3002 PART 12 Endocrinology and Metabolism

and sometimes breast development, cyclical hematuria, and/or phallic

development at puberty. Progressive regression of the ovarian and/

or testicular component can occur over time. Gender identity varies

in OTDSD but often aligns with assigned sex. Risk of GCC is also

elevated in OTDSD (~3%) and may occur in the ovarian or testicular

component. Infertility is typical (especially in 46,XX testes with no Y

chromosome), but births have occurred via ovum or sperm from individuals with other forms of OTDSD.

DISORDERS OF GONADAL AND

PHENOTYPIC SEX

Disorders of gonadal and phenotypic sex can result in reduced androgen production or action in individuals with a 46,XY karyotype (46,XY

DSD) or excess androgen production in individuals with a 46,XX karyotype (46,XX DSD) (Table 390-1). These conditions cover a spectrum

of phenotypes ranging from phenotypic females with a Y chromosome

to phenotypic males with a 46,XX karyotype to individuals with atypical genitalia. Karyotype is a useful starting investigation for diagnosis

but does not define an individual’s gender.

■ 46,XY DSD

Underandrogenization of the 46,XY fetus reflects defects in androgen

production or action. It can result from disorders of testis development,

defects of androgen synthesis, or resistance to testosterone and DHT

(Table 390-1).

Disorders of Testis Development • TESTICULAR DYSGENESIS

Complete gonadal dysgenesis (CGD, Swyer’s syndrome) is associated

with streak gonads, müllerian structures (due to insufficient AMH/

MIS secretion), and a complete absence of androgenization. Phenotypic females with this condition often present because of absent

pubertal development and are found to have a 46,XY karyotype. Serum

sex steroids, AMH/MIS, and inhibin B are low, and LH and FSH are

elevated. Individuals with CGD typically identify as female. The risk of

GCC is high, and intraabdominal gonads should be removed. In contrast, patients with partial gonadal dysgenesis (PGD, dysgenetic testes)

may produce enough MIS to regress the uterus and sufficient testosterone for partial androgenization and, therefore, usually present in the

newborn period with atypical genitalia, highlighting the spectrum of

features that are typically seen with many DSDs.

Testicular dysgenesis can result from disruption of testis-promoting

genes (e.g., WT1, SF1, SRY, SOX9, MAP3K1, DHH, DHX37, and others)

or, rarely, duplication of chromosomal loci containing “antitestis” genes

(e.g., DAX1) (Table 390-4). Among these, deletions or mutations of

SRY and heterozygous mutations of SF1 (NR5A1) or DHX37 appear to

be most common but still account collectively for <30% of cases. Associated clinical features may be present, reflecting additional functional

roles for these genes. For example, renal dysfunction occurs in patients

with specific WT1 mutations (Denys-Drash and Frasier’s syndromes),

primary adrenal insufficiency occurs in a minority of patients with

disruption of SF1, and severe cartilage abnormalities (campomelic

dysplasia) are the predominant clinical feature of pathogenic variants

in SOX9. A family history of DSD, hypospadias, infertility, or early

menopause is important because variations in some genes (e.g., SF1/

NR5A1, SOX8) can be associated with a range of reproductive phenotypes. SF1 variants are sometimes inherited from a mother in a sexlimited dominant manner (which can mimic X-linked inheritance),

and a woman may later develop primary ovarian insufficiency because

of the effect of SF1 on the ovary. Gender identity can be variable in

PGD. Dysgenetic testes have an increased risk of GCC. For descended

testes, monitoring via physical examination is appropriate. If testes are

intraabdominal and not able to be brought down, they may be removed

to prevent GCC (risk up to 35% if intraabdominal). Dysgenetic testes

may or may not produce sufficient testosterone for puberty. In those

who identify as male, testosterone replacement may be needed. In

those who identify as female, estrogen replacement will be needed for

female-typical pubertal development and ongoing sex steroid requirements. Absent (vanishing) testis syndrome (bilateral anorchia) reflects

regression of the testis during development. The absence of müllerian

structures indicates adequate secretion of AMH early in utero. Usually,

androgenization of the external genitalia is normal. The etiology is

often unknown but sometimes associated with pathogenic variants

in DHX37. These individuals can be offered testicular prostheses and

androgen replacement in adolescence and typically identify as male.

Disorders of Androgen Synthesis Defects in the pathway that

regulates androgen synthesis (Fig. 390-4) cause underandrogenization

of the 46,XY fetus (Table 390-1). Müllerian regression is unaffected

because Sertoli cell function is preserved, and no uterus is found.

These conditions can present with a spectrum of genital appearances,

ranging from female-typical external genitalia or clitoromegaly in some

individuals to penoscrotal hypospadias or a small phallus in others.

LH RECEPTOR Mutations in the LH receptor (LHCGR) cause Leydig

cell hypoplasia and androgen deficiency, due to impaired actions of

human chorionic gonadotropin in utero and LH late in gestation and

during the neonatal period. As a result, testosterone and DHT synthesis are reduced.

STEROIDOGENIC ENZYME PATHWAYS Mutations in steroidogenic

acute regulatory protein (StAR) and CYP11A1 affect both adrenal and

gonadal steroidogenesis (Fig. 390-4) (Chap. 386). Affected individuals

(46,XY) usually have severe early-onset salt-losing adrenal failure and a

female phenotype, although later-onset milder variants are increasingly

reported. Defects in 3β-hydroxysteroid dehydrogenase type 2 (HSD3B2)

also cause adrenal insufficiency in severe cases, but the accumulation

of dehydroepiandrosterone (DHEA) has a mild androgenizing effect,

resulting in atypical genitalia or hypospadias. Salt loss occurs in

many but not all children. Patients with CAH due to 17α-hydroxylase

(CYP17A1) deficiency have variable underandrogenization and develop

hypertension and hypokalemia due to the potent salt-retaining effects

of corticosterone and 11-deoxycorticosterone. Patients with complete

loss of 17α-hydroxylase function often present as phenotypic females

who do not enter puberty and are found to have inguinal testes and

hypertension in adolescence. Some mutations in CYP17 selectively

impair 17,20-lyase activity without altering 17α-hydroxylase activity,

leading to underandrogenization without mineralocorticoid excess and

hypertension. Disruption of the coenzyme, cytochrome b5 (CYB5A),

can present similarly, and methemoglobinemia is usually present.

Mutations in P450 oxidoreductase (POR) affect multiple steroidogenic

enzymes, leading to reduced androgen production and a biochemical

pattern of apparent combined 21-hydroxylase and 17α-hydroxylase

deficiency, sometimes with skeletal abnormalities (Antley-Bixler craniosynostosis). Defects in 17β-hydroxysteroid dehydrogenase type 3

(HSD17B3) and 5α-reductase type 2 (SRD5A2) interfere with the

synthesis of testosterone and DHT, respectively. These conditions

are characterized by minimal or absent androgenization in utero, but

some phallic development can occur during adolescence due to the

action of other enzyme isoforms. Individuals with 5α-reductase type 2

deficiency have normal wolffian structures and usually do not develop

breast tissue. At puberty, the increase in testosterone induces muscle

mass and other virilizing features despite DHT deficiency. DHT gel

can improve prepubertal phallic growth in patients raised as male.

Prevention of testosterone exposure (by gonadectomy or pubertal

suppression) in adolescence and estrogen replacement at puberty can

be considered in individuals who identify as female. Disruption of

alternative pathways to fetal DHT production might also present with

46,XY DSD (AKR1C2/AKR1C4).

Disorders of Androgen Action • androgen insensitivity

syndrome Pathogenic variants in the androgen receptor cause

resistance to androgen (testosterone, DHT) action or the androgen

insensitivity syndrome (AIS). AIS is a spectrum of disorders that affects

at least 1 in 100,000 46,XY individuals. Because the androgen receptor

is X-linked, only 46,XY offspring are affected. The condition is usually

inherited from a mother who carries the sequence variant but can also

arise de novo. XY individuals with complete AIS (formerly called testicular feminization syndrome) have a female phenotype, normal breast

development (due to aromatization of testosterone), a short vagina but

no uterus (because AMH/MIS production is normal), scanty pubic

3003 Sex Development CHAPTER 390

and axillary hair, and typically a female gender identity and sex role

behavior. Gonadotropins and testosterone levels can be low, normal,

or elevated, depending on the degree of androgen resistance and the

contribution of estradiol to feedback inhibition of the hypothalamicpituitary-gonadal axis. AMH/MIS levels in childhood are normal or

high. CAIS sometimes presents as inguinal hernias (containing testes)

in childhood or more often with primary amenorrhea in late adolescence. In the past, gonadectomy was recommended in childhood, but

due to the low risk of malignancy (~2%), increasingly this is delayed,

and gonads are left in situ until breast development is complete.

