Up-Regulation of SOX9 in Sertoli Cells from Testiculopathic Patients Accounts for Increasing Anti-Mullerian Hormone Expression via Impaired Androgen Receptor Signaling

Kuo-Chung Lan,Yen-Ta Chen,Hong-Yo Kang,Yung-Chiao Chang,Hsin-Jung Lin,Ko-En Huang,Chawnshang Chang,Claude Beaudoin

doi:10.1371/journal.pone.0076303

Abstract

BackgroundTestosterone provokes Sertoli cell maturation and represses AMH production. In adult patients with Sertoli-cells-only syndrome (SCOS) and androgen insensitivity syndrome (AIS), high level of AMH expression is detected in Sertoli cells due to defect of androgen/AR signaling.ObjectiveWe postulated that up-regulation of SOX9 due to impairment of androgen/AR signaling in Sertoli cells might explain why high level of anti-Mullerian hormone (AMH) expression occur in these testiculopathic patients.MethodsBiological research of testicular specimens from men with azoospermia or mouse. The serum hormone levels were studied in 23 men with obstructive azoospermia, 33 men with SCOS azoospermia and 21 volunteers with normal seminograms during a period of 4 years.Immunohistochemical staining and reverse-transcription PCR were used to examine the relationships among AR, SOX9 and AMH expression in adult human and mouse testes. The ability of AR to repress the expression of SOX9 and AMH was evaluated in vitro in TM4 Sertoli cells and C3H10T1/2 cells.ResultsSCOS specimens showed up-regulation of SOX9 and AMH proteins but down-regulation of AR proteins in Sertoli cells. The mRNA levels of AR were significantly lower and the SOX9, AMH mRNA levels higher in all SCOS patients compared to controls (P< 0.05). The testosterone levels in the SCOS patients were within the normal range, but most were below the median of the controls. Furthermore, our in vitro cell line experiments demonstrated that androgen/AR signaling suppressed the gene and protein levels of AMH via repression of SOX9.ConclusionsOur data show that the functional androgen/AR signaling to repress SOX9 and AMH expression is essential for Sertoli cell maturation. Impairment of androgen/AR signaling promotes SOX9-mediated AMH production, accounts for impairments of Sertoli cells in SCOS azoospermic patients.

Highlights

Androgen and the androgen receptor (AR) have been shown to play critical roles in testis function [1,2,3]
Consistent with previous reports [2,27,31], specific AR immunostaining was detected in the nuclei of Sertoli, Leydig, and peritubular myoid cells of adult human testis with normal spermatogenesis, but not in the germ cells (Figure 1A)
SOX9 proteins were exclusively detected in human testicular Sertoli cells nuclei (Figure 1B, E and H) and anti-Mullerian hormone (AMH) proteins were detected in the cytoplasm of Sertoli cells; neither signal was detected in Leydig or peritubular myoid cells (Figure 1C, F and I)

Summary

Introduction

Androgen and the androgen receptor (AR) have been shown to play critical roles in testis function [1,2,3]. A lack of AR expression in Sertoli cells was found to account for the absence of AMH repression during early human testicular development [12]. In adult patients with Sertoli-cells-only syndrome (SCOS) and androgen insensitivity syndrome (AIS), high level of AMH expression is detected in Sertoli cells due to defect of androgen/AR signaling. Objective: We postulated that up-regulation of SOX9 due to impairment of androgen/AR signaling in Sertoli cells might explain why high level of anti-Mullerian hormone (AMH) expression occur in these testiculopathic patients. Conclusions: Our data show that the functional androgen/AR signaling to repress SOX9 and AMH expression is essential for Sertoli cell maturation. Impairment of androgen/AR signaling promotes SOX9-mediated AMH production, accounts for impairments of Sertoli cells in SCOS azoospermic patients

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