Yih Harng Chong

Serum anti-Mullerian hormone (AMH) levels correlate with infrarenal aortic diameter in healthy older men: is AMH a cardiovascular hormone?

Anti-Müllerian hormone (AMH) is a gonadal hormone present in the blood in men and pre-menopausal women. AMH regulates male sexual differentiation but has no putative function in adulthood. In recent studies, high AMH levels are associated with absence of cardiovascular disease in men and smaller atherosclerotic burden in monkeys. Mechanistically, AMH has downstream convergence with known regulators of the cardiovascular system, while the specific receptor for AMH is present in murine aorta and the human heart. Our primary objective was to examine whether AMH levels in healthy men correlated with the physical characteristics of their aorta. Our secondary aim was to document whether men with distinct vascular disorders expressed different levels of AMH. Serum AMH assayed by ELISA in 153 men (54-93 years) free from vascular disease inversely correlated with the ultrasonographic diameters of the distal- (r=-0.22, P=0.006) and mid-infrarenal aorta (r=-0.26, P=0.008). This association was similar in magnitude but opposite to that of body surface area (largest known determinant of aortic diameter) and independent of known cardiovascular risk factors. This relationship is specific to AMH, as inhibin B, a Sertoli cell hormone-like AMH, did not correlate with aortic diameter (r=-0.04, P=0.66) despite partially correlating with AMH. Among men with known vascular disease, higher AMH levels were associated with varicose vein disease, while men with higher levels of AMH were under-represented in the abdominal aortic aneurysm relative to the healthy cohort. These findings identify AMH as a novel putative regulator of the cardiovascular system.

Enzyme-linked immunosorbent assay measurements of antimüllerian hormone (AMH) in human blood are a composite of the uncleaved and bioactive cleaved forms of AMH

OBJECTIVE To determine whether the Beckman Coulter antimüllerian hormone (AMH) Gen II enzyme-linked immunosorbent assay (ELISA) detects the uncleaved precursor (proAMH) and/or the active cleaved form (AMHN,C) of AMH. DESIGN Technical investigation. SETTING Community study. PATIENT(S) Healthy boys and male and female adult volunteers. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Assay of AMH and Western blot analysis of captured forms of AMH. RESULT(S) In blood, AMH in blood consists of both proAMH, the inactive uncleaved precursor, and AMHN,C, the enzyme-cleaved, receptor-competent form. The Gen II AMH ELISA detected both recombinant proAMH and AMHN,C. The noncovalent association of the two cleavage fragments of AMHN,C appears to be necessary for ELISA detection because recombinant free AMHC and AMHN were undetectable. Spike-recovery experiments showed that proAMH was not completely recovered from serum unless it was prediluted 1 hour before the assay. CONCLUSION(S) The leading ELISA for AMH provides a composite value of two biologically distinct forms of AMH. It is not known whether proAMH and AMHN,C have identical relationships to ovarian reserve, antral follicle counts, or other aspects of ovarian function. Hence, future research into the physiology and clinical utility of AMH should consider the two forms separately.

Elderly Men Have Low Levels of Anti-Müllerian Hormone and Inhibin B, but with High Interpersonal Variation: A Cross-Sectional Study of the Sertoli Cell Hormones in 615 Community-Dwelling Men

The Sertoli cells of the testes secrete anti-Müllerian hormone (Müllerian inhibiting Substance, AMH) and inhibin B (InhB). AMH triggers the degeneration of the uterine precursor in male embryos, whereas InhB is part of the gonadal-pituitary axis for the regulation of sperm production in adults. However, both hormones are also putative regulators of homeostasis, and age-related changes in these hormones may therefore be important to the health status of elderly men. The levels of AMH in elderly men are unknown, with limited information being available about age-related changes in InhB. We have therefore used ELISAs to measure Sertoli cell hormone levels in 3 cohorts of community-dwelling men in New Zealand. In total, 615 men were examined, 493 of which were aged 65 or older. Serum AMH and InhB levels inversely correlated with age in men older than 50 years (p<0.001) but not in the younger men. A minority of elderly men had undetectable levels of AMH and InhB. The variation in hormone levels between similarly aged men increased with the age of men. AMH and InhB partially correlated with each other as expected (r = 0.48, p<0.001). However, the ratio of the two Sertoli hormones varied significantly between men, with this variation increasing with age. Elderly men selected for the absence of cardiovascular disease had AMH levels similar to those of young men whereas their InhB levels did not differ from aged-matched controls. These data suggests that Sertoli cell number and function changes with age, but with the extent and nature of the changes varying between men.

The Daily Profiles of Circulating AMH and INSL3 in Men are Distinct from the Other Testicular Hormones, Inhibin B and Testosterone.

