Sex Hormones In Females And Related Disorders

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1 Sex Hormones In Females And Related Disorders (see previous newsletter article for males) Sex Hormone production and measurement in females Follicle Stimulating Hormone () Luteinizing Hormone (), Oestradiol and Progesterone The hypothalamus produces gonadotropin-releasing hormone (GnRH) which stimulates anterior pituitary production of and ; stimulates ovarian follicle development and oestrogen production during the follicular phase; the midcycle peak of stimulates ovulation (ovulation occurs about 24 hours after peak ) and thereafter stimulates progesterone production in the luteal phase by the corpus luteum (see Figure 1 opposite). This complex and varying hormone pattern can make interpretation difficult for the occasional requester. In particular, specimens taken around midcycle may be problematic as the normal rise in (and resulting increased / ratio) may be misdiagnosed as Polycystic Ovary Syndrome (PCOS) and if, as occasionally happens at midcycle, and both rise similarly, menopause or ovarian failure may be misdiagnosed (for typical follicular, midcycle and luteal patterns see tables opposite). Timing of specimen: For the above reasons a day 2-4 specimen is best for assessing the sex hormones for most purposes unless proof of ovulation is required. In this case a day 21 (assuming a 28 day cycle) progesterone of greater than 20 nmol/l confirms ovulation. If cycle length is usually longer or shorter than 28 days, take the sample 7 days before the usual end of the cycle. This is because the luteal phase generally takes 14 days regardless of overall cycle length. Endocrine Cycle Ovarian Histology Endometrial Histology Body Temp. ºC Follicular recruitment Menses Follicular Phase Days Typical Follicular Pattern 4 U/L (1-10) 2 U/L (2-7) Oest 255 pmol/l (77-920) Prog 0.5 nmol/l (<1.0) Typical Midcycle Pattern 11 U/L (6-17) 42 U/L (9-74) Oest 832 pmol/l ( ) Prog 1 nmol/l (<1.0) Typical Luteal Pattern 3 U/L (1-9) 3 U/L (1-9) Oest 450 pmol/l ( ) Prog 25 nmol/l ( ) Dominant follicle Figure 1. Monthly Ovarian Cycle Adapted from Harrisons Principles of Internal Medicine, 11th ed. Ovulation Luteal Phase Early morning specimens have higher androgen (due to adrenal diurnal rhythm) and prolactin levels than afternoon specimens. Prolactin levels are also higher during midcycle. Useful hints: E 2 Corpus luteum Remember to request a sex hormone binding globulin (SHBG) level if testosterone is measured (most testosterone is bound to SHBG and is thus inactive). This enables the free testosterone or free androgen index (FAI) to be calculated; these are more useful than testosterone alone. Hormones are measured using immunological techniques and this means that results using different methods are not interchangeable. P

2 This is particularly important with HCG levels when used to diagnose or monitor tumours. Most HCG assays used for pregnancy assessment are not suitable for tumour use as tumour-derived HCG is often structurally different to pregnancy-derived HCG; tumours may be missed if the reason for the test request is not clear on the request form. Clinipath Pathology, however, uses an HCG assay (Roche) which has been certified for both pregnancy and tumour use. Prolactin and Galactorrhoea Prolactin is produced by the anterior pituitary; it rises during pregnancy and functions by stimulating milk production and inhibiting and secretion. This inhibits ovulation and fertility which is desirable during the perinatal period and during breast-feeding. After delivery prolactin elevations are related to episodes of suckling. Pathological elevation of prolactin is the commonest hypothalamic-pituitary endocrine disorder and can cause galactorrhoea, reduced libido, amenorrhoea and infertility and will ultimately result in osteoporosis if not treated. Mild to moderate prolactin elevations (<2000 miu/l) in the non-pregnant/non-breast-feeding patient are usually due to stress, midcycle, medications (particularly psychotropic medications eg antipsychotics and antidepressants; H2 antagonists eg cimetidine; calcium channel blockers & oestrogens) PCOS and breast stimulation and less commonly due to hypothyroidism, pituitary stalk damage, renal failure and acromegaly. With these moderate elevations we advise a repeat sample taken a week later after resting 20 minutes, and if unexplained elevation persists, prolactinoma should be excluded by pituitary testing (TSH, T4, growth hormone, IGF1,,, oestradiol, progesterone and 9am cortisol) and pituitary imaging. Psychiatric patients on antipsychotics should have prolactin checked routinely as chronic elevations over 1000 miu/l inhibit ovulation, reduce oestrogen levels and cause osteoporosis. Consideration should be given to changing medications if levels exceed 1000 miu/l. Elevations of over 2000 miu/l are usually due to microprolactinomas, while elevations of greater than 5000 and typically around miu/l are seen in macroprolactinomas (tumours >1cm), however, antipsychotic medications and anti nausea drugs (eg stemetil and domperidone) can also occasionally cause elevations of this order. Macroprolactin is a confusingly named condition which describes a chronic elevation of prolactin due to incidental binding to it by antibodies in the patient s circulation. These antibodies prevent prolactin being cleared; prolactin levels thus rise but the hormone-antibody complex is generally inactive. Macroprolactin is thus usually considered an incidental and not a pathological finding. At Clinipath Pathology we routinely exclude macroprolactin if the prolactin level is over 1000 IU/L, or if it is repeatedly elevated to a lesser extent. Galactorrhoea is considered abnormal if it persists more than 6 months after delivery or discontinuation of breast-feeding. Bloody or non-milky discharges are not considered to be true galactorrhoea and local breast pathology, including possible breast cancer, should be excluded. True galactorrhoea is usually associated with elevated prolactin (see above for causes) and fasting morning specimens should be checked for this. Those women with galactorrhoea, normal menses and normal prolactin probably have increased sensitivity of the breast to normal prolactin levels. Investigation of galactorrhoea: Morning prolactin and IGF1 (rarely acromegally can present with galactorrhoea and normal prolactin) should be checked. If prolactin is elevated, then TSH, T4, U&Es, GH, IGF1, cortisol, oestradiol and progesterone should be measured. Pituitary imaging may also be required. Puberty and its disorders in girls Normal Puberty: The normal onset of puberty is between 8-13 years and the mean age of menarche in Western populations is years. Puberty occurs because of rising levels of sex hormones. and oestradiol levels may be fairly high in the neonatal period (and may even cause isolated breast development in some infant girls) but are then suppressed during childhood till the onset of puberty. Normal sequence and timing of puberty: Breast development may be the earliest manifestation of puberty Clinipath Pathology Newsletter / Summer 2011

