You are on page 1of 7

LEADING ARTICLE 401

Determination of sex (through duplication of Xp21) and


....................................................................................... Wnt4 (through duplication of 1p35),
have been associated with impaired

Early assessment of ambiguous


gonadal development and undervirilisa-
tion in a small number of karyotypic 46
XY males.
genitalia Mutations or duplications in the
various genes responsible for gonadal
A L Ogilvy-Stuart, C E Brain differentiation and the subsequent
development of the internal and exter-
................................................................................... nal genital phenotype genes may be
responsible for gonadal dysgenesis and
A multidisciplinary problem in some cases complete sex reversal
(table 1).

T
o discover that there is uncertainty the differentiation of the gonad. The Wnt4 is also expressed in the
about the sex of one’s newborn differentiation of the gonad in turn Müllerian ducts and in the absence of
baby is devastating and often determines the development of both anti-Müllerian hormone (AMH) (also
incomprehensible for most parents. It the internal genital tracts and the known as Müllerian inhibiting sub-
is paramount that clear explanations external genitalia and thus phenotypic stance) and testosterone, Müllerian
and investigations are commenced sex, which occurs later in development structures develop, while the Wolffian
promptly, and that no attempt is made (from about 5–6 weeks of gestation). ducts involute.4 AMH promotes regres-
to guess the sex of the baby. Extreme Both male and female genitalia differ- sion of Müllerian structures and as the
sensitivity is required, and ideally the entiate from the same structures along only source of AMH in the fetus is the
baby should be managed in a tertiary the urogenital ridge. At about 4 weeks testes, the absence of a uterus in a baby
centre by a multidisciplinary team after fertilisation, primordial germ cells with ambiguous genitalia is evidence
including a paediatric endocrinologist migrate from the yolk sac wall to the that there has been functional testicular
and a paediatric urologist. Early involve- urogenital ridge that develops from the tissue (Sertoli cell) present. Testosterone
ment of a clinical psychologist with mesonephros. The urogenital ridge also produced from Leydig cells promotes
experience in this field should be man- contains the cells that are the precursors differentiation of the Wolffian ducts and
datory. Other professionals including for follicular or Sertoli cells and steroid hence the internal male genitalia (vas
geneticists and gynaecologists may also producing theca and Leydig cells. The deferens, epididymis, and seminal vesi-
become involved. There must be access ‘‘indifferent’’ gonads form on the geni- cles).
to specialist laboratory facilities and tal ridges. Testosterone is converted to dihydro-
experienced radiologists. The incidence The development of the fetal adrenals testosterone [DHT] by the enzyme 5a-
of genital ambiguity that results in the and gonads occur in parallel, as before reductase. DHT masculinises the exter-
child’s sex being uncertain is 1 per migration, the potential steroidogenic nal genitalia from about 6 weeks gesta-
4500,1 although some degree of male cells of both originate from the meso- tion, and the degree of masculinisation
undervirilisation, or female virilisation nephros. There are many genes and is determined by the amount of fetal
may be present in as many as 2% of live transcription factors that are expressed androgen present (irrespective of
births.2 in both tissues (for example, SF1 and source) and the ability of the tissues to
Parents require reassurance that DAX1), and hence mutations in these respond to the androgens.
either a male or female gender will genes may affect both adrenal and Defects in any part of this pathway
definitely be assigned. However the gonadal development (fig 1). In addi- (including gene mutations and chromo-
outcome of some of the investigations tion, WT1 is expressed in the kidney and somal abnormalities (for example,
may take some weeks, and registration gonad, hence the association of Wilms’ 46XY/46XX, 45X/46XY), inappropriate
of the child’s birth should be deferred tumour and gonadal dysgenesis in hormone levels, or end-organ unrespon-
until gender has been assigned. This Denys-Drash syndrome, for example. siveness) may result in genital ambi-
may require communication with the The undifferentiated gonad is capable guity, with undervirilisation of an XY
Registrar of Births, and a skilled clinical of developing into either an ovary or a individual, virilisation of an XX indivi-
psychologist will help the parents in testis. The theory that the ‘‘default’’ dual, or the very rare true hermaphro-
deciding what to tell family and friends programme generates an ovary is prob- dite (an individual with both ovarian
in the interim. It is also helpful (if ably not correct, although the exact role tissue with primary follicles and testi-
appropriate) to reassure the parents that of ‘‘ovarian determining’’ genes in cular tissue with seminiferous tubules
their child is otherwise healthy. humans is unclear at present. In con- which may be in separate gonads or
While not all intersex conditions are trast, testicular development is an active ovotestes).
apparent at birth (for example, complete process, requiring expression of the
androgen insensitivity may only become primary testis determining gene SRY, CLINICAL ASSESSMENT OF
apparent in a child with a testis within and other testis forming genes such as AMBIGUOUS GENITALIA
an inguinal hernia, or at puberty with SOX9. Transcription factors such as SF1 History
primary amenorrhoea and lack of and WT1 are also required for develop- The history should include details of the
androgen hair), only those presenting ment of the undifferentiated gonad, as pregnancy, in particular the use of any
with genital ambiguity at birth will be well as for the activation of the other
considered in this article. male pathway genes required for testis Abbreviations: AMH, anti-Müllerian hormone;
An understanding of sex determina- development and the consequent devel- CAH, congenital adrenal hyperplasia; DHEA,
tion and differentiation is essential to opment of male internal and external dehydroepiandrosterone; DHEAS,
dehydroepiandrosterone sulphate; DHT,
direct appropriate investigations and to genitalia.3 dihydrotestosterone; EUA, examination under
establish a diagnosis. DAX1 and Wnt 4 are two genes that anaesthesia; hCG, human chorionic
Genetic sex is determined from the may act to ‘‘antagonise’’ testis develop- gonadotrophin; StAR, steroidogenic acute
moment of conception and determines ment. Over-expression of DAX1 regulatory protein

