult to justify because none of the available measures of ovarian estrogen production is completely reliable, for reasons already described. A serum FSH contributes to a more confident clinical assessment of ovarian function and helps differentiate patients with common chronic anovulatory conditions from those with more severe hypogonadism who otherwise may go unrecognized and who require further specific evaluation, counseling, or treatment. For example, when evaluation suggests marginal levels of estrogen production (e.g., serum estradiol 30-40 pgmL or scant bleeding after a progestin challenge), a low serum FSH can identify those who merit further evaluation to exclude pituitary and hypothalamic disease, as described below. Conversely, in women with normal levels of estrogen production, a moderately elevated FSH level (e.g., 10-15 IU/L) can reveal a diminished ovarian reserve in women who may be afforded the chance to actively pursue their reproductive goals before the opportunity is lost, if alerted to their advanced reproductive aging. Like the other measures of ovarian function, the serum FSH concentration must be interpreted carefully, in its clinical context. FSH levels can fluctuate unpredictably, particularly during the years immediately preceding the menopause, regardless whether it occurs prematurely or at the usual age.

When evaluation reveals clear evidence of normal ovarian estrogen production and the serum FSH level also is normal, the diagnosis of chronic anovulation is established. Hyperprolactinemia is one of the most common causes of anovulation and amenorrhea and, although less common, thyroid disorders are easily identified and treated. Measurement of the serum prolactin and thyroid-stimulating hormone (TSH) concentrations are therefore justified in all women with amenorrhea. For efficiency, both can be measured along with the serum FSH and estradiol levels, at the outset of the evaluation. When all are normal, no further evaluation is required.

Besides thyroid and prolactin disorders, common and likely causes of chronic anovulation include PCOS, obesity, stress or exercise, and reproductive aging. In all but the last, anovulation can be attributed to a dysfunctional HPO axis in which gonadotropin secretion is sufficient to stimulate follicular development and estrogen production, but the system lacks the coordination required to achieve ovulation. Women with classical PCOS usually are easily recognized because they also exhibit signs of hyperandrogenism, unlike most women whose chronic anovulation relates solely to weight gain or obesity; the pathophysiology of the two disorders is complex and is discussed at length in Chapter 12 (PCOS) and Chapter 19 (obesity). Severe hirsutism or signs of virilization warrant additional specific evaluation to exclude enzyme deficiencies, androgen-secreting tumors, and Cushing’s syndrome, as described in Chapter 13. The diagnosis of anovulation relating to emotional, nutritional, or physical stress is made by exclusion, but often is suggested by the medical history and physical examination. The management of chronic anovulation associated with thyroid disorders and hyperprolactinemia is summarized here.

The newest ultra-sensitive TSH assays now in common use detect both primary hypothyroidism (elevated TSH) and primary hyperthyroidism (low TSH); either may result in chronic anovulation and amenorrhea. Although only a few patients presenting with amenorrhea will have a thyroid disorder that is not clinically apparent, their exclusion and
treatment are so simple that routine measurement of TSH is justified; a return of ovulatory cycles typically follows the restoration of normal thyroid hormone levels. Any abnormal TSH value should be confirmed and accompanied by measurement of serum thyroxine (tetra-iodothyronine; T4, or free T4) to better define the nature and extent of the thyroid disorder. An elevated TSH with a normal free T4 concentration indicates a subclinical hypothyroidism, best viewed as a compensated state wherein normal levels of T4 are maintained, but only under increased levels of pituitary stimulation. Although observation and periodic re-evaluation are reasonable in patients with subclinical hypothyroidism, because not all will develop frank hypothyroidism, treatment is warranted in those with menstrual dysfunction or infertility. In those with a low TSH and a normal free T4 level, serum tri-iodothyronine (T3) should be measured; an elevated T3 can identify hyperthyroidism that otherwise might escape detection. When the T3 also is normal, a subclinical hyperthyroidism is likely and should be followed carefully. On rare occasions, both TSH and free T4 levels are low, suggesting a secondary hypothyroidism of pituitary origin that requires additional evaluation to determine the cause and whether other pituitary functions also are affected.

A few women with hypothyroidism will develop a secondary hyperprolactinemia and even galactorrhea. The likelihood of hyperprolactinemia increases with the duration of hypothyroidism; galactorrhea is more common in young women with higher prolactin levels.⁵ The mechanism probably involves both the gradual depletion of hypothalamic dopamine (the putative prolactin-inhibiting factor) and constant stimulation of pituitary lactotropes by thyrotropin-releasing hormone (TRH), which may cause pituitary hypertrophy or hyperplasia and sometimes even enlargement or erosion of the sella turcica.^6,7 Although hormone levels rapidly normalize with appropriate treatment, the disappearance of breast secretions in those with galactorrhea is gradual and can take several months. Patients with primary hypothyroidism and hyperprolactinemia may present with either primary or secondary amenorrhea.⁸

Although premature follicular depletion is the cause in almost all cases, additional specific evaluation is indicated to exclude chromosomal and other genetic abnormalities and autoimmune disease that may have important potential health implications for the patient and other members of her family. The elements and purpose of the expanded diagnostic evaluation are summarized here. The known causes of ovarian failure and the disorders deserving specific consideration and exclusion are discussed at greater length in a later section of this chapter.