Subsequently, the adolescent or young adult should be counseled

about the risk of malignancy and the option for gonadectomy (with

estrogen replacement), especially because early detection of premalignant changes by imaging or biomarkers is currently not possible. The

use of graded dilators in adolescence is often sufficient to dilate the

vagina for sexual activity.

Partial AIS (Reifenstein’s syndrome) results from androgen receptor

mutations that maintain residual function. Patients often present in

infancy with penoscrotal hypospadias and undescended testes and

with gynecomastia at the time of puberty. Gender identity can be variable. At puberty, testes produce testosterone with variable phenotypic

development. For those who identify as male, high-dose testosterone

has been given to support development if puberty does not progress,

but long-term data are limited. For those raised as female, testosterone

TABLE 390-4 Selected Genetic Causes of 46,XY Disorders of Sex Development (DSDs)

GENE INHERITANCE GONAD UTERUS EXTERNAL GENITALIA ASSOCIATED FEATURES

Disorders of Testis Development

WT1 AD Dysgenetic testis +/– Female or ambiguous Wilms’ tumor, renal abnormalities, gonadal tumors (WAGR,

Denys-Drash and Frasier’s syndromes)

SF1/NR5A1 AR/AD (SL) Dysgenetic testis/Leydig

dysfunction

+/– Female, ambiguous or

male

Primary adrenal failure (rare); hyposplenia (rare); primary

ovarian insufficiency in female (46,XX) relatives

SRY Y Dysgenetic testis or

ovotestis

+/– Female or ambiguous

SOX9 AD Dysgenetic testis or

ovotestis

+/– Female or ambiguous Campomelic dysplasia

MAP3K1 AD (SL) Dysgenetic testis +/– Female or ambiguous

DHX37 AD Dysgenetic testis +/– Female, ambiguous or

male

Testicular regression syndrome

DHH AR Dysgenetic testis/Leydig

dysfunction

+ Female Minifascicular neuropathy

Other causes of testicular dysgenesis include: DMRT1, CBX2, SOX8, ZNRF3, GATA4, and ZFPM2 (congenital heart disease); ARX (X-linked lissencephaly); TSPYL1

(sudden infant death); MYRF (diaphragmatic hernia); ESR2/NR3A2, SAMD9 (MIRAGE syndrome); MAMLD1, dupXp21, dup1p35, del9p24, del10q23

Disorders of Androgen Synthesis

LHCGR AR Testis – Female, ambiguous or

micropenis

Leydig cell hypoplasia

DHCR7 AR Testis – Variable Smith-Lemli-Opitz syndrome: coarse facies, second-third toe

syndactyly, failure to thrive, developmental delay, cardiac and

visceral abnormalities

STAR AR Testis – Female or ambiguous Congenital lipoid adrenal hyperplasia (primary adrenal

insufficiency)

CYP11A1 AR Testis – Female or ambiguous Primary adrenal insufficiency

HSD3B2 AR Testis – Ambiguous CAH, primary adrenal insufficiency ± salt loss, partial

androgenization due to ↑ DHEA

CYP17A1 AR Testis – Female or ambiguous CAH, hypertension due to ↑ corticosterone and

11-deoxycorticosterone, except in isolated 17,20-lyase

deficiency

CYB5A AR Testis – Ambiguous Apparent isolated 17,20-lyase deficiency; methemoglobinemia

POR AR Testis – Ambiguous or male Mixed features of 21-hydroxylase deficiency and

17α-hydroxylase/17,20-lyase deficiency, sometimes associated

with Antley-Bixler craniosynostosis

HSD17B3 AR Testis – Female or ambiguous Partial androgenization at puberty, ↑ androstenedione-totestosterone ratio

SRD5A2 AR Testis – Ambiguous or micropenis Partial androgenization at puberty, ↑ testosterone-todihydrotestosterone ratio

AKR1C2

(AKR1C4)

AR Testis – Female or ambiguous Decreased fetal DHT production

Disorders of Androgen Action

Androgen

receptor

X Testis – Female, ambiguous,

micropenis or normal

male

Phenotypic spectrum from complete androgen insensitivity

syndrome (female external genitalia) and partial androgen

insensitivity (ambiguous) to normal male genitalia and infertility

Abbreviations: AD, autosomal dominant; AKR1C2, aldo-keto reductase family 1 member 2; AR, autosomal recessive; ARX, aristaless related homeobox, X-linked; CAH,

congenital adrenal hyperplasia; CBX2, chromobox homologue 2; CYB5A, cytochrome b5; CYP11A1, P450 cholesterol side-chain cleavage; CYP17A1, cytochrome P450 family

17 subfamily A member 1; DAX1, dosage sensitive sex-reversal, adrenal hypoplasia congenita on the X chromosome, gene 1; DHEA, dehydroepiandrosterone; DHCR7, sterol 7

δ reductase; DHH, desert hedgehog; DMRT1,doublesex and mab3-related transcription factor 1; GATA4, GATA binding protein 4; HSD17B3, 17β-hydroxysteroid dehydrogenase

type 3; HSD3B2, 3β-hydroxysteroid dehydrogenase type 2; LHR, LH receptor; MAP3K1, mitogen-activated protein kinase kinase kinase 1; MIRAGE, myelodysplasia, infection,

restriction of growth, adrenal hypoplasia, genital phenotypes, and enteropathy; MYRF, myelin regulatory factor; POR, P450 oxidoreductase; SF1, steroidogenic factor 1; SL,

sex-limited; SOX8, SRY-related HMG-box gene 8; SOX9, SRY-related HMG-box gene 9; SRD5A2, 5α-reductase type 2; SRY, sex-related gene on the Y chromosome; StAR,

steroidogenic acute regulatory protein; TSPYL1, testis-specific Y-encoded-like protein 1; WAGR, Wilms’ tumor, aniridia, genitourinary anomalies, and mental retardation;

WNT4, wingless-type mouse mammary tumor virus integration site, 4; WT1, Wilms’ tumor–related gene 1; ZFPM2, zinc finger protein, multitype 2; ZNRF3, zinc and ring finger 3.

3004 PART 12 Endocrinology and Metabolism

Cholesterol

StAR

Pregnenolone

(Cholesterol

 side chain

 cleavage enzyme)

CYP11A1

Congenital

 adrenal

 hyperplasia

 and

 46,XX

 androg-

 enization

46,XY

 underandrog-

 enization

 only

CYP17,

(17,20-lyase), CYB5A

(Cytochrome b5)

Congenital

 adrenal

 hyperplasia

 and 46,XY

 underandrog-

 enization

HSD17B3

 (17β-hydroxysteroid

 dehydrogenase 3)

Progesterone

(3β-hydroxysteroid

 dehydrogenase 2)

CYP21A2

 (21-hydroxylase)

SRD5A2

 (5α-reductase)

HSD3B2

CYP11B1

 (11-hydroxylase)

17-hydroxyprogesterone

11-deoxycortisol Androstenedione

Cortisol Testosterone

Glucocorticoid

 Pathway

Androgen Pathway

CYP17 (17α-hydroxylase)

LH

 (testis)

ACTH

 (adrenal)

Dihydrotestosterone

FIGURE 390-4 Simplified overview of glucocorticoid and androgen synthesis pathways. Defects in CYP21A2

and CYP11B1 shunt steroid precursors into the androgen pathway and cause androgenization of the 46,XX

fetus. Testosterone is synthesized in the testicular Leydig cells and converted to dihydrotestosterone

peripherally. Defects in enzymes involved in androgen synthesis result in underandrogenization of the 46,XY

fetus. StAR, steroidogenic acute regulatory protein.

effects at puberty can be prevented (by gonadectomy or pubertal suppression) and female-typical puberty induced with estrogen. They also

have an increased risk of GCC, again raising the question of if and

when to perform gonadectomy. Azoospermia and male-factor infertility also have been described in association with mild loss-of-function

mutations in the androgen receptor.