www.archdischild.com
402 LEADING ARTICLE

drugs (table 2) that may cause virilisa-


tion of a female fetus and details of any
previous neonatal deaths (which might
point to an undiagnosed adrenal crisis).
A history of maternal virilisation may
suggest a maternal androgen secreting
tumour or aromatase deficiency. A
detailed family history should be taken,
including whether the parents are con-
sanguineous (which would increase the
probability of an autosomal recessive
condition) or if there is a history of geni-
tal ambiguity in other family members
(for example, an X-linked recessive con-
dition such as androgen insensitivity).

Examination
The general physical examination
should determine whether there are
any dysmorphic features and the gen-
eral health of the baby. A number of
syndromes are associated with ambig-
uous genitalia, for example, Smith-
Lemli-Opitz syndrome (characterised
by hypocholesterolaemia and elevated
7-dehydrocholesterol levels, and result-
ing from mutations affecting 7-dehy-
drocholesterol reductase), Robinow
syndrome, Denys-Drash syndrome, and
Beckwith-Wiedemann syndrome. Mid-
line defects may point towards a
hypothalamic-pituitary cause for micro-
penis and cryptorchidism.
Hypoglycaemia may indicate cortisol
deficiency secondary to hypothalamic-
pituitary or adrenocortical insufficiency.
The state of hydration and blood pres-
sure should be assessed as various forms
Figure 1 Normal sexual differentiation. SRY, sex determining region on Y chromosome; TDF,
of congenital adrenal hyperplasia (CAH)
testis determining factor; AMH, anti-Müllerian hormone; T, testosterone; DHT, dihydrotestosterone;
WT1, Wilms’ tumour suppressor gene; SF1, steroidogenic factor 1; SOX9, SRY-like HMG-box; can be associated with differing degrees
Wnt4, Wnt = a group of secreted signalling molecules that regulate cell to cell interactions during of salt loss, varying degrees of virilisa-
embryogenesis; DAX1, DSS-AHC critical region on the X chromosome. tion in girls or undervirilisation in boys,

Table 1 Consequences of mutations/deletions and duplications/translocations of genes involved in gonadal differentiation


Chromosome Gonadal Sex reversal/genital Müllerian
location development Associated disorder ambiguity development

Gene mutation or deletion (loss of function)


WT1 11p13 Dysgenesis (R and =) WAGR syndrome Genital ambiguity (=) Variable (=)
Denys-Drash syndrome Sex reversal or genital Variable (=)
ambiguity (=)
Frasier syndrome Sex reversal (=) Yes (=)
SF1 9q33 Dysgenesis (=) Adrenal failure Sex reversal (= Yes (=)
SRY Yp11.3 =R ovary Sex reversal or genital = (variable)
ambiguity (=)
DAX1 Xp21 = Dysgenesis Adrenal failure and No No
hypogonadotrophic
hypogonadism/impaired
spermatogenesis
SOX9 17q24.3–25.1 = Dysgenesis or Campomelic dysplasia Sex reversal or genital Variable
ovary/ovotestis ambiguity (=)
AMH 19p 13.3–13.2 Normal Yes (=)

Gene duplication or translocation (gain of function)


SRY Y fragment Yp11.3 RR testis Sex reversal or genital No
translocation ambiguity (R)
DAX1 duplication dupXp21 = Dysgenesis or Sex reversal or genital Variable
ovary/ovotestis ambiguity (=)
Wnt 4 duplication Dup 1p35 = Dysgenesis Genital ambiguity (=) Yes
SOX9 duplication dup17q24.3–25.1 RR Testis Genital ambiguity (R) No

WAGR, Wilms’ tumour, aniridia, genitourinary anomalies, mental retardation; Denys-Drash (exonic mutations) WT, diffuse mesangial sclerosis; Frasier (intronic
mutations) no WT, focal segmental glomerulosclerosis. Other abbreviations as for fig 1.