When there is no clear explanation for hypogonadotropic hypogonadism (e.g., significant physical, nutritional, or emotional stress) or for hyperprolactinemia (e.g., medications), further evaluation with imaging is indicated to exclude tumors and to help distinguish between pituitary and hypothalamic causes. The method of choice is MRI (with gadolinium contrast) because it is more sensitive and accurate than other imaging techniques for detection of abnormalities within and near the sella turcica.⁶³ MRI can demonstrate the nearby optic chiasm and also can detect blood, allowing hemorrhage and vascular abnormalities to be distinguished from other sellar mass lesions. Most sellar masses are pituitary adenomas, which account for 10% of all intracranial neoplasms. Other less common mass lesions in or near the sella include benign tumors (craniopharyngioma, hamartoma, meningioma), pituitary hyperplasia (thryotroph or gonadotroph hyperplasia due to long-standing primary hypothyroidism or gonadal failure), malignant tumors (germ cell, sarcoma, chordoma, carcinoma, lymphoma), metastases (lung, breast), cysts (Rathke’s cleft, arachnoid, dermoid), pituitary abscess, lymphocystic hypophpysitis, sarcoidosis, tuberculosis, and carotid arteriovenous fistula.

Although mass lesions are the most obvious abnormality to be excluded, other rare possibilities include Sheehan syndrome (pituitary infarct resulting from hypotension associated with postpartum hemorrhage), infiltrative hemosiderosis relating to frequent transfusions or hereditary hemochromatosis, traumatic brain injury,⁶⁴ and mutations in the GnRH receptor.⁶⁵

In the absence of any demonstrable mass lesion in the sellar region or relevant history suggesting another specific cause for pituitary damage, there is no need to perform any additional specific pituitary function tests. The clinical signs and symptoms associated with different types of functional and nonfunctional pituitary tumors and other specific pituitary causes of gonadotropin deficiency are discussed in a later section of this chapter.

The hymen is formed by invagination of the posterior wall of the urogenital sinus and usually ruptures spontaneously during the perinatal period. Although most cases of
imperforate hymen occur sporadically, reports of families with several affected members suggest that some cases may have a genetic and heritable cause.⁶⁹

Typically, patients with an imperforate hymen present at the expected time of menarche with complaints of cyclic perineal, pelvic or abdominal pressure or pain that results from the gradual accumulation of obstructed menstrual flow (cryptomenorrhea), and exhibit otherwise normal, symmetrical secondary sexual development for age. They also may present with acute urinary retention due to compression of the urethra and bladder by a grossly distended lower vagina.⁷⁰ The genital examination reveals no obvious vaginal orifice and a thin, often bulging, blue perineal membrane at the inferior limit of a palpable, fluctuant mass (hematocolpos).

The treatment of women with an imperforate hymen centers on providing relief of symptoms related to accumulated menstrual fluid and debris. Definitive surgery should be accomplished as soon as possible because delay can lead to infertility due to inflammatory changes and to the development of severe endometriosis. Surgical correction of an imperforate hymen is straightforward. The classical procedure is to make a simple cruciate incision in the hymen to the base of the hymeneal ring and to excise its central portion to allow drainage of sequestered menstrual fluid and subsequent normal menstruation. Alternatively, to avoid any risk of damage to the hymeneal ring (a sign of virginity important to some individuals and in some cultures), a sterile puncture can be made in the center of the distended membrane and enlarged to approximately 0.5 cm in diameter to allow insertion of a 16F Foley catheter. After thorough drainage of the vagina via irrigation with sterile saline, the catheter is left in place for approximately 2 weeks to allow further drainage from the vagina and upper genital tract. A single dose of prophylactic antibiotics is prudent and estrogen cream applied locally to the hymeneal ring helps to encourage re-epithelialization.⁷¹

A transverse vaginal septum results when the vaginal plate, formed from the fused sinovaginal bulbs, fails to break down or canalize during embryogenesis. As could be expected, girls with a transverse vaginal septum or cervical atresia, like those with an imperforate hymen, generally present at or soon after the age of expected menarche with complaints of cyclic pelvic or abdominal pain due to obstructed menses and exhibit symmetrical, age-appropriate secondary sexual development. Physical examination reveals a normal vaginal orifice, a shortened vagina of varying length, no visible cervix, and a palpable hematocolpos in the proximal vaginal segment above the obstruction and/or a pelvic mass resulting from hematometra and hematosalpinges. A Valsalva maneuver will cause distention at the intoitus in those with an imperforate hymen, but not in those with a transverse vaginal septum or cervical atresia, and can help to distinguish the two. Imaging is necessary to define the anatomy of the disorder but laboratory investigation generally is not required. Pelvic ultrasonography can reveal the level and extent of the hematocolpos and any associated hematometra or hematosalpinges. However, abdominal/pelvic MRI provides greater anatomical detail and is recommended to more clearly define the length of the atretic segment between the lower and upper vaginas,^72,73 information that is essential to planning surgical treatment. The temptation to insert a needle for diagnostic purposes must be resisted to avoid the risk of converting a hematocolpos into a pyocolpos. In rare instances, laparoscopy may be required to clarify the anatomy of the developmental anomaly. Transverse vaginal septum and cervical atresia may be accompanied by abnormalities of the upper reproductive tract, such as absent segments or atresia of the fallopian tubes or unilateral absence of the fallopian tube and ovary.⁷⁴ Unfortunately, chronic retrograde menstruation frequently results in pelvic endometriosis and adhesions, which can be severe.