3 (precedes sex hair development in 60% of girls) and is due to conversion of adrenal androgens to oestrogens. Increased secretion of adrenal steroids (adrenarche) especially DHEA, results in increased sex hair and apocrine sweat odour. Ovarian maturation (gonadarche) normally follows adrenarche. Menarche usually follows 2-3 years after the onset of breast development, and cycles are usually irregular for about a year varying between 21 to 45 days. Most cycles are regular by 2-4 years post menarche. Initiation of puberty: Puberty is triggered by initiation of pulsatile release of GnRH from the hypothalamic pulse generator (approximately neurones derived from the olfactory bulb) which then stimulates and release and consequent ovarian development. It is thought that increasing insulin resistance in childhood initiates pulsatile GnRH release; this view is supported by the findings that obese girls experience puberty sooner than normals; that menarche occurs at a critical body weight rather than age; and that the age of menarche is now younger than in previous generations. Apart from the influence of body weight, genetic factors account for much of the variation in age of puberty both in a familial and ethnic context (eg African Americans enter puberty a year before Caucasian girls). Precocious puberty in girls: This was defined (some 40 years ago in the UK) as the development of secondary sexual characteristics before 8 years of age. The earlier onset of puberty today has resulted in recommendations that this be reduced to less than 7 years in Caucasians and less than 6 years in African Americans. Causes: Precocious puberty may be GnRH mediated (ie accelerated normal mechanism of puberty) or GnRH independent (pseudopuberty). GnRH mediated: Early maturation of the GnRH mechanism occurs., and oestradiol levels are elevated for age. Most cases of precocious puberty (85%) are due to idiopathic early activation of GnRH pulsatile secretion (commoner in girls than boys) which results in early sexual development, increased growth rate and advanced bone age. Treatment with GnRH agonists down-regulates pituitary and secretion, slows progression of puberty and thus can prevent early menarche and closure of the epiphyses and short adult stature. PCOS may cause early puberty by increasing insulin resistance as outlined above. Occasionally lesions of the hypothalamus (tumours, infections, inflammation) may trigger onset of puberty by prematurely triggering GnRH release from the hypothalamic pulse generator. GnRH independent (pseudopuberty): In these cases, excess sex hormone production is not due to early maturation of the normal mechanism but to other causes. Oestrogens and/or androgens are elevated but and are suppressed. GnRH agonists are thus ineffective and other drugs e.g. oestrogen receptor blockers are required if the primary cause cannot be eliminated. Congenital adrenal hyperplasia: excess adrenal androgens are produced due to an enzyme deficiency in the pathway to cortisol synthesis. 21 hydroxylase deficiency is commonest and 17 OH progesterone is thus elevated. Presentation in girls is with hirsutism and virilisation, which may mimic precocious puberty. Oestrogen or androgen producing tumours of ovary or adrenal HCG producing tumours (HCG stimulates ovarian oestrogen production) McCune Albright syndrome (due to a G protein defect, and receptors in the ovary are constitutively active and oestrogen production is stimulated.) Exogenous oestrogen and androgen intake. (Foods, medications, alternative therapies).