■ OTHER DISORDERS AFFECTING 46,XY MALES

Persistent müllerian duct syndrome is the presence of a uterus in an

otherwise phenotypic male. This condition can result from pathogenic variants in AMH or its receptor (AMHR2). The uterus may be

removed, but only if damage to the vasa deferentia and blood supply

to the testes can be avoided. Isolated hypospadias occurs in ~1 in

250 males. Most cases are idiopathic, although evidence of penoscrotal hypospadias, poor phallic development, and/or bilateral cryptorchidism requires investigation for an underlying DSD (e.g., partial

gonadal dysgenesis, mild defect in testosterone action, or even severe

forms of 46,XX CAH). Unilateral undescended testes (cryptorchidism)

affect >3% of boys at birth. Orchidopexy should be considered if the

testis has not descended by 6–9 months of age. Bilateral cryptorchidism

occurs less frequently and should raise suspicion of gonadotropin

deficiency or DSD. Syndromic associations and intrauterine growth

retardation also occur relatively frequently in association with impaired

testicular function or target tissue responsiveness, but the underlying

etiology of many of these conditions is unknown.

■ 46,XX DSD

Androgenization of the 46,XX fetus occurs when the gonad (ovary)

contains androgen-secreting testicular tissue or after increased androgen exposure, which is usually adrenal in origin (Table 390-1).

46,XX Testicular DSD Testicular tissue can

develop in 46,XX testicular DSD (46,XX males)

most often following translocation of SRY. This

may be diagnosed with karyotype/phenotype

discordance or later in life during evaluation for

hypogonadism or infertility. Individuals with

this condition develop testes with normal testosterone production, leading to external male

phenotype in utero, and produce AMH/MIS to

regress müllerian structures. They have azoospermia due to lack of the AZF region of the Y

chromosome. Progressive testicular regression

and hypogonadism are common. Gender identity

is typically male.

46,XX OTDSD Ovotestes (or testes) can also

develop in individuals with a 46,XX karyotype

following upregulation of SOX9 or SOX3 or

defects in RSPO1, NR2F2, WT1, or SF1/NR5A1

(Table 390-5). OTDSD is discussed above under

“Disorders of Chromosomal Sex.”

Increased Androgen Exposure • 21-

hydroxylase deficiency (congenital

adrenal hyperplasia) The classic form

of 21-hydroxylase deficiency (21-OHD) is the

most common cause of CAH (Chap. 386), and

it is the most common cause of androgenization in chromosomal 46,XX females (incidence

between 1 in 10,000 and 1 in 15,000) (Table

390-5). Affected individuals are homozygous

or compound heterozygous for severely disruptive sequence variants in the gene (CYP21A2)

encoding the enzyme 21-hydroxylase. Impaired

21-hydroxylase activity prevents adrenal glucocorticoid and mineralocorticoid synthesis, thus

shunting steroid precursors into the androgen

synthesis pathway (Fig. 390-4). Increased androgen synthesis in utero causes androgenization of

the 46,XX fetus in the first trimester. Atypical

genitalia are seen at birth, with varying degrees of clitoral enlargement

and labial fusion.

A salt-wasting crisis usually manifests between 5 and 21 days of life

and is a potentially life-threatening event that requires urgent fluid

resuscitation and steroid treatment. Thus, a diagnosis of 21-OHD

should be considered in any baby with atypical genitalia with bilateral

nonpalpable gonads. Males (46,XY) with 21-OHD have no genital

abnormalities at birth but are equally susceptible to adrenal insufficiency and salt-losing crises. Excess androgen production can cause

gonadotropin-independent precocious puberty in males with 21-OHD.

Patients with nonclassic 21-OHD produce normal amounts of cortisol and aldosterone but at the expense of producing excess androgens.

Symptoms may include hirsutism, menstrual dysfunction, subfertility,

and recurrent miscarriages. This is one of the most common recessive

disorders in humans, with an incidence as high as 1 in 100–500 in

many populations and 1 in 27 in Ashkenazi Jews of Eastern European

origin.

TREATMENT

Congenital Adrenal Hyperplasia

Acute salt-wasting crises require fluid resuscitation, IV hydrocortisone, and correction of hypoglycemia. Once the patient is

stabilized, glucocorticoids must be given to correct the cortisol

insufficiency and suppress ACTH stimulation, thereby preventing

further virilization, rapid skeletal maturation, adrenal rest tumor

formation, and the development of polycystic ovaries. Mineralocorticoid replacement may be needed, along with salt supplements in

early life. In childhood, treatment is also titrated carefully to prevent

3005 Sex Development CHAPTER 390

TABLE 390-5 Selected Genetic Causes of 46,XX Disorders of Sex Development (DSDs)

GENE INHERITANCE GONAD UTERUS EXTERNAL GENITALIA ASSOCIATED FEATURES

Testicular/Ovotesticular DSD

SRY Translocation Testis or ovotestis – Male or ambiguous

SOX9 dup17q24 Testis or ovotestis – Male or ambiguous

SF1/NR5A1 (codon 92) AD Testis or ovotestis +/– Male or ambiguous

WT1 (zinc finger 4) AD Testis or ovotestis +/– Male or ambiguous

Other causes of testicular/ovotesticular DSD include: COUP-TF2/NR2F2 (congenital heart disease), RSPO1 (palmar plantar hyperkeratosis, squamous cell skin

carcinoma), WNT4 (SERKAL syndrome), dysregulation/duplication of SOX3 (Xq27)

Increased Androgen Synthesis

HSD3B2 AR Ovary + Clitoromegaly CAH, primary adrenal insufficiency, mild

androgenization due to ↑ DHEA

CYP21A2 AR Ovary + Ambiguous CAH, phenotypic spectrum from severe salt-losing

forms associated with adrenal insufficiency to simple

virilizing forms with compensated adrenal function, ↑

17-hydroxyprogesterone

POR AR Ovary + Ambiguous or female Mixed features of 21-hydroxylase deficiency and

17α-hydroxylase/17,20-lyase deficiency, sometimes

associated with Antley-Bixler craniosynostosis

CYP11B1 AR Ovary + Ambiguous CAH, hypertension due to ↑ 11-deoxycorticosterone

CYP19 AR Ovary + Ambiguous Maternal virilization during pregnancy, absent breast

development at puberty

Abbreviations: ACTH, adrenocorticotropin; AD, autosomal dominant; AR, autosomal recessive; CAH, congenital adrenal hyperplasia; COUP-TF2, chicken ovalbumin upstream

promoter transcription factor 2; CYP11B1, 11β-hydroxylase; CYP19, aromatase; CYP21A2, 21-hydroxylase; DHEA, dehydroepiandrosterone; HSD3B2, 3β-hydroxysteroid

dehydrogenase type 2; POR, P450 oxidoreductase; RSPO1, R-spondin 1; SERKAL, sex reversion, kidneys, adrenal and lung dysgenesis; SF1, steroidogenic factor 1; SOX3,

SRY-related HMG-box gene 3; SOX9, SRY-related HMG-box gene 9; SRY, sex-related gene on the Y chromosome; WT1, Wilms’ tumor–related gene 1.

impairment of linear growth. In the future, different forms of glucocorticoid replacement and multimodal therapies may improve

treatment options. See Chap. 386 for detailed discussion of hormone replacement.

Individuals with 46,XX CAH due to classic 21-OHD historically

underwent genitoplasty in infancy, but if and when these procedures should be performed is debated. Concerns have arisen about

the importance of assent/consent by the individual for genital surgery, potential long-term side effects related to sexual function and

ability to achieve orgasm, and the increased incidence of nonfemale

gender identity. Surgical options include vaginoplasty and clitoroplasty. When vaginoplasty is performed in infancy, surgical revision

or vaginal dilation may still be needed in adolescence or adulthood

and, if deferred, may be necessary for menstrual flow or intercourse.