In all instances, every effort should be made to incise and drain the sequestered menstrual fluid from below, at the level of the obstruction. Even in complicated circumstances, continuity of the lower genital tract usually can be achieved successfully. Operative excision of painful masses from above risks damage to the bladder, ureters, and rectum and unnecessarily removes distended but otherwise healthy reproductive organs. The surgical management of a transverse vaginal septum can be challenging, and because they also are infrequently encountered, often requires consultation with specialists having the necessary training and experience. Simply described, the procedure involves excision of the septum or dissection of the atretic segment and primary anastamosis of the margins of the lower and upper vaginal canals over the site of the defect. Atretic segments of greater length may require application of a graft to bridge the gap between the lower and uppper vaginas. Because septa that appear relatively thin by physical examination and MRI may be significantly larger after decompression of the proximal hematocolpos, preparations for surgery should consider the possibility that a graft may be required.

The best surgical management for rare women with cervical atresia is controversial. Ideally, the goal would be to create a functional vagina and to preserve the uterus and fertility, but experience has proven that such heroic efforts may be associated with serious postoperative complications such as peritonitis and sepsis, recurrent obstruction, and persistent infertility, prompting many to view hysterectomy as the best management option. However, conservative surgical treatment is reasonable to consider in selected individuals. The best candidates are those recognized early, before they develop severe pelvic endometriosis and adhesions, having a well-developed lower vagina,⁷⁵ although successful reconstruction and pregnancy can be achieved even in those who also require vaginoplasty.^76,77

The failure of müllerian development is a relatively common cause of primary amenorrhea, much more frequently encountered than AIS and second only to gonadal dysgenesis in prevalence;⁷⁸ in Finland, the incidence is approximately 1 in 5,000 newborn girls.⁷⁹ The cause is unknown. Although usually sporadic, some cases of müllerian agensis are associated with chromosomal translocations or occur in familial aggregates, suggesting a genetic basis for the disorder. Logically, müllerian agenesis might be attributed to an activating mutation in the gene encoding antimüllerian hormone (AMH) or its receptor, causing excess AMH activity. Inactivating mutations in these genes causing persistence of müllerian structures in otherwise normally virilized males have been described.^80,81 However, no activating mutations have been identified in patients with mullerian agenesis.⁸² The prevalence of a mutation in the galactose-1-phosphate uridyl transferase (GALT) gene (different from that associated with classical galactosemia) is increased in daughters with müllerian agenesis and their mothers.⁸³ The observation suggests that errors in fetal or maternal galactose metabolism resulting in increased intrauterine galactose exposure may have adverse effects on müllerian development, consistent with studies in rodents wherein a high galactose diet during pregnancy delayed vaginal opening in female offspring.⁸⁴ Given the relationship between classical galactosemia and premature ovarian failure, patients with müllerian agenesis who carry such a variant GALT gene mutation may be at increased risk for the same.

Patients with müllerian agenesis typically present in late adolescence or as young adults, well after menarche was expected, with primary amenorrhea as their only complaint. They exhibit normal, symmetrical breast and pubic hair development, no visible vagina, and have no symptoms or signs of crytomenorrhea because the rudimentary uteri contain no functional endometrium. However, in approximately 10%, functional islands of endometrium may result in a hematometra and symptoms of cyclic pain.^85,86 Two forms
of the disorder have been described. Type A is characterized by symmetrical, muscular, rudimentary uteri, and normal fallopian tubes, and Type B by asymmetrical rudimentary uteri and absent or hypoplastic fallopian tubes.⁸⁷ In the great majority of patients with müllerian agenesis, the ovaries are entirely normal, but one or both also may be undescended, hypoplastic, or associated with an inguinal hernia. Urologic anomalies are relatively common (15-40%), particularly in Type B müllerian agenesis, and include unilateral renal agenesis, ectopic or horseshoe kidney, and duplication of the collecting system(s).^85,86 Skeletal malformations involving the vertebrae, the ribs, or the pelvis are observed in 10-15% of patients; some of the more common abnormalities include hemivetebrae leading to scoliosis and the Klippel-Feil syndrome, characterized by a short neck, low hairline, limited range of motion, and sometimes pain and neurologic symptoms, all relating to one or more fused cervical vertebrae.

Although müllerian agenesis usually can be diagnosed by medical history and physical examination alone, additional evaluation is warranted to establish the diagnosis and to identify any of the urologic (renal ultrasonography) and skeletal anomalies (spinal X-rays) associated with the disorder. After puberty, a serum testosterone concentration in the normal female range effectively excludes AIS (discussed below). However, because patients with müllerian agenesis can exhibit characteristics similar to those observed in some types of male pseudohermaphroditism, a karyotype is justified and definitive. When examination raises suspicion that a uterine structure may be present, imaging is indicated. Ultrasonography may help to define the size and symmetry of any pelvic reproductive organs, but MRI is more accurate and is indicated when doubt remains.^88,89 Laparoscopy usually is not necessary for diagnosis of müllerian agenesis. Although imaging frequently does not agree completely with surgical observations, detailed knowledge of the pelvic anatomy is not often needed.⁹⁰ Surgery generally is indicated only in those with symptoms relating to hematometra, endometriosis, or a hernia into the inguinal canal.