www.archdischild.com
LEADING ARTICLE 403

Table 2 Disorders of sex differentiation in babies with CAH. The gestation of the
baby can cause confusion—in preterm
Virilisation of XX females girls the clitoris and labia minora are
Increased fetal adrenal androgen production relatively prominent and in boys, the
Congenital adrenal hyperplasia (3b-hydroxysteroid dehydrogenase deficiency, 21-hydroxylase
deficiency, 11b-hydroxylase deficiency)
testes are usually undescended until
Androgen secreting tumour about 34 weeks gestation.
Placental aromatase deficiency
Fetal gonadal androgen production
True hermaphrodite with both testicular and ovarian tissue
INVESTIGATIONS
Transplacental passage of maternal androgens Initial investigations ascertain whether
Drugs administered during pregnancy: e.g. progesterone, danazol the child is an undervirilised male or a
Maternal androgen secreting tumour, luteoma of pregnancy virilised female and from these a differ-
Other causes ential diagnosis can be established and
Dysmorphic syndromes
Prematurity—prominent clitoris subsequent investigations planned
Bisexual gonads (fig 3). The principal aims of the initial
Hermaphroditism. Usual genotype 46XX investigations are to determine the most
appropriate sex of rearing and in a
Undervirilisation of XY males
Testicular dysgenesis/malfunction
genetic female, to exclude CAH and
Pure XY gonadal dysgenesis avert a salt losing crisis. Rarer forms of
Mixed gonadal dysgenesis–45X/46XY. May be associated with gene mutations on SRY, SOX9, or CAH (steroidogenic acute regulatory
WT1 genes protein (StAR) deficiency, 3b-hydroxys-
Dysgenetic testis
Testicular regression syndromes
teroid dehydrogenase deficiency) may
True agonadism, rudimentary testis syndrome also present with an adrenal crisis in
Biosynthetic defect—decreased fetal androgen biosynthesis undervirilised males.
Leydig cell hypoplasia (LH deficiency or LH receptor defect)
Testosterone biosynthesis (non-virilising CAH): (StAR, 3b-HSD, 17a-OHD/17–20 lyase, Smith-Lemli-
Opitz syndrome) Karyotype
5a-reductase deficiency As the differential diagnosis and a
Deficient synthesis or action of AMH—persistent Müllerian duct syndrome. May be due to mutations number of subsequent investigations
in AMH or AMH receptor gene or SF1 gene will depend on the genetic sex, an
End organ unresponsiveness
Androgen receptor and post-receptor defects (complete and incomplete androgen insensitivity urgent karyotype should be sent. Rapid
syndrome) FISH studies using probes specific for
Others the X (DX1) and Y (SRY) chromosomes
Urogenital malformations can provide useful information,
Dysmorphic syndromes
Exogenous maternal oestrogens
although a full karyotype is required
for confirmation and exclusion of
mosaicism. The latter may be tissue
specific and may not be apparent from
blood, but only from skin or gonadal
biopsies. In cases with a mosaic karyo-
or hypertension (table 3). Although the detect the presence of a gonad. They type, for example, XO/XY or XX/XY, the
cardiovascular collapse with salt loss may be situated high in the inguinal gonads are often different. This may
and hyperkalaemia in congenital adre- canal so careful examination is required. involve one being a streak gonad and
nal hyperplasia does not usually occur A unilateral palpable gonad may be a the other a testis or ovotestis, or two
until between the 4th to 15th day of life testis, ovotestis, or rarely an ovary ovotestes with different ovarian and
and so will not be apparent at birth in a within an inguinal hernia. Congenital testicular components.
well neonate, it should be anticipated adrenal hyperplasia must be excluded in
until CAH has been excluded. Jaundice a phenotypically male baby with bilat- Determination of internal anatomy
(both conjugated and unconjugated) eral undescended testes. As with the external genitalia, the
may be caused by concomitant thyroid The length of the phallus should be degree of virilisation of the internal
hormone or cortisol deficiency. The determined. The normal term newborn genital tracts is defined by Prader stages
urine should be checked for protein as penis is about 3 cm (stretched length I–V (fig 2). The anatomy of the vagina
a screen for any associated renal anom- from pubic tubercle to tip of penis) with or a urogenital sinus and uterus may be
aly (for example, Denys-Drash/Frasier micropenis less than 2.0–2.5 cm, determined by ultrasound, and if neces-
syndromes). although this does vary slightly depend- sary, further delineation by EUA/cysto-
ing on ethnic origin.5 Chordee should be scopy or urogenital sinogram.
Examination of external genitalia noted as this may decrease the apparent Ultrasound is also useful in excluding
and Prader staging length of the phallus and the penis may associated renal anomalies, particularly
Examination of the external genitalia be ‘‘buried’’ in some cases. The presence if Denys-Drash syndrome is suspected or
determines whether gonads are palpable of hypospadias and the position of the proven. It may also be used to visualise
and the degree of virilisation or Prader urethral meatus should be determined. the adrenal glands. Ultrasound may also
stage. Prader stages I–V describe The urine may be coming from more locate inguinal gonads, although it is
increasing virilisation from a phenotypic than one orifice. The degree of fusion not sensitive for intra-abdominal
female with mild cliteromegaly (stage I) and rugosity of the labioscrotal folds gonads. Ultrasound sensitivity and
to a phenotypic male with glandular should be noted and the presence or accuracy depend on probe resolution;
hypospadias (stage V) (fig 2). absence of a separate vaginal opening an experienced ultrasonographer is
If both gonads are palpable they are determined. required. Identification of the gonads
likely to be testes (or ovotestes) which Hyperpigmentation, especially of the may require magnetic resonance ima-
may be normal or dysgenetic. Flat finger genital skin and nipples occurs in the ging (MRI) or laparoscopy. These latter
palpation from the internal ring milking presence of excessive ACTH and pro- investigations are also useful for deter-
down into the labial scrotal folds may opiomelanocortin and may be apparent mining the anatomy of the internal