Current clinical practice guidelines recommend that parents be

informed of all surgical options including the option to defer surgery. Women with 21-OHD frequently develop polycystic ovaries

and have subfertility. The latter occurs due to multiple factors

including anatomic barriers, hormone imbalances, and psychological effects of the condition. Preconception genetic counseling is recommended. Due to concerns about fetal neurologic development,

prenatal treatment with dexamethasone to prevent androgenization

of a fetus is currently not recommended unless in a study protocol

that allows long-term follow-up of all children treated.

The treatment of other forms of CAH (including in 46,XY individuals) includes mineralocorticoid and glucocorticoid replacement for salt-losing conditions (e.g., StAR, CYP11A1, HSD3B2),

suppression of ACTH drive with glucocorticoids in disorders

associated with hypertension (e.g., CYP11B1), and appropriate

sex hormone replacement in adolescence and adulthood, when

necessary.

OTHER CAUSES Increased androgen synthesis can also occur in

CAH due to defects in POR, 11β-hydroxylase (CYP11B1), and 3βhydroxysteroid dehydrogenase type 2 (HSD3B2) and with mutations in

the genes encoding aromatase (CYP19). Increased androgen exposure

in utero can occur with maternal virilizing tumors, luteomas, and

ingestion of androgenic compounds.

■ OTHER DISORDERS AFFECTING 46,XX FEMALES

Congenital absence of the vagina occurs in association with müllerian

agenesis or hypoplasia as part of the Mayer-Rokitansky-Kuster-Hauser

(MRKH) syndrome. This diagnosis should be considered in otherwise

phenotypic females with primary amenorrhea. Associated features

include renal (agenesis) and cervical spinal abnormalities.

■ GLOBAL CONSIDERATIONS

The approach to a child or adolescent with atypical genitalia or another

DSD requires cultural sensitivity, as the concepts of sex and gender

vary widely around the world. Rare genetic DSDs can occur more

frequently in specific populations (e.g., 5α-reductase type 2 in the

Dominican Republic). Different forms of CAH also show ethnic and

geographic variability. In many countries, appropriate biochemical

tests may not be readily available, and access to appropriate forms of

treatment and support may be limited.

■ FURTHER READING

Ahmed SF et al: Turner HE. Society for Endocrinology UK Guidance

on the initial evaluation of a suspected difference or disorder of sex

development (Revised 2021). Clin Endocrinol (Oxf) 818, 2021.

Cools M et al: Caring for individuals with a difference of sex development (DSD): A Consensus Statement. Nat Rev Endocrinol 14:415, 2018.

Gravholt CH et al: Clinical practice guidelines for the care of girls

and women with Turner syndrome: Proceedings from the 2016 Cincinnati International Turner Syndrome Meeting. Eur J Endocrinol

177:G1, 2017.

Merke DP, Auchus RG: Congenital adrenal hyperplasia due to

21-hydroxylase deficiency. N Engl J Med 383:1248, 2020.

Mongan NP et al: Androgen insensitivity syndrome. Best Pract Res

Clin Endocrinol Metab 29:569, 2015.

Zitzmann M et al: European Academy of Andrology guidelines on

Klinefelter syndrome: Endorsing organization: European Society of

Endocrinology. Andrology 9:145, 2021.

3006 PART 12 Endocrinology and Metabolism

The male reproductive system regulates sex differentiation, androgenization, and the hormonal changes that accompany puberty, ultimately

leading to spermatogenesis and fertility. Under the control of the pituitary hormones—luteinizing hormone (LH) and follicle-stimulating

hormone (FSH)—the Leydig cells of the testes produce testosterone

and germ cells are nurtured by Sertoli cells to divide, differentiate, and

mature into sperm. During embryonic development, testosterone and

dihydrotestosterone (DHT) induce the wolffian duct and virilization of

the external genitalia. During puberty, testosterone promotes somatic

growth and the development of secondary sex characteristics. In the

adult, testosterone is necessary for spermatogenesis, libido and normal sexual function, and maintenance of muscle and bone mass. This

chapter focuses on the physiology of the testes and disorders associated

with decreased androgen production, which may be caused by gonadotropin deficiency or by primary testis dysfunction. Infertility occurs

in ~5% of men and is increasingly amenable to treatment by hormone

replacement or by using sperm transfer techniques. For further discussion of sexual dysfunction, disorders of the prostate, and testicular cancer, see Chaps. 397, 87, and 88, respectively.

DEVELOPMENT AND STRUCTURE

OF THE TESTIS

The fetal testis develops from a single bipotential progenitor cell

population in the undifferentiated gonad after expression of a genetic

cascade that is initiated by the gene encoding SRY (sex-related gene on

the Y chromosome) (Chap. 390). SRY, whose expression is regulated by

histone modification and DNA methylation, induces differentiation of

Sertoli cells, which surround germ cells and, together with peritubular

myoid cells, form testis cords that will later develop into seminiferous

tubules. Fetal Leydig cells and endothelial cells migrate into the gonad

from the adjacent mesonephros but may also arise from interstitial

cells that reside between testis cords. Fetal Leydig cells atrophy after

birth and do not contribute to the origin of adult Leydig cells, which

originate from undifferentiated progenitor cells that appear in the testis after birth and acquire full steroidogenic function during puberty.

Testosterone produced by the fetal Leydig cells supports the growth

and differentiation of Wolffian duct structures that develop into the

epididymis, vas deferens, and seminal vesicles. Testosterone is also

converted to DHT (see below), which induces formation of the prostate and the external male genitalia, including the penis, urethra, and

scrotum. Testicular descent through the inguinal canal is controlled in

part by Leydig cell production of insulin-like factor 3 (INSL3), which

acts via a receptor termed great (G protein–coupled receptor affecting

testis descent). Sertoli cells produce Müllerian inhibiting substance

(MIS), which causes regression of the Müllerian structures, including

the fallopian tube, uterus, and upper segment of the vagina.

NORMAL MALE PUBERTAL DEVELOPMENT

Puberty commonly refers to the maturation of the reproductive axis and

the development of secondary sex characteristics. In addition to reproductive hormones, it requires a coordinated response of multiple hormonal systems including metabolic signals (e.g., leptin), as well as the

adrenal and growth hormone (GH) axes (Fig. 391-1). The development

of secondary sex characteristics is initiated by adrenarche, which usually

occurs between 6 and 8 years of age when the adrenal gland begins to

produce greater amounts of androgens from the zona reticularis, the

principal site of dehydroepiandrosterone (DHEA) production. The sex

maturation process is greatly accelerated by the activation of the hypothalamic-pituitary axis and the production of gonadotropin-releasing

hormone (GnRH). The GnRH pulse generator in the hypothalamus

391 Disorders of the Testes and

Male Reproductive System

Shalender Bhasin, J. Larry Jameson

is active during fetal life and early infancy but is restrained until the

early stages of puberty by a neuroendocrine brake imposed by the

inhibitory actions of glutamate and γ-aminobutyric acid (GABA)

in the mediobasal hypothalamus and neuropeptide Y. Although the

pathways that initiate reactivation of the GnRH pulse generator at the

onset of puberty remain incompletely understood, mounting evidence

supports involvement of GPR54, a G protein–coupled receptor that

binds an endogenous ligand, kisspeptin. Individuals with mutations of

GPR54 fail to enter puberty, and experiments in primates demonstrate

that infusion of the ligand is sufficient to induce premature puberty.

Kisspeptin signaling plays an important role in mediating the feedback

action of sex steroids on gonadotropin secretion and in regulating the

tempo of sexual maturation at puberty. Leptin, a hormone produced by

adipose cells, plays a permissive role in the resurgence of GnRH secretion

at the onset of puberty, as leptin-deficient individuals also fail to enter

puberty (Chap. 401). The adipocyte hormone leptin, the gut hormone

ghrelin, neuropeptide Y, and kisspeptin integrate the signals originating

in energy stores and metabolic tissues with mechanisms that control

onset of puberty through regulation of GnRH secretion. Energy deficit

and excess and metabolic stress are associated with disturbed reproductive maturation and timing of puberty.