The primary goal of treatment in women with müllerian agenesis—creation of a functional vagina—can be accomplished with a variety of methods, when the time is appropriate. In the large majority of cases, progressive vaginal dilation as originally described by Frank⁹¹ and later by others,⁹² is an appropriate and effective first choice. In motivated patients, the technique is highly successful and can create a functional vagina within 3 to 6 months.⁹³ The procedure involves applying pressure to the point of moderate discomfort for an interval of 20-30 minutes daily, using commercially available vaginal dilators. Initially, pressure is directed posteriorly, to create a shallow pouch. After approximately 2 weeks, pressure shifts to the usual axis of the vagina. After the desired depth is achieved, dilators of increasing diameter will expand the vagina to a functional size. A variation on the technique uses a tight-fitting garment to hold the dilator in place, maintaining pressure by leaning forward on a bicycle seat mounted on a stool, or even on a bicycle.⁹⁴

Operative treatment of women with müllerian agenesis generally can be reserved for those who are unable or unwilling to dedicate themselves to a program of progressive vaginal dilation and for those in whom earnest efforts fail. The traditional McIndoe procedure for surgical creation of a neovagina involves dissection of the rectovaginal space and placement of a skin graft, held in place with a soft mold until the graft becomes established.⁹⁵ Subsequently, regular intercourse or vaginal dilation must be maintained to avoid risk of fibrosis and loss of function. The alternative Vecchietti operation involved internal stretching of the vaginal dimple after surgical abdominal and vaginal dissection of the vesicorectal space.⁹⁶ A modification of the Vecchietti operation, performed laparoscopically, has emerged as an attractive and effective option for surgical creation of a neovagina.^97,98 The procedure employs a specially designed system including a springloaded traction device, positioned on the abdomen, connected to the tip of a dilator, positioned at the introitus, via paired threads introduced with a needle inserted through small incisions in the lower abdomen and guided beneath the peritoneum to penetrate
the introitus in the midline. Tension on the threads is adjusted to maintain traction on the dilator, which is drawn upward gradually, invaginating the tissue to produce a neovagina 7-8 cm in depth over an interval of 7-10 days. Thereafter, standard vaginal dilators are employed to further extend and expand the vagina to functional dimensions over a period of a few weeks. Evidence indicates that the procedure results in a quality of sexual experience comparable to that in a sample of healthy, age-matched women of equivalent cultural and social status.⁹⁸

Reassurance and support are important elements of the management of vaginal/müllerian agenesis. Affected women should be counseled that although they are infertile, normal sexual function can be expected and that genetic offspring can be achieved by in vitro fertilization (IVF) using oocytes retrieved from their own normal ovaries and the sperm of their chosen partner, with subsequent transfer of embryos to a gestational surrogate.^99,100 An analysis of 34 live births resulting from IVF in 58 women with müllerian agenesis revealed no evidence to suggest a dominant pattern of inheritance and demonstrated that IVF, combined with gestational surrogacy, is a realstic option for patients with the disorder.^100,101

Complete AIS (testicular feminization) is a form of male pseudohermaphroditism, the term referring to the gonadal sex (male) and the contrasting phenotype (female). The disorder, discussed in greater detail in Chapter 9 as a cause of abnormal sexual development, is the third most common cause of primary amenorrhea, after gonadal dysgenesis and müllerian agenesis. Patients with AIS have a normal male karyotype (46,XY) and testes that produce both testosterone and AMH. However, an inactivating mutation in the gene encoding the intracellular androgen receptor (located on the long arm of the X chromosome, Xq) results in an end organ insensitivity to androgen actions that prevents normal masculinization of the internal and external genitalia during embryonic development. Consequently, the external genitalia are those of a female (absent androgen action), the cervix and uterus are absent (due to normal AMH action), and the vagina is short and ends blindly (derived only from the urogenital sinus).

Patients with complete AIS appear normal at birth. Growth and development during childhood also are generally normal, although overall height usually is above average and the body habitus somewhat eunuchoid (long arms, large hands and feet). At puberty, the breasts develop, driven by estrogen derived from the peripheral conversion of high circulating testosterone levels, unopposed by the actions of androgen. The breasts may become relatively large and have subtle abnormalities; lacking the actions of progesterone, they have little glandular tissue, small nipples and pale areolae. The labia minora usually are underdeveloped and the vagina is short and ends blindly. Pubic and axillary hair does not develop, due to the absence of androgen stimulation. The testes may be intra-abdominal, but often are partially descended; more than half of patients with complete AIS have an inguinal hernia. The testes frequently are palpable in the inguinal canals, most commonly at the level of the external inguinal ring. They generally resemble any cryptorchid testes but may be nodular. After puberty, the testes contain immature seminiferous tubules lined by immature germ cells and Sertoli cells, with no evidence of spermatogenesis.