www.archdischild.com
404 LEADING ARTICLE

Table 3 Forms of CAH causing genital ambiguity in the newborn period, but AMH is undetectable in
female serum until the onset of puberty.
StAR protein AMH may prove to be a more sensitive
Type of CAH deficiency 3b-HSD 17a-OHD 11b-OHD 21-OHD marker for the presence of testicular
Genital = = = R R tissue than serum testosterone levels,
ambiguity both before and after the neonatal
Salt wasting Yes Yes Hypertension Hypertension Yes androgen surge, and, consequently,
Enzyme P-450cc 3b-HSD P-450c17 P-450c11 P-450c21
may obviate the need for hCG stimula-
Steroidsq None DHEA, DOC, DOC, 11- 17-OHP, A4
17-OH-preg corticosterone deoxycortisol tion in the evaluation of certain intersex
SteroidsQ All Aldosterone, Cortisol, Cortisol Cortisol, disorders.9 Similarly, basal inhibin B has
testosterone, testosterone aldosterone been shown to predict the testosterone
cortisol response to hCG in boys, and therefore
Defective gene StAR HSD3B2 CYP17 CYP11B1 CYP21
Chromosome 8p11.2 1p13.1 10q24.3 8q24.3 6p21.3
may give reliable information about
both the presence and function of the
StAR protein, steroidogenic acute regulatory protein deficiency, otherwise known as lipoid hyperplasia/ testes. Furthermore, inhibin B levels
cholesterol desmolase deficiency; 3b-HSD, 3b-hydroxysteroid dehydrogenase deficiency; 17a-OHD, have been shown to demonstrate spe-
17a-hydroxylase deficiency; 11b-OHD, 11b-hydroxylase deficiency; 21-OHD, 21-hydroxylase
cific alterations in patients with genital
deficiency; DOC, 11-deoxycorticosterone; DHEA, dehydroepiandrosterone; 17-OHP, 17-
hydroxyprogesterone; 17-OH-preg, 17-hydroxypregnenolone; A4, androstenedione. ambiguity which may aid the differen-
tial diagnosis of male undervirilisation.10
However, these assays are not routinely
available in the UK.
genitalia, and gonadal and genital skin (testosterone secretion normally rises
biopsies can be performed at the time of in the fourth week of life, peaking at Assessment of gonadotrophins
laparoscopy. 1–3 months6–8) and proceeding with the Assessment of the gonadotrophins may
investigations as promptly as possible. give useful information. Raised basal
Testosterone response to human While some would advocate that this gonadotrophins are consistent with pri-
chorionic gonadotrophin (hCG) test is deferred beyond 2 weeks of age, mary gonadal failure. The gonadotro-
To determine whether functioning this may not be necessary. In the new- phin response to GnRH is difficult to
Leydig cells are present (that is, capable born period an increase in testosterone, interpret in the prepubertal child unless
of producing testosterone in response to reaching adult male levels, would nor- exaggerated, which is consistent with
LH), an hCG stimulation test is under- mally be expected after hCG. gonadal failure but adds little to the
taken. This is also used to delineate a The hCG test can be extended to a basal levels alone. Pituitary failure
block in testosterone biosynthesis from three week test if the three day hCG would give a flat response but is not
androstenedione (17b-hydroxysteroid test is inconclusive. The same dose of diagnostic, and hypothalamic abnorm-
dehydrogenase deficiency) or conver- hCG is administered twice weekly for alities such as Kallmann’s syndrome are
sion of testosterone to DHT (5a-reduc- three weeks and testosterone, DHT, not excluded by a ‘‘normal’’ response
tase deficiency). There are a variety of and androstenedione samples taken The optimal time to assess this is within
protocols for the hCG test, but essen- 24 hours after the last hCG injection. the first six months of life when there is
tially it involves taking baseline samples The clinical response in terms of testi- a gonadal surge in both sexes (girls
for testosterone and its precursors dehy- cular descent and change in the size of greater than boys), and the axis is hence
droepiandrosterone (DHEA) (or DHEA the phallus and frequency of erections maximally responsive.
sulphate (DHEAS)) and androstene- should be documented. Photographs
dione and its metabolite (DHT). One to pre- and post-hCG, may be helpful.
Assessment of adrenal function
three intramuscular injections of high Urine steroid profile
dose hCG (500–1500 IU) are given at Anti-Müllerian hormone (AMH) and Output of adrenal steroids will be low in
24 hour intervals and repeat androgen inhibin B levels adrenal insufficiency. In CAH specific
samples are taken at 72 hours or While hCG stimulation tests the func- ratios of metabolites will be altered,
24 hours after the last injection. The tion of the Leydig cells, both AMH and depending on the level of the enzyme
neonatal gonad is more active than at inhibin B are secreted by the Sertoli block.
any time in childhood until puberty, cells. Inhibin B is detectable for the first
reaching a peak at about a 6–8 weeks of six months, rising again at puberty.
Synacthen test
age. A balance needs to be reached AMH levels are high in human male
A standard short Synacthen test is
between performing the hCG test when serum postnatally for several years
useful in the following situations:
the gonad is normally most active before declining during the peripubertal
N Suspected CAH where a peak 17-
hydroxyprogesterone (17-OHP) of
100–200 nmol/l is suggestive of 21-
hydroxylase deficiency. (Higher
reference ranges for preterm infants.)
N Suspected CAH to assess adrenal
cortical reserve (measurement of
cortisol levels).