The testes secrete four hormones (anti-Müllerian hormone, insulin-like peptide 3, Inhibin B and testosterone) from two endocrine cell types. It is unknown whether anti-Müllerian hormone and insulin-like peptide 3 levels have a diurnal variation, and if so, whether they covary during the day with testosterone and InhB. Sera were obtained from 13 men at 00:00, 06:00, 09:00, 12:00, 14:00, 17:00 and 19:00 hours and the levels of their testicular hormones measured by ELISA. A second cohort of 20 men was similarly examined with blood drawn at 19:00 and the following 06:00. Anti-Müllerian hormone levels exhibited a subtle diurnal pattern with a 19:00 peak that was 4.9% higher on average than the 06:00 nadir (p = 0.004). The decrease in anti-Müllerian hormone coincided with a rise in testosterone and InhB, but there was no association between the person-to-person variation in the diurnal patterns of anti-Müllerian hormone and testosterone or Inhibin B. Insulin-like peptide 3 had no diurnal pattern, with only minor sporadic variation between time points being observed in some men. In conclusion, the diurnal and sporadic variation of each testicular hormone is distinct, indicating that the major regulation is at the level of the hormone rather than at the endocrine cell type. Consequently, the balance of the hormones being released by the testes has complex variation during the day. The physiological significance of this will vary depending on which combinations of testicular hormones that the target cells respond to.

Anti-Müllerian hormone level in older women: detection of granulosa cell tumor recurrence.

Objective To determine whether anti-Müllerian hormone (AMH) production resumes during normal late menopausal aging. Anti-Müllerian hormone has been proposed as a specific serum marker for adult granulosa cell tumors. Materials and Methods Serum AMH from 21 elderly postmenopausal women (mean age, 77 years) and 9 young women (mean age, 22 years) were measured by ultrasensitive immunoassay. Results Both median (0 pmol/L) and mean (0.48 pmol/L) serum AMH values for the elderly women were below the level of detection for the immunoassay kit. Three of the 21 participants had minimally detectable level of AMH (1.13–2.76 pmol/L). The cohort of young women had expected normal values of AMH as measurable by the same immunoassay kit. Conclusions Serum AMH values were negligible for postmenopausal women older than 65 years. This extends the normative data for AMH to 108 years old, providing a reference range for the detection of granulosa cell tumors in postmenopausal women.

Relative levels of the proprotein and cleavage‐activated form of circulating human anti‐Müllerian hormone are sexually dimorphic and variable during the life cycle

Anti‐Müllerian hormone (AMH) is a gonadal hormone, which induces aspects of the male phenotype, and influences ovarian follicular recruitment. AMH is synthesized as a proprotein (proAMH), which is incompletely cleaved to the receptor‐competent AMHN,C. AMH ELISAs have not distinguished between proAMH and AMHN,C; consequently, the physiological ranges of circulating proAMH and AMHN,C are unknown. A novel proAMH ELISA has been used to assay serum proAMH in humans. Total AMH was also measured, enabling the AMHN,C concentration to be calculated. Stored serum from 131 boys, 80 younger, and 106 older men were examined, with serum from 14 girls and 18 women included for comparison. The mean levels of proAMH and AMHN,C in pM were respectively: boys (253, 526), men (7.7, 36), elderly men (5.7, 19), girls (3.3, 15), and women (5.2, 27) (boys vs. men, P < 0.001; girls vs. women, P = 0.032). The proportion of proAMH as a percentage of total AMH (API) was approximately twofold higher in boys than men (P < 0.001) with little overlap between the ranges, with girls also exhibiting lesser cleavage of their AMH than women (P < 0.001). The API varied within each population group. In young men, the API did not correlate with circulating levels of the other testicular hormones (testosterone, InhB, and INSL3). In conclusion, the cleavage of circulating AMH varies extensively within the human population, with most individuals having significant levels of proAMH. The physiological and clinical relevance of circulating proAMH needs to be established.

The Testicular Hormones AMH, InhB, INSL3, and Testosterone Can Be Independently Deficient in Older Men

Background Late-onset hypogonadism is symptomatically diverse and not fully explained by circulating testosterone level. The adult testes secrete four distinct hormones (testosterone, AMH, INSL3, and InhB) into the circulation. Testosterone and InhB have proven dynamic regulation, with limited information available for AMH and INSL3. During aging, there is cellular senescence, which may underlie the diversity of hypogonadism. This leads to the postulate that the relative levels (profile) of the four testicular hormones in older men are variable and cannot be evaluated by the measurement of one hormone. Methods 111 men aged 19-50 years and 98 men aged 70-90 years were examined. The circulating levels of the testicular hormones were measured using ELISAs, and the variation in the levels of hormones was analyzed by various correlative analyses. Results All four hormones were largely or totally independent. Some men were deficient in multiple hormones, but no man had multiple elevated hormones. The average hormonal levels were lower in older men, with diverse profiles of the four testicular hormones. Hence, some men had one or more hormones below the reference range, with testosterone the most conserved. Consequently, testosterone levels were not indicative of the complete state of the endocrine testes. Conclusions The four hormones vary independently of each other, in younger and older men. This indicates that they are regulated dynamically rather than influenced by endocrine cell number. Older men exhibited diverse profiles of low levels of testicular hormones, suggesting that the testes age differently between men. Testosterone alone inadequately describes gonadal states.