Patients with complete AIS most commonly present after the age of puberty in late adolescence or as young adults with primary amenorrhea. They exhibit asymmetrical secondary sexual development (breast development with absent or scant pubic hair), a short vagina with no visible cervix, and have no other symptoms or complaints. They also may be recognized at birth or in childhood when they may present with an inguinal mass or hernia,
particularly when the disorder is reasonably suspected because other family members such as a sister or maternal aunt are affected. Diagnosis usually is not difficult. Patients with complete AIS generally are easily distinguished from those with müllerian agenesis by the absence of pubic and axillary hair, and from those with an imperforate hymen or transverse vaginal septum by the absence of a uterus and symptoms relating to obstructed menstrual flow. In some cases, the presence of some pubic hair, due to incomplete penetrance, can be confusing or misleading. A serum testosterone concentration easily distinguishes patients with AIS because levels are normal or modestly elevated above the range observed in normal males and well above the normal range for females. Serum LH levels also are elevated, reflecting androgen insensitivity at the hypothalamic-pituitary level. A karyotype (46,XY) firmly establishes the diagnosis.

In rare cases of incomplete androgen insensitivity, the sensitivity to androgens is greater. Consequently, pubic hair growth may accompany breast development and the clitoris may enlarge, or a phallus even may be present.¹⁰² In other rare individuals having a deficiency of the enzyme 17b-hydroxysteroid dehydrogenase (type 3), which catalyzes the conversion of androstenedione to testosterone in testicular Leydig cells, the clinical presentation may be similar, but due to impaired testosterone production rather than abnormalities in the androgen receptor. When necessary, the two disorders can be differentiated by molecular analysis of the genes encoding the androgen receptor (AR) and the enzyme (17HSDB3).^103,104

The treatment of patients with complete AIS has two major components, one focusing on creation of a functional vagina, and another relating to the risk for developing malignancy in the cryptorchid testes. In patients with AIS, the options for creation of a neovagina are the same as in those with müllerian agenesis—progressive vaginal dilation and vaginoplasty. The short but distinct vagina observed in most patients speeds the progress of efforts at vaginal dilation. Good results also can be expected with surgical treatment, when necessary.¹⁰⁵ Gonadectomy is indicated because the incidence of neoplasia in cryptorchid testes is relatively high. In one early series of 50 cases, 11 malignancies, 15 adenomas, and 10 benign cysts were observed: a 22% incidence of malignancy and a 52% overall incidence of neoplasia.¹⁰⁶ More recent series suggest a lower 5-10% overall incidence of gonadal tumors.^{50,102,107,108} Whereas gonadectomy is recommended at time of diagnosis in other intersex states such as XY gonadal dysgenesis (Swyer syndrome), it is better delayed in those with AIS, for two reasons. First, the smooth pubertal development that results from endogenous hormone production is difficult to achieve with exogenous hormone treatment, and second, gonadal tumors develop less often in patients with AIS and rarely before puberty. Therefore, gonadectomy and hormone therapy (physiologic estrogen treatment) generally are best postponed until after pubertal development is complete, by approximately age 16-18. Complete AIS is the only exception to the rule that gonads with a Y chromosome should be removed as soon as a diagnosis is made. Gonadectomy usually can be accomplished endoscopically with relative ease when the testes reside within the abdomen, and via inguinal incisions when they are partially descended.^109,110 In patients with the incomplete form of AIS, surgery should not be postponed because prompt gonadectomy will prevent further unwanted virilization.

In years now past, conventional wisdom warned against unthinking and “needless” disclosure of the true gonadal and chromosomal sex to patients with complete AIS for fear of undermining gender identity, but that attitude has changed. Although infertile, patients with AIS have a completely female gender identity that should be reinforced, not questioned. We strongly advocate combining a truthful education with appropriate psychological counseling for both patient and parents. Patients want, deserve, and appreciate a fuller understanding of themselves and the disorder. Moreover, because the public now has ready access to sophisticated medical information, secrecy also is no longer a practical possibility. One excellent resource is the Androgen Insensitivity Syndrome Support Group, headquartered in the United Kingdom (http://www.aissg.org/).

Severe cervical stenosis with complete outflow obstruction is a rare complication of cervical conization procedures or other surgical treatments for cervical intraepithelial neoplasia. When cervical stenosis causes symptoms, worsening dysmenorrhea or prolonged light staining or spotting after menses are the most common complaints; amenorrhea is a rare occurrence. Compared to other methods for limiting blood loss during conization, elective suturing appears to increase the risk for subsequent cervical stenosis and amenorrhea.¹¹¹ In women with amenorrhea who have had a previous conization or other cervical surgery or ablative treatment, simple uterine sounding will establish the diagnosis of cervical stenosis and transvaginal ultrasonography will reveal any associated hematometra. The treatment for cervical stenosis is careful dilation, ideally performed under ultrasound guidance. Temporary placement of a urinary or specialized balloon catheter for an interval of approximately 2 weeks provides ongoing drainage of the uterine cavity and may help to prevent recurrence.¹¹²