The dose of Synacthen is dependent on


local protocol (example 36 mg/kg or a
standard dose of 62.5 mg intravenously
Figure 2 Differential virilisation of the external genitalia using the staging system of Prader, from or intramuscularly). Cortisol and 17-
normal female (left) to normal male (right). Sagittal (upper panel) and perineal (lower panel) views OHP levels are normally measured at 0
shown. and 30 minutes only as the 60 minute

www.archdischild.com
LEADING ARTICLE 405

CAH are DHEA, androstenedione, tes-


tosterone, ACTH, and plasma renin
activity, all of which may be increased.
From day 3 of life, the ratio of urinary
steroid metabolites will be altered
depending on the site of the block,
and is very helpful in the diagnosis of
21-OHD and the rarer forms of CAH. A
skilled ultrasound examination will
show normal internal genitalia, but an
EUA may be required to confirm the
presence of normal Müllerian struc-
tures, and to show the level of entry
of the vagina into the urogenital
sinus, in the case of a single perineal
opening.
The excessive androgen may be gona-
dal in origin—usually from an ovotestis
or testis in a true hermaphrodite. While
the commonest karyotype of the true
hermaphrodite is 46XX (70.6%), the
next commonest karyotype is a chromo-
somal mosaicism containing a Y chro-
mosome (usually 46XX/46XY) (20.2%).11
Figure 3 Investigation flow plan for assessment of ambiguous genitalia.
It is important to check fibroblast and/or
gonadal genotype in these babies, which
may contain a mosaic cell line.
Transplacental transfer of androgen
value does not usually contribute The androgens may be derived from the may rarely occur if the mother has an
further. A normal basal value, or even fetal adrenal gland (CAH and placental androgen secreting tumour (she may be
a normal stimulated response does not aromatase deficiency), fetal gonad (the virilised as a result) or from drugs given
exclude the evolution of adrenal insuf- testis or ovotestis in true hermaphrodit- during pregnancy. Placental aromatase
ficiency, and may need to be repeated ism), or rarely, exogenously via trans- deficiency, by inhibiting the conversion
depending on clinical suspicion. A basal placental passage from the mother of androgens to oestrogens, may cause
ACTH level may be helpful, but in most (adrenal or ovarian tumours). virilisation of a female fetus, and in
laboratories the turnaround time is Absence of palpable gonads in addition, maternal virilisation occurs
slower than for cortisol. association with otherwise apparently from placental transfer of the excessive
male genitalia should always alert one fetal androgens.12
Skin and gonadal biopsies to the possibility of a virilised female. By Table 2 lists conditions that cause
Genital skin biopsies (2–4 mm) per- far the commonest cause of this is virilisation of a female fetus.
formed at the time of examination CAH. Of the enzyme defects that
under anaesthesia (EUA) or genito- cause virilisation in female fetuses, 21- Genital ambiguity with a 46XY
plasty are useful to establish cell lines hydroxylase deficiency is the most fre- karyotype indicates an
for androgen receptor binding assays quent (accounting for 90–95%, UK undervirilised male
and analysis of 5a-reductase activity. incidence 1/15–20 000). Diagnosis is This is a genetically XY male with two
The cell line is also a source of genomic made on a raised serum 17-hydroxypro- testes, but whose genital tract fails to
DNA and RNA and/or subsequent mole- gesterone level. This level may be differentiate normally. There are numer-
cular and functional studies. Karyotype increased in the first 48 hours in nor- ous presentations of genital anomaly,
analysis for the presence of mosaicism mal babies, and may be significantly from apparently normal female (Prader
may be indicated. Gonadal biopsies are increased in sick and preterm babies in I) with a palpable gonad through to an
essential when considering diagnostic the absence of CAH. The other enzyme apparently normal male with hypospa-
categories such as dysgenesis and true defects that can cause virilisation of dias (Prader V).
hermaphroditism. A detailed histo- female fetuses are 11b-hydroxylase The three main diagnostic categories
pathological report is essential. As spe- deficiency (where the 11-deoxycortisol are: testicular dysgenesis/malfunction, a
cial treatment of the samples may be levels will be increased) and less com- biosynthetic defect, and end-organ
required, prior discussion with the monly 3b-hydroxysteroid dehydrogen- unresponsiveness (table 2).
pathologist or genetics laboratory ase deficiency (diagnosed by raised If the gonads are palpable, they are
should take place. 17-hydroxypregnenolone and dehydro- likely to be testes (or rarely ovotestes).
epiandrosterone) (table 3). Investigations are directed at determin-
INTERPRETATION OF RESULTS If CAH is confirmed, the electrolytes ing the anatomy of the internal genita-
Genital ambiguity with a 46XX need to be watched closely as salt lia, and establishing whether the
karyotype indicates a virilised wasting occurs in 70% of cases of 21- testicular tissue is capable of producing
female hydroxylase deficiency, usually between androgens. Investigations that may help
The female fetus with ovaries and days 4 and 15. Mineralocorticoid defi- with the former include pelvic ultra-
normal internal genitalia has been ciency induces a rise in serum potas- sound, examination under anaesthetic
exposed to excessive testosterone, and sium levels (usually the first sign of salt with cystoscopy and laparoscopy.
hence dihydrotestosterone (by conver- wasting) and sodium levels fall. Urinary Genital skin biopsy specimens can be
sion of testosterone by 5a-reductase) sodium levels will be inappropriately taken at the time of endoscopies.
which virilises the external genitalia. high. Further confirmatory tests for Occasionally urogenital sinogram or