The early stages of puberty are characterized by nocturnal surges

of LH and FSH. Growth of the testes is usually the first clinical sign

of puberty, reflecting an increase in seminiferous tubule volume.

Increasing levels of testosterone deepen the voice and stimulate muscle

growth. Conversion of testosterone to DHT leads to growth of the

external genitalia and pubic hair. DHT also stimulates prostate and

facial hair growth and initiates recession of the temporal hairline. The

growth spurt occurs at a testicular volume of ~10–12 mL. GH increases

early in puberty and is stimulated in part by the rise in gonadal steroids. GH increases the level of insulin-like growth factor 1 (IGF-1),

which enhances linear bone growth. The prolonged pubertal exposure

to gonadal steroids (mainly estradiol) ultimately induces epiphyseal

closure and limits further bone growth.

REGULATION OF TESTICULAR FUNCTION

■ REGULATION OF THE HYPOTHALAMICPITUITARY-TESTIS AXIS IN ADULT MAN

Pulsatile secretion of GnRH in the hypothalamus is regulated by

the KNDy neurons through the release of kisspeptin, neurokinin B

(NKB), and dynorphin (Fig. 391-2). Kisspeptin binds to the kisspeptin

(GPR54) receptors in the cell bodies of the GnRH neurons as well as in

the GnRH nerve terminals in the median eminence to induce pulsatile

GnRH secretion into the portal blood. As a component of this autocrine/paracrine loop, NKB released by the KNDy neurons activates

NK3R to stimulate kisspeptin release. KNDy neurons also produce

dynorphin A, which inhibits basal as well as NKB-stimulated kisspeptin release through the mediation of K-type opioid receptor. The

8 9 10 11 12

4–6

2 3 4 5

2 3 4 5

10–12 15–25

13

Age (years)

Tanner stages

Height velocity

Testicular

 volume (mL)

Genitalia

Pubic hair

14 15 16 17

FIGURE 391-1 Pubertal events in males. Sexual maturity ratings for genitalia and

pubic hair and divided into five stages.

3007 Disorders of the Testes and Male Reproductive System CHAPTER 391

negative feedback effects of testosterone, estradiol, and progesterone

are mediated through KNDy neurons in the preoptic area by inhibition

of kisspeptin release.

Hypothalamic GnRH regulates the production of the pituitary

gonadotropins LH and FSH (Fig. 391-2). GnRH is released in discrete

pulses approximately every 2 h, resulting in corresponding pulses of

LH and FSH. These dynamic hormone pulses account in part for the

wide variations in LH and testosterone, even within the same individual. LH acts primarily on the Leydig cell to stimulate testosterone

synthesis. The regulatory control of androgen synthesis is modulated

by dynamic integration of the feedforward elements exerted on the

testis by LH and FSH and the feedback exerted by testosterone and

estrogen on both the hypothalamus and the pituitary. FSH acts on the

Sertoli cell to regulate spermatogenesis and the production of Sertoli

products such as inhibin B, which acts to selectively suppress pituitary

FSH. Despite these somewhat distinct Leydig and Sertoli cell–regulated

pathways, testis function is integrated at several levels: GnRH regulates

both gonadotropins; spermatogenesis requires high levels of testosterone; and numerous paracrine interactions between Leydig and Sertoli

cells are necessary for normal testis function.

■ THE LEYDIG CELL: ANDROGEN SYNTHESIS

LH binds to its seven-transmembrane G protein–coupled receptor

to activate the cyclic AMP pathway. Stimulation of the LH receptor

induces steroid acute regulatory (StAR) protein, along with several

steroidogenic enzymes involved in androgen synthesis. LH receptor

mutations cause Leydig cell hypoplasia or agenesis, underscoring the

importance of this pathway for Leydig cell development and function.

The rate-limiting process in testosterone synthesis is the transport of

intracellular cholesterol by the StAR protein to the inner mitochondrial

membrane. Mutations of the StAR protein are associated with congenital lipoid adrenal hyperplasia, a rare form of congenital adrenal hyperplasia (CAH) characterized by very low adrenal and gonadal steroids.

Peripheral benzodiazepine receptor, a mitochondrial cholesterol-binding

protein, is also an acute regulator of Leydig cell steroidogenesis. The

major enzymatic steps involved in testosterone synthesis are summarized in Fig. 391-3. After cholesterol transport into the mitochondrion,

the formation of pregnenolone by CYP11A1 (side chain cleavage

enzyme) is a limiting enzymatic step. The 17α-hydroxylase and the

17,20-lyase reactions are catalyzed by a single enzyme, CYP17A1; posttranslational modification (phosphorylation) of this enzyme and the

presence of specific enzyme cofactors, such as cytochrome B, confer

17,20-lyase activity selectively in the testis and zona reticularis of the

adrenal gland. Although CYP17A1 is able to catalyze the conversion of

progesterone to 17α-hydroxyprogesterone, most of δ4-androstenedione

in humans is not derived from 17α-hydroxyprogesterone but rather from

the conversion of 17α-hydroxypregnenolone to DHEA in the δ5 pathway

and further conversion of DHEA to δ4-androstenedione. Abiraterone

is a dual inhibitor of 17α-hydroxylase and 17,20-lyase activities, which

play an important role in androgen synthesis in castration-resistant

prostate cancers. Testosterone can be converted to the more potent DHT

by a family of steroid 5α-reductase enzymes, or it can be aromatized to

estradiol by CYP19 (aromatase). At least two isoforms of steroid 5αreductase, SRD5A1 and SRD5A2, have been described; all known

patients with 5α-reductase deficiency have had mutations in SRD5A2,

the predominant form in the prostate and the skin. Finasteride predominantly inhibits SRD5A2, whereas dutasteride is a dual inhibitor of both

SRD5A1 and SRD5A2. DHT can also be derived through the backdoor

pathway in which 17α-hydroxyprogesterone is converted to androsterone and eventually to DHT. Recent reports of mutations in the AKR1C2/4

genes in undervirilized 46,XY individuals suggest that the backdoor pathway for DHT formation, which was originally described in the tammar

wallaby, is active in the human fetal testis. The placental progesterone

serves as a substrate for the synthesis of androsterone via the backdoor

pathway, which is then converted to DHT in the genital tubercle.

Testosterone Transport and Metabolism In males, 95% of circulating testosterone is derived from testicular production (3–10 mg/d).

Direct secretion of testosterone by the adrenal and the peripheral

conversion of androstenedione to testosterone collectively account

for another 0.5 mg/d of testosterone. Only a small amount of DHT

(70 μg/d) is secreted directly by the testis; most circulating DHT is

derived from peripheral conversion of testosterone. Most of the daily

production of estradiol (~45 μg/d) in men is derived from aromatasemediated peripheral conversion of testosterone and androstenedione.

– +

+

+

–

Kiss

Preoptic area

Pulsatile

GnRH

secretion Median

eminence

Negative

feedback

by sex

steroids

Tunica albuginea

Seminiferous

tubules

Interstitial

Leydig cells

(testosterone)

Anterior

pituitary

Vas deferens

Testosterone

Inhibin B

E2

DHT

Pulsatile

LH, FSH

secretion

LH

FSH Epididymis

Sertoli cell

(Inhibin B)

Spermatid

Spermatogonium

GnRH

GnRH

neuron

KNDY

neurons

KOR

Dyn

Dyn

NKB

NK3R

Kiss

GPR54

Arcuate nucleus

FIGURE 391-2 Hypothalamic-pituitary-gonadotropin axis, structure of testis,

seminiferous tubule. DHT, dihydrotestosterone; Dyn, dynorphin A; E2

, 17β-estradiol;

FSH, follicle-stimulation hormone; GnRH, gonadotropin-releasing hormone; GPR54,

G protein–coupled receptor 54 for kisspeptin; Kiss, kisspeptin; KNDY, kisspeptin,

neurokinin B, dynorphin neurons; LH, luteinizing hormone; NKB, neurokinin B; NK3R,

neurokinin 3 receptor.