Asherman syndrome, first described by Joseph Asherman in 1948 and called “amenorrhoea traumatica,”¹¹³ results from intrauterine adhesions that obstruct or obliterate the uterine cavity, as a consequence of trauma. Risk for developing intrauterine adhesions is increased by inflammation, as may result from endometritis or retained products of conception, and when the endometrium is relatively thin and inactive, as it is during the postpartum period. Consequently, most cases arise in close temporal proximity to a pregnancy and are associated with surgical trauma, primarily curettage.¹¹⁴ In the original series of 29 cases described by Asherman, 11 had a previous postpartum hemorrhage, 15 a spontaneous abortion, 2 an elective termination, and 1 had a hydatidiform mole.¹¹³ Asherman syndrome is an uncommon but recognized complication of cesarean section, abdominal or hysteroscopic myomectomy or metroplasy, and uterine artery embolization,¹¹⁵ Elective endometrial ablation procedures for the management of menorrhagia frequently result in amenorrhea, by intent, but most do not have intrauterine adhesions. End organ endometrial damage and adhesions also may result from intrauterine infections such as tuberculosis and schistosomiasis, which are rare in the United States but not in other regions of the world.^116,117

Although Asherman syndrome from any cause may result in amenorrhea, most women with intrauterine adhesions present with dysmenorrhea, hypomenorrhea, infertility, or recurrent pregnancy loss, rather than amenorrhea. The diagnosis of Asherman syndrome is based primarily on a high index of suspicion, based on history. In women whose history suggests the possibility, scant or no withdrawal bleeding after sequential treatment with exogenous estrogen (e.g., conjugated equine estrogens 1.25 mg daily for 21 days) and progestin (e.g., medroxyprogesterone acetate 10 mg daily for the last 5-7 days) can demonstrate end organ endometrial failure and corroborate the clinical suspicion.

However, some form of imaging ultimately is required to establish the diagnosis. Transvaginal or transabdominal ultrasonography may reveal a hematometra, but such findings are surprisingly rare. Sonohysterography or hysterosalpingography (HSG) provide more specific information regarding the location and extent of adhesions that partially or completely obliterate or obstruct the endometrial cavity or the cervical canal,¹¹⁸ and hysteroscopy is definitive. A variety of classification schemes have been proposed to describe the extent of intrauterine adhesions and to predict treatment outcomes,^{119,120,121,122,123} and ¹²⁴ but none has been validated. Diagnosis of genital tuberculosis is made by endometrial biopsy (histopathology or culture) or with a nucleic acid-based test performed on an endometrial aspirate. Diagnosis of shistosomiasis is made by identifying the eggs of the parasite in urine, feces, rectal scrapings, menstrual discharge, or the endometrium.

Operative hysteroscopy is the primary method for treatment of intrauterine adhesions that may be lysed by scissors, electrodissection, or with a laser; most prefer sharp dissection, which may have less risk for causing further injury. Simultaneous laparoscopy or transabdominal ultrasonography provides useful guidance when dense scar tissue makes it difficult to enter the uterine cavity and can help to maintain orientation in a grossly distorted cavity, reducing the risk of uterine perforation. Most now advocate insertion of an intrauterine balloon catheter (left in place for approximately 7-10 days) after adhesiolysis to keep the walls of the uterine cavity separated during healing and decrease the risk of recurrence.¹²⁵ Treatment with a broad-spectrum antibiotic (e.g., doxycycline 100 mg twice daily) and a non-steroidal antiinflammatory drug help to minimize risk of infection and uterine cramping while the catheter remains in place. High dose exogenous estrogen treatment (2.5 mg conjugated equine estrogens 2-3 times daily, or its equivalent) for approximately 4 weeks after surgery generally is recommended to encourage rapid endometrial re-epithelialization and proliferation; treatment with a progestin during the final week, if followed by menses, demonstrates a return of function. Despite these measures, recurrence rates are relatively high, ranging from 20% to over 60% in severe cases,¹²⁶ and repeated procedures often are required to restore a normal uterine cavity.¹²⁰ The surgical outcome can be assessed by postoperative HSG or by early “second-look” office hysteroscopy, which also provides the means to lyse any early recurring adhesions when still filmy.¹¹⁴ Menstrual function can be restored in most cases (52-88%),¹²⁶ and among infertile women, live birth rates after hysteroscopic adhesiolysis generally have ranged between 25 and 35%.¹²⁷ As might be expected, outcomes tend to correlate with the severity of adhesions.^{119,120,126,128} In those who do achieve pregnancy, the risks of preterm labor, placenta accreta, placenta previa, and postpartum hemorrhage are increased.¹¹⁴

Gonadal dysgenesis is defined as an incomplete or defective formation of the gonads, resulting from a disturbance in germ cell migration or organization, caused by structural or numerial sex chromosome abnormalities or mutations in the genes involved in formation of
the urogenital ridge and sexual differentiation of the bipotential gonad. Gonadal dysgenesis is among the most common causes of primary amenorrhea (approximately 30-40%). Due to the absence of ovarian follicles or their accelerated depletion during embryogenesis or the first few years of life, the gonads contain only stroma and appear as fibrous streaks. The large majority of patients with gonadal dysgenesis have an obvious abnormality involving an X chromosome. Aproximately 25% of affected individuals have a normal 46,XX karyotype and may harbor a more subtle abnormality invoving one or more specific genes on the X chromosome that are required for normal ovarian function; some with 46,XX gonadal dysgenesis also have neurosensory deafness, the combination known as Perrault syndrome. By far, the most common form of gonadal dysgenesis is Turner syndrome.