www.archdischild.com
406 LEADING ARTICLE

MRI can be helpful. Laparoscopy/laparo- This condition is rare in the UK, but consistent with X linkage. The majority
tomy and gonadal biopsy may be recognised in the Dominican Republic, of XY infants with undervirilisation
required. where individuals are often raised as remain unexplained.
female and convert to male in puberty,
Testicular dysgenesis/malfunction when body habitus and psychosexual
DNA ANALYSIS
A 46 XY karyotype, with low basal and orientation becomes male. Virilisation
Many of the causes of genital ambiguity
hCG stimulated testosterone and low improves but is incomplete.
have a genetic basis, and in these cases
testosterone precursors suggests A biochemical diagnosis is made by
genetic counselling will be required. For
either gonadal dysgenesis (which may showing a ratio of T:DHT .30 after
example, androgen insensitivity syn-
require laparoscopy and testicular puberty or following hCG stimulation
biopsy) or lipoid CAH (caused by an drome is X-linked recessive, and CAH
before puberty, and the ratio of 5a:5b
abnormality in the steroidogenic acute is autosomal recessive. In addition,
metabolites in a urine steroid profile will
regulatory (StAR) protein) identification and characterisation of a
be increased after 6 months of age. The
In the latter condition total adrenal number of mutations of the genes
urinary 5a:5b metabolites can also be
failure is confirmed on Synacthen test, used if the gonads have been removed. involved in sexual differentiation has
electrolytes, and urinary steroids. The diagnosis is confirmed by screening resulted in DNA tests which can be
Because of the other associated gene for mutations in the 5a-reductase type II used in prenatal diagnosis. Iden-
defects (table 1), there may be other gene (5RD5A2). tification of carriers will facilitate
anomalies such as bony dysplasias or genetic counselling.
renal anomalies, which should be A 46XY karyotype, low basal and hCG
looked for. stimulated testosterone levels with Congenital adrenal hyperplasia
The poorly functioning testicular tis- increased testosterone precursors Several laboratories within the UK will
sue is likely to give a subnormal indicates a testosterone biosynthetic undertake DNA analysis for this condi-
testosterone response to hCG and basal defect tion. Once the diagnosis has been made,
gonadotrophins will usually be Those forms of CAH that cause under- a DNA sample from the proband should
increased, consistent with primary virilisation of male genitalia include be taken. If a gene mutation is identi-
gonadal failure. In addition, the dysge- 17a-hydroxylase/17,20-lyase deficiency fied, the carrier status of the parents
netic testes may secrete inadequate and 3b-hydroxysteroid dehydrogenase may be determined and the family
amounts of anti-Müllerian hormone, deficiency (table 3). should be counselled about the possibi-
and Müllerian structures may be present The conversion of androstenedione to lity of antenatal screening of subsequent
(although often hypoplastic) in children testosterone occurs predominantly in pregnancies, and of steroid treatment of
with gonadal dysgenesis and a 46XY the gonad and a post-hCG stimulated the mother in an attempt to reduce
karyotype. ratio of androstenedione to testosterone virilisation of a subsequent affected
A mosaic karyotype, for example 45X/ of .20:1 suggests 17b-hydroxysteroid female fetus. In the UK, antenatal
46XY suggests gonadal dysgenesis. dehydrogenase deficiency. A urine ster- diagnosis and treatment is offered as
There is a very variable phenotype both oid profile is generally not helpful in this part of a national British Society for
in terms of internal and external geni- diagnosis before puberty. Molecular Paediatric Endocrinology and Diabetes
talia, which is not dependent on the analysis of the HSD III gene supported study monitoring efficacy,
percentage of each karyotype as deter- (HSD17B3) is sought as confirmation short and long term side effects, and
mined by lymphocyte analysis. Over of the diagnosis. outcome measures.
90% of individuals with prenatally diag-
nosed 45X/46XY karyotype have a nor- End-organ unresponsiveness
mal male phenotype, suggesting most A 46XY karyotype with genital Androgen insensitivity
individuals with this karyotype escape ambiguity, normal or increased basal DNA analysis is now being undertaken
detection and that an ascertainment and peak testosterone on hCG test, in Cambridge as part of a molecular
bias exists towards those with clinically and a normal T:DHT ratio points to genetics service in collaboration with
evident abnormalities.13 14 partial androgen insensitivity Professor Ieuan Hughes. Samples are
There is a variable phenotype, and sex of only analysed following collection of
Biosynthetic defect rearing depends on the degree of phallic clinical, biochemical, and histological
A 46XY karyotype, low basal and peak development, and sometimes cultural data that are consistent with an andro-
testosterone level on hCG testing, considerations. The child may benefit gen insensitive pathophysiology.
often with increased gonadotrophins from a trial of topical DHT cream or
suggests a diagnosis of an inactivating intramuscular testosterone (for example
Suspected 5a-reductase deficiency
mutation of the LH receptor (Leydig using 12.5–25 mg, monthly for three
and 17b-hydroxysteroid
cell hypoplasia) months) on penile growth to help
dehydrogenase deficiency
This condition is associated with a anticipate response in puberty.
DNA samples in patients with suspected
variable phenotype from a completely The diagnosis is suggested by showing
5a-reductase deficiency and 17b-hydro-
phenotypic female to undervirilisation an abnormality in androgen binding in
xysteroid dehydrogenase deficiency can
of varying degrees.15 genital skin fibroblasts, or a mutation in
also be processed through the
the androgen receptor gene. This
Cambridge laboratory.
A 46XY karyotype with normal or requires DNA and a genital skin biopsy,
increased basal and peak testosterone taken at the time of EUA or genital
on hCG test, and an increased T:DHT surgery. Despite clear evidence of a Unusual cases of sexual ambiguity
ratio is seen in 5a-reductase phenotype consistent with partial Mutations of developmental genes such
deficiency androgen PAIS and normal production as DAX1, SOX9, and WT1 may account
DHT dependent virilisation of external and metabolism of androgens, only a for rarer cases of sexual ambiguity.
genitalia is deficient, resulting in a small minority of patients are found to have Samples should be taken and DNA
phallus and perineal hypospadias. an androgen receptor gene mutation. extracted and either stored or forwarded
Wolffian structures are normal but The likelihood of finding a mutation is to the relevant laboratories, depending
spermatogenesis is usually impaired.16 increased if there is a family history on clinical suspicion.

