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      Anti Müllerian Hormone (AMH) level and expression in mural and cumulus cells in relation to age

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          Abstract

          Background

          Serum AMH is declining with age and is highly associated with ovarian follicular reserve and disordered folliculogenesis. However, the precise role of AMH in the process of human follicular aging has still to be determined.

          Aim

          This study investigates AMH level in the follicular fluid (FF) and mRNA expression pattern in cumulus and mural granulosa cells of human ovarian follicles in relation to age.

          Methods

          We conducted a prospective study. Sixty-eight women undergoing In vitro fertilization (IVF) treatment were enrolled in the study. We obtained FF, mural and cumulus granulosa cells from large preovulatory follicles (17-20 mm) of 21–35 years old women (n = 40) and 40–45 years old women (n = 28) during oocyte pickup.

          Results

          Higher level of AMH mRNA expression in cumulus cells was observed in the older age group compared to the younger (P <0.01). In accordance with AMH mRNA expression results, FF AMH protein levels were significantly higher in the older group than in the younger group (4.7 ± 1.1 ng\ml and 2.3 ± 0.2 ng\ml respectively, p < 0.002).

          Conclusions

          AMH is highly expressed and secreted from cumulus GCs of advanced age patients. This remarkable correlation between AMH mRNA levels in cumulus cells in respect to age suggests that AMH may be involved in follicular aging process.

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          Most cited references18

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          Anti-Müllerian hormone expression pattern in the human ovary: potential implications for initial and cyclic follicle recruitment.

          Anti-Müllerian hormone (AMH) is a member of the transforming growth factor-beta superfamily, which plays an important role in both ovarian primordial follicle recruitment and dominant follicle selection in mice. However, the role of AMH in folliculogenesis in humans has not been investigated in detail. In the present study, AMH expression was assessed using immunohistochemistry in ovarian sections, obtained from healthy regularly cycling women. To this end, a novel monoclonal antibody to human AMH was developed. AMH expression was not observed in primordial follicles, whereas 74% of the primary follicles showed at least a weak signal in the granulosa cells. The highest level of AMH expression was present in the granulosa cells of secondary, preantral and small antral follicles
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            Anti-Müllerian hormone inhibits initiation of primordial follicle growth in the mouse ovary.

            Recruitment of primordial follicles is essential for female fertility; however, the exact mechanisms regulating this process are largely unknown. Earlier studies using anti-Müllerian hormone (AMH)-deficient mice suggested that AMH is involved in the regulation of primordial follicle recruitment. We tested this hypothesis in a neonatal ovary culture system, in which ovaries from 2-d-old C57Bl/6J mice were cultured for 2 or 4 d in the absence or presence of AMH. Ovaries from 2-d-old mice contain multiple primordial follicles, some naked oocytes, and no follicles at later stages of development. We observed that in the cultured ovaries, either nontreated or AMH-treated, follicular development progressed to the same extent as in in vivo ovaries of comparable age, confirming the validity of our culture system. However, in the presence of AMH, cultured ovaries contained 40% fewer growing follicles compared with control ovaries. A similar reduction was found after 4 d of culture. Consistent with these findings, we noted lower inhibin alpha-subunit expression in AMH-treated ovaries compared with untreated ovaries. In contrast, expression of AMH ligand type II receptor and the expression of oocyte markers growth and differentiation factor 9 and zona pellucida protein 3 were not influenced by AMH. Based on the results, we suggest that AMH inhibits initiation of primordial follicle growth and therefore functions as an inhibitory growth factor in the ovary during these early stages of folliculogenesis.
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              Which follicles make the most anti-Mullerian hormone in humans? Evidence for an abrupt decline in AMH production at the time of follicle selection.

              Anti-Müllerian hormone (AMH) is exclusively produced by granulosa cells (GC) of the developing pre-antral and antral follicles, and AMH is increasingly used to assess ovarian function. It is unclear which size follicles make the most AMH (total content) and are the main contributors to circulating AMH concentrations. To determine AMH gene expression in GC (q-RT-PCR) and follicular AMH production (Elisa and RIA) in relation to follicular development, 87 follicles (3-13 mm diameter) including both GC and the corresponding follicular fluid (FF) were collected in connection with fertility preservation of human ovaries. Further, follicle number and diameter, graded in 1 mm increments, were determined by 3D ultrasound in 113 women in their natural menstrual cycle to determine follicle number and diameter in relation to circulating AMH levels. This study demonstrates for the first time a positive association between AMH gene expression in human and both total follicular fluid AMH (P < 0.02) and follicular fluid AMH concentration (P < 0.01). AMH gene expression and total AMH protein increased until a follicular diameter of 8 mm, after which a sharp decline occurred. In vivo modelling confirmed that 5-8 mm follicles make the greatest contribution to serum AMH, estimated for the first time in human to be 60% of the circulating concentration. Significant positive associations between gene expression of AMH and FSHR, AR and AMHR2 expression (P < 0.00001 for all three) and significant negative association between follicular fluid AMH concentration and CYP19a1 expression were found (P < 0.0001). Both AMH gene expression (P < 0.02) and follicular fluid concentration of AMH (P < 0.00001) correlated negatively with estradiol concentration.
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                Author and article information

                Contributors
                kedem2001@gmail.com
                yuval.yung@sheba.health.gov.il
                gil.yerushalmi@gmail.com
                jigalh@hotmail.com
                ettiemaman@gmail.com
                mirithan@gmail.com
                Rina.Hemi@sheba.health.gov.il
                Raoul.orvieto@sheba.health.gov.il
                jdor@netvision.net.il
                Ariel.Hourvitz@sheba.health.gov.il
                Journal
                J Ovarian Res
                J Ovarian Res
                Journal of Ovarian Research
                BioMed Central (London )
                1757-2215
                11 December 2014
                11 December 2014
                2014
                : 7
                : 1
                : 113
                Affiliations
                [ ]IVF unit, Department of Obstetrics and Gynecology, Sheba Medical Center, Tel Hashomer, affiliated with the Sackler Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel
                [ ]Institute of Endocrinology, Sheba Medical Center, Tel Hashomer, affiliated with the Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv, Israel
                Article
                113
                10.1186/s13048-014-0113-3
                4269874
                25500128
                08973819-623c-4180-ba8f-74d8de69dfc1
                © Kedem et al.; licensee BioMed Central Ltd. 2014

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 22 September 2014
                : 26 November 2014
                Categories
                Research
                Custom metadata
                © The Author(s) 2014

                Obstetrics & Gynecology
                Obstetrics & Gynecology

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