Turner syndrome is a well known and thoroughly studied disorder, classically associated with a 45,X karyotype, but also with an assortment of other structural X chromosome abnormalities (deletions, ring and iso-chromosomes), any of which may be present in all or only in some of the cells of the body (mosaicism), depending on the stage of embryonic development at the time they arise. The disorder is discussed thoroughly in Chapter 9, as a cause of abnormal sexual development, and is more briefly summaried here.

The classical phenotype of Turner syndrome includes short stature, absent sexual development, a webbed neck, low set ears and posterior hairline, widely-spaced nipples (“shield chest”) short fourth metacarpals, and an increased carrying angle at the elbow (“cubitus valgus”). Evidence indicates that the specific phenotype of patients with Turner syndrome relates, in part, to the parental origin of their X chromosome; most with a 45,X karyotype retain the maternal X.¹²⁹

If not recognized by phenotype or poor growth during childhood, patients with Turner syndrome generally present at or near the time of expected puberty with primary amenorrhea and absent secondary sexual development. The diagnosis of Turner syndrome generally can be made easily, based on the phenotype and findings of hypergonadotropic hypogonadism. A karyotype is definitive, and specifically indicated, in part because it may reveal a cell line containing a Y chromosome otherwise not suspected or identified (e.g., 45,X/46,XY); approximately 5% of women with Turner syndrome have a karyotype containing all or part of a Y chromosome.¹³⁰ Futher analysis with fluorescence in situ hybridization (FISH) using one or more probes specific for segments of the Y chromosome will identify another 5% having occult Y chromosome material.^130,131 Whereas it is important to identify a Y chromosome because affected individuals are at significant increased risk for developing gonadoblastoma (20-30%), that risk appears lower (5-10%) in women with Turner syndrome, and limited to those having detectable Y chromosome on their karyotype. FISH analysis is most clearly indicated for those exhibiting any evidence of virilization or having a chromosomal fragment of uncertain origin (discussed in chapter 9).¹³²

Mosaicism in women with Turner syndrome has important clinical implications besides those relating to a cell line containing a Y chromosome. In those with a mosaic 46,XX cell line (e.g., 45,X/46,XX), the gonad may contain functional ovarian cortical tissue, resulting in some degree of sexual development, or even menses and the possibility of pregnancy. Approximately 15% of patients with Turner syndrome begin but do not complete pubertal development and approximately 5% complete puberty and begin menstruation.¹³³ As might be expected, the phenotype varies, with some appearing normal and attaining normal stature before experiencing ovarian failure when the limited supply of follicles is exhausted. Natural pregnancies do occur in women with Turner syndrome, but they are rare
and associated with a relatively high risk for sex chromosome aneuploidy and spontaneous abortion.

Women with Turner syndrome may have a wide variety of medical problems having health implications as or more important than those relating to or resulting directly from hypogonadism.¹³² Approximately one-third has cardiovascular anomalies, including a bicuspid aortic valve, coarctation of the aorta, mitral valve prolapse, and aortic aneurysm. Renal anomalies also are common and include horsehoe kidney, unilateral renal agenesis or pelvic kidney, rotational abnormalities, and partial or complete duplication of the collecting system(s). Autoimmune disorders are common in Turner syndrome and include thyroiditis, type 1 diabetes, autoimmune hepatitis and thrombocytopenia, and celiac disase. Hearing loss also is common.^134,135 Consequently, additional and periodic medical evaluation is indicated and should include the following¹³²:

Average intellectual performance is within the normal range,¹³⁶ although the prevalence of attention-deficit/hyperactivity disorder (ADHD) is increased in girls with Turner syndrome.¹³⁷ Overall mortality is increased approximately 3-fold and relates primarily to circulatory disease (e.g., hypertension), diabetes, liver and renal disease.¹³⁸ Overall cancer risks in women with Turner syndrome are similar to those in the general population, but the incidence of CNS tumors, bladder cancer, and endometrial cancer may be increased, and the risk of breast cancer is decreased.¹³⁹ With early diagnosis and treatment with growth hormone (GH), a final height greater than 150 cm (59 inches) can be achieved in most patients with Turner syndrome. The most important determinants of final height are the dose of GH and the duration of treatment before estrogen therapy begins. Treatment with GH generally should begin as soon as height falls below the fifth percentile of normal female growth and must be individualized, according to response.¹³²

Treatment with estrogen must be timed carefully, with the goals of minimizing its adverse effects on growth and adult height and inducing puberty at an approximately normal age. Ideally, estrogen therapy should begin no later than age 15 and not before age 12 when growth is a priority, unless height already has been maximized. Estrogen therapy should begin at a low dose (e.g., 0.25-0.5 mg micronized estradiol or its equivalent), increasing gradually at intervals of 3-6 months according to response (Tanner stage, bone age), with the goal of completing sexual maturation over a period of 2-3 years. When vaginal bleeding first occurs, or after 12-24 months of estrogen therapy, a progestin (e.g., medroxyprogesterone acetate) should be added to the treatment regimen to complete development, prevent dysfunctional bleeding, and to protect the endometrium from the effects of unopposed estrogen.¹³²