3008 PART 12 Endocrinology and Metabolism

Circulating testosterone is bound predominantly to sex hormone–

binding globulin (SHBG) and albumin (Fig. 391-4) and, to a lesser

extent, to cortisol-binding globulin (CBG) and orosomucoid. SHBG

binds testosterone with much greater affinity than albumin, CBG,

and orosomucoid. The binding proteins regulate the transport and

bioavailability of testosterone. SHBG circulates as a dimer, and testosterone’s binding to SHBG involves intermonomeric allostery such

that neither the conformation nor the binding affinity of the two

monomers is equivalent. Similarly, estradiol binding to SHBG involves

bidirectional, intermonomeric allostery that changes the distribution

of both monomers among various energy and conformational states.

Intermonomeric allostery offers a mechanism to extend the binding

range of SHBG and regulate hormone bioavailability as sex hormone

concentrations vary widely during life. Human serum albumin (HSA)

contains multiple, allosterically coupled binding sites for testosterone.

Testosterone shares these binding sites on HSA with free fatty acids.

Commonly used drugs such as ibuprofen and antibiotics can displace

testosterone from HSA under various physiologic states or disease conditions, affecting its bioavailability. SHBG concentrations are decreased

by androgens, obesity, diabetes mellitus, hypothyroidism, nephrotic

syndrome, and genetic factors. Conversely, estrogen administration,

hyperthyroidism, many chronic inflammatory illnesses, infections

such as HIV or hepatitis B and C, aging, and the use of some anticonvulsants are associated with high SHBG concentrations.

Testosterone is metabolized predominantly in the liver, although

some degradation occurs in peripheral tissues, particularly the prostate and the skin. In the liver, testosterone is converted by a series

of enzymatic steps that involve 5α- and 5β-reductases, 3α- and

3β-hydroxysteroid dehydrogenases, and 17β-hydroxysteroid dehydrogenase into androsterone, etiocholanolone, DHT, and 3α-androstanediol.

17 18

19

16

1514

13 12 11

10

B

C D

9 2

1

OH 3

4

5

6

7

8

A

Cholesterol

Pregnenolone Progesterone

CYP17A1 17α-hydroxylase

17α-hydroxylase

17β-hydroxysteroid

dehydrogenase 3 17β-HSD 3

17 hydroxy

allopregnanolone CYP17A1 CYP17A1

AKRIC2/4

17 hydroxydihydroprogesterone

allopregnenolone 3β-hydroxy

steroid dehydrogenase 2

3β HSD 1

Steroid

5α-reductase type 1

SRD5AI

Steroid 5α-reductase type 2

SRD5A2

17 hydroxypregnenolone 17-hydroxyprogesterone

Androsterone

17β hydroxy

steroid

dehydrogenase

3

17β-hydroxy

steroid

dehydrogenase 3

HSD17B3

AKRICI-4

HSD17B3

HSD17B6

AKRIC2

Androstanedione

δ4 Androstenedione

CYP17

Cholesterol side chain

cleavage enzyme

CYP11A1

StAR

CYP19

CYP17A1

CYB5

DHEA

Androstenediol Testosterone

Aromatase

Estradiol

Androstanediol

OH

O

OH

O

H OH

OH

HSD17B6

17β-hydroxysteroid

dehydrogenase 6

5α-dihydrotestosterone

Classical Pathway Alternate Backdoor Pathway

17β hydroxy

steroid

dehydrogenase 6

FIGURE 391-3 The biochemical pathway in the conversion of 27-carbon sterol cholesterol to androgens and estrogens.

3009 Disorders of the Testes and Male Reproductive System CHAPTER 391

These compounds undergo glucuronidation or sulfation before being

excreted by the kidneys.

Mechanism of Androgen Action Testosterone exerts some of its

biologic effects by binding to androgen receptor (AR), either directly

or after its conversion to DHT by the steroid 5α reductase. The actions

of testosterone on the Wolffian structures, skeletal muscle, erythropoiesis, and bone in men do not require its obligatory conversion to

DHT. However, the conversion of testosterone to DHT is necessary

for the masculinization of the urogenital sinus and genital tubercle.

Aromatization of testosterone to estradiol mediates additional effects

of testosterone on bone resorption, epiphyseal closure, sexual desire,

vascular endothelium, and fat. DHT can also be converted in some tissues by the combined actions of the 3α-hydroxysteroid dehydrogenase

and 3β-hydroxysteroid dehydrogenase to 5α-androstane-3β,17β-diol,

which is a high-affinity ligand and agonist of estrogen receptor β.

5α-DHT is further converted in some cell types to 5α-androstane3α,17β-diol, a modulator of GABAA receptors.

The AR is structurally related to the nuclear receptors for estrogen,

glucocorticoids, and progesterone (Chap. 377). The AR, a 919–amino

acid protein with a molecular mass of ~110 kDa, is encoded by a gene

on the long arm of the X chromosome. A polymorphic region in the

amino terminus of the receptor, which contains a variable number of

glutamine and glycine repeats, modifies the transcriptional activity of

the receptor. The AR protein is distributed in both the cytoplasm and

the nucleus. The ligand binding to the AR induces conformational

changes that allow the recruitment and assembly of tissue-specific

cofactors and causes it to translocate into the nucleus, where it binds to

specific androgen response elements in the DNA or other transcription

factors already bound to DNA. Thus, the AR is a ligand-regulated transcription factor that regulates the expression of androgen-dependent

genes in a tissue-specific manner. Testosterone binds to AR with half

the affinity of DHT. The DHT-AR complex also has greater thermostability and a slower dissociation rate than the testosterone-AR complex.

However, the molecular basis for selective testosterone versus DHT

actions remains incompletely explained. Some androgen effects, such

as those on the smooth muscle, may be mediated by nongenomic

AR signal transduction pathways. The nongenomic actions of testosterone involve direct activation of kinase signaling cascades such as

mitogen-activated protein kinase and the cyclic AMP response element

binding protein transcription factor. Some effects of testosterone on

cell proliferation and autophagy require the mediation of GPRC6A.

■ THE SEMINIFEROUS TUBULES:

SPERMATOGENESIS

The seminiferous tubules are convoluted, closed loops with both ends

emptying into the rete testis, a network of progressively larger efferent

ducts that ultimately form the epididymis (Fig. 391-2). The seminiferous tubules total ~600 m in length and compose about two-thirds

of testis volume. The walls of the tubules are formed by polarized

Sertoli cells that are apposed to peritubular myoid cells. Tight junctions between Sertoli cells create the blood-testis barrier. Germ cells

compose the majority of the seminiferous epithelium (~60%) and are

intimately embedded within the cytoplasmic extensions of the Sertoli

cells, which function as “nurse cells.” Germ cells progress through

characteristic stages of mitotic and meiotic divisions. A pool of type

A spermatogonia serve as stem cells capable of self-renewal. Primary

spermatocytes are derived from type B spermatogonia and undergo

meiosis before progressing to spermatids that undergo spermiogenesis

(a differentiation process involving chromatin condensation, acquisition of an acrosome, elongation of cytoplasm, and formation of a

tail) and are released from Sertoli cells as mature spermatozoa. The

complete differentiation process into mature sperm requires 74 days.

Peristaltic-type action by peritubular myoid cells transports sperm into

the efferent ducts. The spermatozoa spend an additional 21 days in the

epididymis, where they undergo further maturation and capacitation.

The normal adult testes produce >100 million sperm per day.

Naturally occurring mutations in FSHβ or in the FSH receptor

confirm an important, but not essential, role for this pathway in spermatogenesis. Females with mutations in FSHβ or the FSH receptor

are hypogonadal and infertile because ovarian follicles do not mature;

males with these mutations exhibit variable degrees of reduced spermatogenesis, presumably because of impaired Sertoli cell function.