www.archdischild.com
LEADING ARTICLE 407

ASSIGNMENT OF SEX OF CONCLUSIONS 3 Ahmed SF, Hughes IA. The genetics of male
undermasulinization. Clin Endocrinol
REARING Genital ambiguity resulting in uncer- 2002;56:1–18.
A decision about the sex of rearing tainty of sex at birth is uncommon. The 4 Vainio S, Heikkila M, Kispert A, et al. Female
should be made as soon as is practic- most frequent cause in a genetic female development in mammals is regulated by Wnt-4
signalling. Nature 1999;379:707–10.
able, usually based on the internal and is CAH, which may be life threatening if 5 Cheng PK, Chanoine JP. Should the definition of
external genital phenotype and the there is a risk of a salt losing crisis. micropenis vary according to ethnicity? Horm Res
results of the various investigations. Reaching a diagnosis, particularly in 2001;55:278–81.
6 Winter JS, Hughes IA, Reyes FI, et al. Pituitary-
Cultural aspects may also be important undervirilised males, is currently not gonadal relations in infancy: 2. Patterns of serum
in cases of severe ambiguity. always possible, but many may have a gonadal steroid concentrations in man from birth
Assignment of sex of rearing can be to two years of age. J Clin Endocrinol Metab
partial androgen insensitivity syndrome 1976;42:679–86.
extremely difficult, particularly as there yet to be defined in molecular terms or 7 Ng KL, Ahmed SF, Hughes IA. Pituitary-gonadal
is a paucity of data on long term milder variants of testicular dysgenesis. axis in male undermasculinisation. Arch Dis Child
outcome in this area. 2000;82:54–8.
Prompt counselling and investigations 8 Davenport M, Brain C, Vandenberg C, et al. The
In the case of a virilised female (with the backup of recognised bio- use of the hCG stimulation test in the endocrine
(usually CAH), there is usually the chemical and genetic laboratories) is evaluation of cryptorchidism. Br J Urol
potential for fertility and these babies 1995;76:790–4.
essential. The decision of sex of rearing 9 Rey RA, Belville C, Nihoul-Fekete C, et al.
are usually raised as girls. This is much and the timing of surgery need careful Evaluation of gonadal function in 107 intersex
easier if the diagnosis of CAH is made consideration within a multidisciplinary patients by means of serum antimullerian
early. hormone measurement. J Clin Endocrinol Metab
environment with full informed consent 1999;84:627–31.
Decisions about nature and timing of of the family. 10 Kubini K, Zachmann M, Albers N, et al. Basal
any surgery are made with the family inhibin B and the testosterone response to
acknowledging the considerable psycho- human chorionic gonadotropin correlate in
logical impact of having a child with ACKNOWLEDGEMENTS prepubertal boys. J Clin Endocrinol Metab
We thank Professor Ieuan Hughes and Dr 2000;85:134–8.
genital ambiguity. There is considerable 11 Krob G, Braun A, Kuhnle. True hermaphroditism:
John Achermann for their helpful comments. geographical distribution, clinical findings,
debate as to the optimal timing of any
Email address for Professor Hughes for chromosomes and gonadal histology. Eur J Pediatr
genital surgery.17 In the presence of mutation analysis and discussion on the 1994;153:2–10.
marked clitoromegaly, clitoral reduction androgen receptor, 5a-reductase deficiency, 12 Shozu M, Akasofu K, Harada T, et al. A new
is usually undertaken in infancy. Lesser and 17b-hydroxysteroid dehydrogenase: cause of female pseudohermaphroditism:
degrees of clitoral enlargement may be placental aromatase deficiency. J Clin Endocrinol
iah1000@cam.ac.uk. Email address for Dr
Metab 1991;72:560–6.
left until puberty when the child can be Achermann for mutation analysis and dis- 13 Chang HJ, Clark RD, Bachman H. Chromosome