Oocyte donation offers the possibility of pregnancy to patients with Turner syndrome, but the cardiovascular demands of pregnancy pose unique and potentially serious risks that must be carefully considered. The risk of death during pregnancy is increased as much as 100-fold, primarily due to complications of aortic dissection or rupture. Risk is greatest for those with preexisting abnormalities such as a bicuspid aortic valve or a dilated aortic root, but even those without such findings remain at risk. Consequently, Turner syndrome generally should be regarded as a relative contraindication to pregnancy. Those expressing
serious interest in oocyte donation must receive thorough evaluation and counseling, and those having any significant cardiac abnormality should be strongly discouraged.¹⁴⁰

Swyer syndrome is a distinctly different and less common form of gonadal dysgenesis, characterized by a 46,XY karyotype. Despite the presence of a Y chromosome, the phenotype is female because the dysgenetic (streak) gonads produce neither AMH nor androgens. Consequently, the vagina, cervix, uterus, and fallopian tubes develop normally and the internal and external genitalia do not masculinize.¹⁴¹ In at least 10-15% of affected individuals, a mutation of the SRY gene (Sex-determining Region of the Y chromosome; located on the short arm, Yp11.3) is the cause.¹⁴² In the remainder, no cause can be determined, although mutations in SRY regulatory elements or in other genes involved in the testis-determining pathway have been implicated (SF1, SOX9, WT1, CMRT1).^143,144 and ¹⁴⁵

Patients with Swyer syndrome generally present after the expected time of puberty with delayed sexual maturation and primary amenorrhea. The presence of pubic hair reflects a normal adrenarche. Evaluation reveals hypergonadotropic hypogonadism, prompting a karyotype that establishes the diagnosis. Gonadectomy is indicated soon after diagnosis due the significant risk for malignant transformation in occult testicular elements (20-30%). Gonadoblastoma is a premalignant germ cell tumor, unique to intersex states like Swyer syndrome, and may contain or give rise to other highly malignant tumors, including dysgerminoma, endodermal sinus tumor, embryonal and choriocarcinomas; evidence suggests they originate from clonal expansion of surviving germ cells in areas of undifferentiated gonadal tissue.¹⁴⁶

Patients with Swyer syndrome exhibit normal growth and intellectual development, have no increased prevalence of any specific medical problems, and require no specific monitoring or treatment beyond that relating to hormone therapy aimed at inducing sexual maturation. The same sequential sex steroid treatment regimen described above for patients with Turner syndrome can be applied successfully in those with Swyer syndrome. Pregnancy, achieved with in vitro fertilization using donor oocytes, is a realistic expectation and has not been associated with any specific risks or complications.¹⁴⁷

Some individuals with primary amenorrhea and gonadal dysgenesis (streak gonads) have a normal 46,XX karyotype, providing indirect evidence that autosomal genes also play a critical role in ovarian differentiation. Affected women are normal in stature and, in most cases, have no apparent somatic anomalies. A wide variety of candidate genes has been identified, primarily via experiments involving murine knock-out models, including several that encode DNA and RNA binding proteins and transcription factors expressed during oogenesis.¹⁴⁸

Premature ovarian failure (POF), traditionally defined as hypergonadotropic hypogonadism and amenorrhea arising before the age of 40, is a heterogeneous disorder that varies widely in cause and phenotype. Whereas the term POF is well-entrenched in the medical literature,
an alternative term—“premature ovarian insufficiency”—has been proposed to more accurately reflect the continuum of decreased ovarian function observed in affected women,¹⁴⁹ acknowledging that many exhibit intermittent ovarian function and ovulation and that 5-10% may conceive and deliver a pregnancy.^150,151

Premature ovarian failure (POF) generally results in secondary amenorrhea at some time after puberty is completed, but also may occur at any time before menarche and is distinguished from gonadal dysgenesis on the basis of ovarian morphology and histology; instead of streak gonads, the ovaries more closely resemble those of postmenopausal women. Approximately 1% of women will develop POF before the age of 40 years. In a cross-sectional survey of women aged 40-55 conducted at seven sites in the United States to determine eligibility for a community-based, multi-ethnic longitudinal sudy of the perimenopause (The Study of Women Across the Nation, SWAN), premature menopause was reported by 1% of Caucasians, 1.4% of African Americans and Hispanics, 0.5% of Chinese, and 0.1% of Japanese women.¹⁵²

Important known causes of POF include numerical and structural chromosomal abnormalities, fragile X (FMR1) premutations, autoimmune disorders, radiation therapy, and chemotherapy. Whereas history alone can identify the latter two, the first three merit specific consideration and exclusion. In these and other rare disorders associated with ovarian failure such as galactosemia, the basic pathophysiology involves accelerated follicular atresia. In other rare genetic disorders, mutations in genes encoding intraovarian regulators, steroidogenic enzymes, gonadotropins, or their receptors result in impaired or abnormal follicular development, but the end result is much the same—functional ovarian failure. In most women with POF, a specific cause cannot be identified, but evidence implicating a number of genetic factors is growing rapidly.¹⁴⁸