Because Sertoli cells produce inhibin B, an inhibitor of FSH, seminiferous tubule damage (e.g., by radiation) causes a selective increase

of FSH. Testosterone reaches very high concentrations locally in the

testis and is essential for spermatogenesis. The cooperative actions of

FSH and testosterone are important in the progression of meiosis and

spermiation. In the prepubertal testis, testosterone alone is insufficient

for completion of spermatogenesis; however, in men with postpubertal onset of gonadotropin deficiency, human chorionic gonadotropin

(hCG) or recombinant LH can reinitiate spermatogenesis without

FSH. FSH and testosterone regulate germ cell survival via the intrinsic

and extrinsic apoptotic mechanisms. FSH may also play an important

role in supporting spermatogonia. Gonadotropin-regulated testicular

RNA helicase (GRTH/DDX25), a testis-specific gonadotropin/androgenregulated RNA helicase, is present in germ cells and Leydig cells and

may be an important factor in the paracrine regulation of germ cell

development. Several cytokines and growth factors are also involved

in the regulation of spermatogenesis by paracrine and autocrine mechanisms. A number of knockout mouse models exhibit impaired germ

cell development or spermatogenesis, presaging possible mutations

associated with male infertility.

The human Y chromosome contains two pseudoautosomal regions

that are located at the two tips of Y chromosome and can recombine

with homologous regions of the X chromosome (Fig. 391-5). The

genes in the pseudoautosomal regions are involved in cell signaling,

transcriptional regulation, and mitochondrial function. Mutations

of genes in pseudoautosomal region 1 are associated with mental

disorders and short stature. The euchromatic part of the Y chromosome that does not recombine with the X chromosome is referred to

as the male-specific region of the Y chromosome (MSY). The MSY

contains nine families of Y-specific multicopy genes; many of these

Y-specific genes are also testis-specific and necessary for spermatogenesis. Microdeletions in several nonoverlapping subregions of the

Y chromosome—AZFa, AZFb, AZFc, and AZFd, which contain many

spermatogenic genes (e.g., RNA-binding motif, RBM; deleted in azoospermia, DAZ)—are associated with oligospermia or azoospermia.

• Masculinization of

external genitalia

• Prostate growth

• Hair growth

• Bone resorption

• Epiphyseal closure

• Hypothalamic/

 pituitary feedback

• Fat mass

• Some vascular and

 behavioral effects

• Libido

Albumin

(33–34%)

Estradiol

• Wolffian duct

• Muscle mass

• Bone formation

• Spermatogenesis

• Erythropoiesis

SHBG

(44–66%)

Free or

unbound

(1–4%)

Testosterone

Excretion

5α-Dihydrotestosterone

(DHT)

Aromatase

(0.3%)

Steroid 5α-reductase

(6–8%)

Testosterone (4–9 mg/d)

Cortisol-binding globulin

Bioavailable Orosomucoid

FIGURE 391-4 Androgen metabolism and actions. SHBG, sex hormone–binding

globulin.

3010 PART 12 Endocrinology and Metabolism

Pseudoautosomal region 1

Euchromatic region of short arm

Euchromatic region of long arm

AZFc AZFb AZFa

Heterochromatic region of long arm

b1/b3 G1/G3

Pseudoautosomal region 2

Centromere

Long arm Yq

Male Specific Region of y (MSY)

Short arm Yp

FIGURE 391-5 Structure of the Y chromosome relevant for spermatogenesis.

Approximately 15% of infertile men with azoospermia and ~6% of

men with severe oligozoospermia harbor a Y microdeletion. Complete

deletions of the AZFa and AZFb subregions are typically associated

with Sertoli cells only and azoospermia and a poor prognosis for

sperm retrieval. In contrast, AZFc subregion microdeletions are typically associated with oligozoospermia and higher success rates for

sperm retrieval. Microdeletion involving the DAZ genes in the AZFc

region is one of the most common Y chromosome microdeletions

in infertile men. Several partial deletions of the AZFc region have

been described including the gr/gr deletion, which is associated with

infertility among Caucasian men in Europe and the Western Pacific

region, whereas the b2/b3 deletion is associated with male infertility

in African and Dravidian men.

TREATMENT

Male Factor Infertility

Treatment options for male factor infertility have expanded greatly

in recent years. Secondary hypogonadism is highly amenable to

treatment with pulsatile GnRH or gonadotropins (see below).

Assisted reproductive technologies, such as in vitro fertilization

(IVF) and intracytoplasmic sperm injection (ICSI), have provided

new opportunities for patients with primary testicular failure and

disorders of sperm transport. Choice of initial treatment options

depends on sperm concentration and motility. Expectant management should be attempted initially in men with mild male factor

infertility (sperm count of 15–20 × 106

/mL and normal motility).

Treatment of moderate male factor infertility (10–15 × 106

/mL

and 20–40% motility) should begin with intrauterine insemination

alone or in combination with treatment of the female partner with

clomiphene or gonadotropins, but it may require IVF with or without ICSI. For men with a severe defect (sperm count of <10 × 106

/

mL, 10% motility), IVF with ICSI or donor sperm has become the

treatment of choice. Yq microdeletions will be transmitted through

ICSI from the affected father to his male offspring if sperm carrying

the Yq microdeletion is used.

CLINICAL AND LABORATORY EVALUATION

OF MALE REPRODUCTIVE FUNCTION

■ HISTORY AND PHYSICAL EXAMINATION

The history should focus on developmental stages such as puberty

and growth spurts, as well as androgen-dependent events such as early

morning erections, frequency and intensity of sexual thoughts, and

frequency of masturbation or intercourse. Although libido and the

overall frequency of sexual acts are decreased in androgen-deficient

men, young hypogonadal men can achieve erections in response to

visual erotic stimuli. Men with acquired androgen deficiency often

report decreased energy and low mood.

The physical examination should focus on secondary sex

characteristics such as hair growth, gynecomastia, testicular

volume, prostate, and height and body proportions. Eunuchoid

proportions are defined as an arm span >2 cm greater than height

and suggest that androgen deficiency occurred before epiphyseal

fusion. Hair growth in the face, axilla, chest, and pubic regions

is androgen-dependent; however, changes may not be noticeable

unless androgen deficiency is severe and prolonged. Ethnicity

also influences the intensity of hair growth (Chap. 394). Testicular volume is best assessed by using a Prader orchidometer.

Testes range from 3.5 to 5.5 cm in length, which corresponds to

a volume of 12–25 mL. Advanced age does not influence testicular size, although the consistency becomes less firm. Asian men

generally have smaller testes than western Europeans, independent of differences in body size. Because of its possible role in

infertility, the presence of varicocele should be sought by palpation while the patient is standing; it is more common on the left

side. Patients with Klinefelter syndrome have markedly reduced

testicular volumes (1–2 mL). In congenital hypogonadotropic

hypogonadism, testicular volumes provide a good index for the degree

of gonadotropin deficiency and the likelihood of response to therapy.

■ GONADOTROPIN AND INHIBIN MEASUREMENTS

LH and FSH are measured using two-site immunoradiometric, immunofluorometric, or chemiluminescent assays, which have very low

cross-reactivity with other pituitary glycoprotein hormones and hCG

and have sufficient sensitivity to measure the low levels present in

patients with hypogonadotropic hypogonadism. In men with a low

testosterone level, an LH level can distinguish primary (high LH)

versus secondary (low or inappropriately normal LH) hypogonadism.

An elevated LH level indicates a primary defect at the testicular level,

whereas a low or inappropriately normal LH level suggests a defect at

the hypothalamic-pituitary level. LH pulses occur about every 1–3 h in

normal men. Thus, gonadotropin levels fluctuate, and samples should

be pooled or repeated when results are equivocal. FSH is less pulsatile

than LH because it has a longer half-life. Selective increase in FSH

suggests damage to the seminiferous tubules. Inhibin B, a Sertoli cell

product that suppresses FSH, is reduced with seminiferous tubule

damage. Inhibin B is a dimer with α-βB subunits and is measured by

two-site immunoassays.

GnRH Stimulation Testing The GnRH test is performed by

measuring LH and FSH concentrations at baseline and at 30 and

60 min after intravenous administration of 100 μg of GnRH. A minimally acceptable response is a twofold LH increase and a 50% FSH

increase. In the prepubertal period or with severe GnRH deficiency,

the gonadotrope may not respond to a single bolus of GnRH because

it has not been primed by endogenous hypothalamic GnRH; in these

patients, GnRH responsiveness may be restored by chronic, pulsatile

GnRH administration. With the availability of sensitive and specific

LH assays, GnRH stimulation testing is used rarely.