Does testicular anti-Müllerian hormone extend life?

Men tend to have shorter lives than women, with the gonads appearing to contributing to this. Gonadal hormones decline from midlife in men, but few elderly men become totally devoid of testicular hormones, the male equivalent of menopause [1]. The systemic metabolic demands created by gonadal hormones are therefore more extended in men than women. When the testicular influence is lessened by castration at a young age, the resulting eunuchs typically outlive intact men by over a decade [2]. This supports the theory that the testes facilitate reproductive success, at the expensive of longevity. Paradoxically, Qayyum and Akbar report in this edition of Endocrine that circulating anti-Müllerian hormone (AMH) in men negatively associates with all cause mortality [3]. Men with higher levels of a gonadal hormone may be outliving men with lower levels of the hormone. This is a novel observation, which opens new lines of investigation. Testosterone is synonymous to the testes for many people, with few being aware that the testes release multiple protein hormones, including AMH, inhibin B and insulin like-3 (INSL3). The levels of these testicular hormones are largely independent in young men, as is the time course of their decline during ageing [1]. Hence, the levels of each testicular hormone are determined by a unique set of regulatory determinants, rather than by a single pan-testicular mechanism. This points to each of the four hormones having distinct functions. Qayyum and Akbar’s study [3] raises the possibility that AMH may partially counterbalances the life-shortening effects of other aspects of the testes. Young men and women initially have similar levels of circulating levels of AMH, but young women lose their AMH in parallel with the decrease in their ovarian reserve. This adds to the paradox: AMH is the first of the ovarian hormones to decline, yet it associates with longevity in men, despite longevity having a female bias. AMH has two distinct faces (reviewed in [4]), and correlative studies do not establish which (if either) of the faces is important. At the start of life, AMH is a male-specific hormone, whose hallmark function is to induce the regression of the Müllerian duct, the uterine anlagen. The immature testes, however, continues to secrete very high levels of AMH into blood until puberty. Consequently, male embryos and boys develop with levels of AMH that do not occur during normal adult physiology or in developing females. The target tissues of AMH during development appear to be broad, but direct evidence of AMH actions is limited to the lungs and the brain. The correlates reported by Qayyum and Akbar [3] may be a reflection of this developmental mechanism, in which case they would be the product of male-specific biology. Men and women have low levels of circulating AMH compared to boys, but the adult levels are capable of signaling in some bioassays. If circulating AMH in adults maintains aspects of their homeostasis, as some animal studies suggest [5], then Qayyum and Akbar’s [3] results are possibly important to both sexes. At the very least, this highlights the potential risks associated with administering the sex steroid hormones alone, without understanding whether the other gonadal hormones modulate or counterbalance the actions of the gonadal steroids. * Ian S. McLennan ian.mclennan@otago.ac.nz

Testosterone in advance age: a New Zealand longitudinal cohort study: Life and Living in Advanced Age (Te Puāwaitanga o Ngā Tapuwae Kia Ora Tonu)

Objectives Serum testosterone (T) levels in men decline with age. Low T levels are associated with sarcopenia and frailty in men aged >80 years. T levels have not previously been directly associated with disability in older men. We explored associations between T levels, frailty and disability in a cohort of octogenarian men. Setting Data from all men from Life and Living in Advanced Age Cohort Study in New Zealand, a longitudinal cohort study in community-dwelling older adults. Participants Community-dwelling (>80 years) adult men excluding those receiving T treatment or with prostatic carcinoma. Outcomes measures Associations between baseline total testosterone (TT) and calculated free testosterone (fT), frailty (Fried scale) and disability (Nottingham Extended Activities of Daily Living scale (NEADL)) (baseline and 24 months) were examined using multivariate regression and Wald’s χ2 techniques. Subjects with the lowest quartile of baseline TT and fT values were compared with those in the upper three quartiles. Results Participants: 243 men, mean (SD) age 83.7 (2.0) years. Mean (SD) TT=17.6 (6.8) nmol/L and fT=225.3 (85.4) pmol/L. On multivariate analyses, lower TT levels were associated with frailty: β=0.41, p=0.017, coefficient of determination (R2)=0.10 and disability (NEADL) (β=−1.27, p=0.017, R2=0.11), low haemoglobin (β=−7.38, p=0.0016, R2=0.05), high fasting glucose (β=0.38, p=0.038, R2=0.04) and high C reactive protein (CRP) (β=3.57, p=0.01, R2=0.06). Low fT levels were associated with frailty (β=0.39, p=0.024, R2=0.09) but not baseline NEADL (β=−1.29, p=0.09, R2=0.09). Low fT was associated with low haemoglobin (β=−7.83, p=0.0008, R2=0.05) and high CRP (β=2.86, p=0.04, R2=0.05). Relationships between baseline TT and fT, and 24-month outcomes of disability and frailty were not significant. Conclusions In men over 80 years, we confirm an association between T levels and baseline frailty scores. The new finding of association between T levels and disability is potentially relevant to debates on T supplementation in older men, though, as associations were not present at 24 months, further work is needed.