4 Investigation of precocious puberty: U&Es, TFTs,,, oestradiol, testosterone, SHBG, DHEAS, androstenedione, 17 OH progesterone. Bone age should also be checked. Imaging may be required depending on findings. Delayed puberty: This is defined as the absence of breast development by 13 years or the absence of menarche by age 16 years. Causes are either failure of the hypothalamic/ pituitary initiation mechanism (hypogonadotrophic) or ovarian failure (hypergonadotrophic). Hypogonadotrophic hypogonadism: Low oestrogen, gonadotrophins are normal or low. Causes 1 & 2 are commonest, others are rare. Constitutional: family history of delayed puberty. Functional: Chronic diseases, hypothyroidism, diabetes, malnutrition, anorexia, excessive exercise. Kallmans syndrome (failure of development of hypothalamic GnRH pulse generator; usually with anosmia, more common in males than females) Tumours/inflammatory lesions of hypothalamus or pituitary (craniopharyngioma, prolactinoma) Hypergonadotrophic hypogonadism: Low oestrogen, elevated and due to ovarian dysgenesis or failure, or defects of steroid hormone synthesis or signalling in the pituitary/gonadal axis. Turners syndrome is the commonest cause. Turner Syndrome (streak ovaries, XO, short stature, webbed neck) Other causes of gonadal dysgenesis Autoimmune oophoritis Previous radiotherapy/ chemotherapy Defects in and receptors or sex steroid synthesis Complete androgen resistance syndromes (patient is XY male but phenotypically female, testosterone levels are in the normal male range.) Investigation of delayed puberty: U&Es, LFTs, TFTs,,, oestradiol, testosterone, SHBG, progesterone, karyotyping, bone age. Amenorrhoea, Menopause and Premature ovarian failure Primary Amenorrhoea: (failure to menstruate by 16 years). The causes of primary amenorrhoea are the same as those causing delayed puberty (covered above) and in addition include anatomical defects of the genitourinary tract. Secondary amenorrhoea: The commonest cause is pregnancy! Investigation of Amenorrhoea: Amenorrhoea lasting more than 3 months should be investigated by U&Es, LFTs, FBC, TFTs, HCG, prolactin,,, oestradiol, testosterone, SHBG, androstenedione and DHEAS. Hormone patterns in amenorrhoea Patients with high oestradiol and progesterone and suppressed and are usually pregnant. The gonadotrophins are suppressed because HCG is driving oestrogen and progesterone synthesis which in turn suppress and. Occasionally this pattern is difficult to distinguish from atypical luteal ovulatory profiles and an HCG check is thus always advisable if not already requested. Clinipath Pathology Newsletter / Summer 2011

5 Pattern A: Typical Pregnancy <1 U/L <1 U/L Oestradiol 3540 pmol/l Progesterone 95 nmol/l Patients with low oestradiol and gonadotrophins have hypothalamic amenorrhoea and causes such as systemic illness, stress, excess exercise, eating disorders, hyperthyroidism, drugs of abuse, pituitary/hypothalamic pathology (especially prolactinoma) must be excluded. This pattern is also typical of oral contraceptive use. Pattern B: Typical Hypothalamic amenorrhoea 1 U/L 1 U/L Oestradiol 40 pmol/l Progesterone 0.3 nmol/l Patients with reduced SHBG and or increased androgens with/without elevated / ratios should be assessed for PCOS. Pattern C: Typical PCOS Testosterone 2.1 nmol/l ( ) SHBG 12 nmol/l (26-110) FAI 17.5 ( ) Free Testosterone 61 pmol/l (3-33) Patients with low oestradiol and elevated are likely to have ovarian failure as typically seen in the menopause. Pattern D: Typical Ovarian Failure or menopause 35 U/L 16 U/L Oestradiol 95 pmol/l A precursor of this pattern is a normal or even high oestradiol with elevated (and sometimes lesser elevation) that is common in the early stages of ovarian failure and menopausal transition and reflects decreasing ovarian responsiveness to. This last-described pattern may be difficult to differentiate from midcycle, however, at midcycle, elevation is usually higher than elevation. Pattern E: Typical Early Ovarian Failure 23 U/L 12 U/L Oestradiol 1423 pmol/l Menopause is defined as the time of the cessation of menses and in Western populations now occurs at a mean of years. Age of menopause is largely genetically determined. Multiparity increases and lower socio-economic status and smoking reduce this age. Premature ovarian failure (POF) is said to occur if onset of failure is before 40 years age and may affect up to 1% of women. Premature ovarian failure may be idiopathic or secondary to ovarian insults (radiotherapy, chemotherapy and possibly mumps) or to the inherited auto-immune polyglandular syndrome which is associated with ovarian and adrenal antibodies and failure of other endocrine systems. A GTT, calcium, TFTs, and 9am cortisol should thus be checked together with ovarian and adrenal antibodies. Note that onset of ovarian failure is frequently relapsing and occasional women may conceive spontaneously after the onset of POF. Androgens, hirsutism, virilisation and PCOS Female androgen production: Most serum testosterone is bound to SHBG and is inactive, thus it is essential to measure SHBG as well as testosterone to try and estimate the free, active fraction (FAI and calculated free testosterone). In normal adult women, about half of testosterone production is ovarian and the remainder adrenal. Ovarian testosterone production decreases at menopause but SHBG also falls due to low oestrogens and the net effect is an increase in the relative amount of free testosterone which may cause hirsutism after menopause. Other androgens are DHEAS (mainly adrenal origin) and androstenedione (adrenal and ovarian origin). Clinical setting: About 10% of women are hirsute (ie. have male pattern hair growth) and most of these are familial/idiopathic or ethnic (darker individuals and Mediterranean origin). Patients with virilisation have in addition acne, male habitus, male pattern balding, deep voice, clitoromegally, etc. Virilisation (unlike hirsutism alone) is usually due to significant pathology. Sudden onset of hirsutism and virilisation is almost always due to androgen secreting tumours. Very high androgen levels are also suggestive of tumour production. Causes of hirsutism and virilisation: PCOS (by far the most common cause) Congenital Adrenal Hyperplasia (mild form of 21 hydroxylase deficiency; patients have elevated 17 OH progesterone) Cushings