■ TESTOSTERONE ASSAYS

Total Testosterone Total testosterone includes both unbound

and protein-bound testosterone and is measured by radioimmunoassays, immunometric assays, or liquid chromatography tandem mass

spectrometry (LC-MS/MS). LC-MS/MS involves extraction of serum

by organic solvents, separation of testosterone from other steroids by

high-performance liquid chromatography and mass spectrometry, and

quantitation of unique testosterone fragments by mass spectrometry.

LC-MS/MS provides accurate and sensitive measurements of testosterone levels even in the low range and has emerged as the method of

choice for testosterone measurement. The use of LC-MS/MS for the

measurement of testosterone in laboratories that have been certified

by the Centers for Disease Control and Prevention’s (CDC) Hormone

Standardization Program for Testosterone (HoST) can ensure that testosterone measurements are accurate and calibrated to an international

standard. A single fasting morning sample provides a good approximation of the average testosterone concentration with the realization

3011 Disorders of the Testes and Male Reproductive System CHAPTER 391

that testosterone levels fluctuate because of its pulsatile, diurnal, and

circannual secretory rhythms. Testosterone is generally lower in the

late afternoon and is reduced by acute illness. The harmonized normal

range for total testosterone, measured using LC-MS/MS in nonobese

populations of European and American men aged 19–39 years, is

264–916 ng/dL. This harmonized reference range can be applied to

values from laboratories that are certified by the CDC’s HoST program.

Alterations in SHBG levels due to aging, obesity, diabetes mellitus,

hyperthyroidism, some types of medications, or chronic illness, or on

a congenital basis, can affect total testosterone levels. Heritable factors

contribute substantially to the population-level variation in testosterone levels, and genome-wide association studies have revealed polymorphisms in the SHBG gene as important contributors to variation

in testosterone levels.

Measurement of Unbound Testosterone Levels Most circulating testosterone is bound to SHBG and to albumin; only 2.0–4% of

circulating testosterone is unbound, or “free.” Free testosterone should

ideally be measured by equilibrium dialysis under standardized conditions using an accurate and reliable assay for total testosterone. The

unbound testosterone concentration also can be calculated from total

testosterone, SHBG, and albumin concentrations. Recent research has

shown that testosterone binding to SHBG is a multistep process that

involves complex allosteric interactions between the two binding sites

within the SHBG dimer; a novel ensemble allosteric model of testosterone’s binding to SHBG dimers provides good estimates of free testosterone concentrations. The previous law-of-mass-action equations

based on linear models of testosterone binding to SHBG used assumptions that have been shown to be erroneous. Tracer analogue methods

are relatively inexpensive and convenient, but they are inaccurate.

The term bioavailable testosterone refers to unbound testosterone plus

testosterone bound loosely to albumin and reflects the concept that

albumin-bound testosterone can dissociate at the capillary level, especially in tissues with long transit time, such as the liver and the brain.

Bioavailable testosterone can be determined by the ammonium sulfate

precipitation method. However, the measurements of bioavailable

testosterone using the ammonium sulfate precipitation are technically

challenging, susceptible to imprecision, and not recommended.

hCG Stimulation Test The hCG stimulation test is performed by

administering a single injection of 1500–4000 IU of hCG intramuscularly and measuring testosterone levels at baseline and 24, 48, 72, and

120 h after hCG injection. An alternative regimen involves three injections of 1500 units of hCG on successive days and measuring testosterone levels 24 h after the last dose. An acceptable response to hCG is a

doubling of the testosterone concentration in adult men. In prepubertal

boys, an increase in testosterone to >150 ng/dL indicates the presence

of testicular tissue. No response may indicate an absence of testicular

tissue or marked impairment of Leydig cell function. Measurement of

MIS, a Sertoli cell product, is also used to detect the presence of testes

in prepubertal boys with cryptorchidism.

■ SEMEN ANALYSIS

Semen analysis is the most important step in the evaluation of male

infertility. Samples are collected by masturbation following a period

of abstinence for 2–3 days. Semen volumes and sperm concentrations vary considerably among fertile men, and several samples may

be needed before concluding that the results are abnormal. Analysis

should be performed within an hour of collection. Using semen samples from >4500 men in 14 countries, whose partners had a time to

pregnancy of <12 months, the World Health Organization (WHO)

has generated the following one-sided reference limits for semen

parameters: semen volume, 1.5 mL; total sperm number, 39 million per

ejaculate; sperm concentration, 15 million per mL; vitality, 58% live;

progressive motility, 32%; total (progressive + nonprogressive) motility,

40%; and morphologically normal forms, 4.0%. Some men with low

sperm counts are nevertheless fertile. Some studies suggest that sperm

counts have declined in recent decades. A variety of tests for sperm

function can be performed in specialized laboratories, but these add

relatively little to the treatment options.

■ TESTICULAR BIOPSY

Testicular biopsy is useful in some patients with oligospermia or azoospermia as an aid in diagnosis and indication for the feasibility of treatment. Using fine-needle aspiration biopsy is performed under local

anesthesia to aspirate tissue for histology. Alternatively, open biopsies

can be performed under local or general anesthesia when more tissue is

required. A normal biopsy in an azoospermic man with a normal FSH

level suggests obstruction of the vas deferens, which may be correctable surgically. Biopsies are also used to harvest sperm for ICSI and to

classify disorders such as hypospermatogenesis (all stages present but

in reduced numbers), germ cell arrest (usually at primary spermatocyte

stage), and Sertoli cell–only syndrome (absent germ cells) or hyalinization (sclerosis with absent cellular elements).

Testing for Y Chromosome Microdeletions Y chromosome

microdeletions are detected by extracting DNA from peripheral blood

leukocytes and using polymerase chain reaction (PCR) amplification

using primers for ~300 sequence-tagged sites on the Y chromosome,

followed by gel electrophoresis to determine whether the DNA

sequences corresponding to the selected Y chromosome markers

are present. However, because these ~300 Y chromosome markers

account for only a small fraction of the 23 million base pairs on the Y

chromosome, a negative test does not exclude microdeletions in other

subregions of the Y chromosome.

DISORDERS OF SEXUAL DIFFERENTIATION

See Chap. 390.

DISORDERS OF PUBERTY

The onset and tempo of puberty vary greatly in the general population

and are affected by genetic, nutritional, and environmental factors.

Although a substantial fraction of the variance in the timing of puberty

is explained by heritable factors, the genes involved remain unknown.

■ PRECOCIOUS PUBERTY

Puberty in boys aged <9 years is considered precocious. Earlier onset

of puberty is associated with increased risk for several cancers, cardiovascular disease, hypertension, type 2 diabetes, hair pigmentation,

and shorter life span. Genome-wide association studies for age of

menarche in girls and age of voice deepening in boys have identified

389 independent loci in girls and 76 independent loci for the timing of

puberty in boys.

Isosexual precocity refers to premature sexual development consistent with phenotypic sex and includes features such as the development of facial hair and phallic growth. Isosexual precocity is divided

into gonadotropin-dependent and gonadotropin-independent causes

of androgen excess (Table 391-1). Heterosexual precocity refers to the

premature development of estrogenic features in boys, such as breast

development.

Gonadotropin-Dependent Precocious Puberty This disorder, called central precocious puberty (CPP), is less common in boys

than in girls. It is caused by premature activation of the GnRH pulse

generator, sometimes because of central nervous system (CNS) lesions

such as hypothalamic hamartomas, but it is often idiopathic. CPP is

characterized by gonadotropin levels that are inappropriately elevated

for age. Because pituitary priming has occurred, GnRH elicits LH

and FSH responses typical of those seen in puberty or in adults. MRI

should be performed to exclude a mass, structural defect, infection, or

inflammatory process. Mutations in kisspeptin, kisspeptin receptor,

and MKRN3, an imprinted gene encoding makorin ring finger protein

3, which is expressed only from the paternally inherited allele, have

been associated with CPP. Loss-of-function mutations in MKRN3

remove the brake that restrains pulsatile GnRH, resulting in precocious

puberty.

Gonadotropin-Independent Precocious Puberty Androgens

from the testis or the adrenal are increased but gonadotropins are low.

This group of disorders includes hCG-secreting tumors; CAH; sex

steroid–producing tumors of the testis, adrenal, and ovary; accidental or