involved with the decision making. The cussion on early gonadal genes or cases mosaicism in 6,000 amniocenteses. Am J Med
timing of any vaginoplasty is dependent associated with adrenal failure: j.achermann Genet 1989;32:506–13.
@ich.ac.uk. 14 Wilson MG, Lin MS, Fujimoto A, et al. The
on the anatomy of the internal and phenotype of 45,X/46,XY mosaicism: an analysis
external genitalia and influenced by Web address for the British Society for
of 92 prenatally diagnosed cases. Am J Hum
Paediatric Endocrinology and Diabetes: Genet 1990;46:156–67.
local practice. However, there has been www.bsped.org.uk. 15 Laue L, Wu S-M, Kudo M, et al. A nonsense
a move away from early vaginoplasty in mutation of the human luteinizing hormone
all infants. It may be appropriate to Arch Dis Child 2004;89:401–407. receptor gene in Leydig cell hypoplasia. Hum Mol
delay surgery until puberty when a doi: 10.1136/adc.2003.011312 Genet 1995;4:1429–33.
16 Hochberg Z, Chayen R, Reiss N, et al. Clinical,
single stage reconstruction can be
...................... biochemical, and genetic findings in a large
undertaken. Recent outcome data on pedigree of male and female patients with 5
clitoral sensation18 and success of early Authors’ affiliations alpha-reductase 2 deficiency. J Clin Endocrinol
A L Ogilvy-Stuart, Neonatal Unit, Rosie Metab 1996;81:2821–7.
vaginoplasty19 20 in adult women who
Hospital, Addenbrooke’s NHS Trust, 17 Creighton S. Surgery for intersex. J R Soc Med
underwent genital surgery in infancy Cambridge CB2 2SW, UK 2001;94:218–20.
and childhood has induced a more C E Brain, Dept of Paediatric Endocrinology, 18 Minto CL, Liao LM, Woodhouse CRJ, et al. The
cautious surgical approach.21 22 effect of clitoral surgery on sexual outcomes in
Great Ormond Street Hospital, Great Ormond individuals who have intersex conditions with
The appropriate sex of rearing of a Street, London WC1N 3JH, UK ambiguous genitalia: a cross sectional study.
very undervirilised male requires as Lancet 2003;361:1252–7.
much information as possible, from the 19 Alizai NK, Thomas DF, Lilford RJ, et al.
Correspondence to: Dr A L Ogilvy-Stuart, Feminizing genitoplasty for congenital adrenal
investigations discussed, as well as Neonatal Unit, Rosie Hospital, Addenbrooke’s hyperplasia: what happens at puberty? J Urol
thorough, multidisciplinary discussions NHS Trust, Cambridge CB2 2SW, UK; 1999;161:1588–91.
involving the parents, urologist, endo- amanda.ogilvy-stuart@addenbrookes.nhs.uk 20 Creighton S, Minto C, Steele SJ. Feminizing
childhood surgery in ambiguous genitalia:
crinologist, geneticist, and a clinical objective cosmetic and anatomical outcomes in
psychologist. It may be appropriate to REFERENCES adolescence. Lancet 2001;358:124–5.
defer gender assignment until the 21 Creighton S, Minto C. Managing intersex. BMJ
1 Hamerton JL, Canning N, Ray M, et al. A 2001;323:1264–5.
results of relevant investigations are cytogenetic survey of 14,069 newborn infants. I. 22 Creighton S, et al. Regarding the consensus
reviewed and the effects of exogenous Incidence of chromosome abnormalities. Clin statement on 21-hydroxylase deficiency from the
androgens have been assessed. The birth Genet 1975;4:223–43. Lawson Wilkins Paediatric Endocrine Society and
2 Blackless M, Charuvastra A, Derryck A, et al. the European Society for Paediatric
registration should be delayed for as How sexually dimorphic are we? Am J Hum Biol Endocrinology [letter]. Hormone Res
long as it takes for the final decision to 2000;12:151–66. 2003;59:262; and J Clin Endocrinol Metab
be made. 2003;88:3455.

www.archdischild.com

You might also like