6 Androgen-secreting tumours of ovary or adrenal Partial form of Testicular Feminisation (XY male with partial androgen resistance e.g. 800 metre gold medallist Caster Semenya) Exogenous androgens Investigations of hirsutism and virilisation: Prolactin, TFTs, Testosterone, SHBG, DHEAS, androstenedione and 17 OH progesterone together with day 2-4,, oestradiol and progesterone. 24 hour urinary cortisol (or dexamethasone suppression) should be checked if Cushings is a clinical possibility. NOTE: Commercial testosterone assays are not sensitive enough to reliably measure testosterone levels at or below the female lower reference limit (ie at or below the female lower normal limit) and should not be relied upon to diagnose androgen deficiency in women. PCOS These patients present with hyperandrogenism, irregular menstruation and infertility. They are commonly overweight or obese and have polycystic ovaries on ultrasound, however polycystic ovaries are not exclusive to PCOS. Insulin resistance, dyslipidemia, glucose intolerance and type 2 diabetes are strongly associated. Incidence is up to 12% and rising with increasing obesity; first degree relatives are at increased risk. Clinically PCOS commonly develops during puberty; hirsutism is the commonest finding followed by menstrual irregularity or amenorrhoea (20% of cases). Anovulation results in infertility. Ultrasound shows polycystic ovaries (>12 follicles per ovary) which reflects failure of developing follicles to ovulate. Polycystic ovaries may, however, also be found in normal women. Obesity is present in more than 50% of cases of PCOS and is typically centripetal (due to visceral fat) with increased waist/hip ratio and accompanying insulin resistance. In these cases high insulin levels correlate with low SHBG and thus free testosterone levels increase. Insulin resistance is also present in non-obese PCOS; the mechanism is uncertain. Patients with severe insulin resistance may develop acanthosis negricans. Glucose tolerance testing is advisable for all PCOS patients (fasting blood glucose may be normal but 2 hour specimen is often abnormal) as these patients are at high risk of diabetes (approximately a third develop type 2 diabetes by age 40 years). Patients are also at high risk of dyslipidaemia (low HDL, high LDL and triglycerides), metabolic syndrome and cardiovascular disease. Pathogenesis and Treatment of PCOS Increased androgens and insulin resistance (present in obese and non-obese PCOS) are probably both involved. Insulin resistance results in reduced SHBG and thus higher free testosterone levels. Ovarian androgen production is also increased due to high levels. Increased androgens cause android fat deposition, which worsens insulin resistance and dyslipidaemia. Treatment is by OCP (suppresses ovarian androgen production and increases SHBG levels, thus reducing free androgen levels) anti-androgens e.g. spironolactone, flutamide and finasteride, and insulin sensitization by weight loss, exercise and metformin. Hormone profile in PCOS Low SHBG, mild to moderate elevation of FAI/free testosterone and mild elevation of testosterone (usually less than 3 nmol/l) are the most reliable biochemical markers, especially in obese patients. (see pattern C) Lean patients with PCOS generally have elevated / ratios >2; these ratios may, however, be normal in obese patients and false positives are seen if testing is done near to midcycle. Testosterone levels > 6nmol/L and DHEAS >20umol/L are suggestive of tumour rather than PCOS production. Clinipath Pathology Newsletter / Summer 2011

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