Updating the Function of Activin A in the Fetal Testis: A New Role in Steroidogenesis

Activin A, Hsd17b3, Hsd17b1, Steroidogenesis, Sertoli cells, testis development

Over the last few decades, an increase in the incidence of male reproductive disorders as diverse as hypospadias, cryptorchidism, testicular germ cell cancer, and low sperm counts has been observed in numerous countries (1). It has been hypothesized that all these disorders share risk factors, suggesting that they are interrelated through a testicular dysgenesis syndrome (TDS) that could be caused by genetic or environmental factors. During fetal life, androgen production is essential to the development of male characteristics, and deficiencies in testosterone occurring within a discrete developmental time window have been suggested to be one of the main causes of TDS (1).

Androgen synthesis is initiated with the conversion of cholesterol into androstenedione, which is converted in turn into testosterone. Leydig cells have long been thought to be solely responsible for testosterone synthesis. Whereas this is true for adult Leydig cells, pioneer studies by Shima et al have shown that, in mice, fetal Leydig cells do not express HSD17B3 and are unable to convert androstenedione to testosterone. The conversion of androstenedione to testosterone rather occurs within the Sertoli cells (2). Furthermore, it was recently shown that murine fetal Sertoli cells also express HSD17B1, and that its loss leads to an increase in the mRNA levels of steroidogenic enzymes expressed in fetal Leydig cells and to the compensatory upregulation of Hsd17b3 levels in Sertoli cells (although intratesticular testosterone levels are unaffected) (3). This suggests that HSD17B1 contributes to testosterone synthesis by the Sertoli cells in fetal testes. Interestingly, HSD17B1 expression was also detected in human Sertoli cells (3), suggesting that Sertoli cells might also contribute to testicular steroidogenesis in men. Despite the evidence supporting the importance of Sertoli cells for testosterone production in the fetal testes, no signaling molecules regulating the Sertoli-related step of steroidogenesis have been identified.

Activin A, a ligand belonging to the transforming growth factor beta (TGFB) superfamily, is a signaling protein formed by the dimerization of two βA subunits (a subunit shared with the Inhibin A and Activin AB heterodimers) encoded by Inhba. A decade ago, two genetic mouse models of Inhba inactivation were generated (4, 5). Despite differences between these two models (Inhba knockout [KO] (5) vs Inhba conditional KO in the testicular somatic cells (4)), both showed that Inhba modulates the proliferation of fetal Sertoli cells (4, 5). The observed phenotype was attributed to the loss of Activin A, as the inactivation of Inha (the gene encoding the other subunit of Inhibin A) leads to postnatal gonadal tumor formation but not to dysgenesis of the fetal testis cords (6), whereas inactivation of Inhbb (the gene encoding the other subunit of Activin AB) slightly affects the coelomic vessel diameter (7). Despite these advances, the mechanisms of Activin A action in the developing testis cords remain to be elucidated.

In this issue of Endocrinology, Whiley et al (8) revisited the Inhba KO model to identify potential Activin A target genes in the developing testes. To do this, they first employed microarray analyses comparing e15.5 whole testes from Inhba KO and wild-type (WT) littermates. This was followed by RNAseq analyses done on e13.5 and e15.5 FACS-sorted testicular somatic cells from Inhba KO × Oct4-GFP littermates. The combination of these two analyses with preexisting datasets revealed that Activin A mainly affected the somatic cells at e15.5. Activin A target genes were identified primarily in the developing Sertoli cells. DAVID analyses further revealed that the genes regulated by Activin A play key roles in the integrity of the cell membrane, lipid biosynthesis/metabolism, and steroid biosynthesis, whereas the expression of sex-determining genes was not regulated by Activin A. Whiley et al further validated the results of their high throughput assays by performing Fluidigm analyses for candidate transcripts on e12.5 to 15.5 testes. Finally, the authors confirmed that Activin A directly affected developing testes by evaluating the expression of candidate genes in cultured newborn mouse testes treated with an Activin inhibitor.

A novel finding obtained from the analyses of the in vivo datasets and subsequent in vitro validation performed by Whiley et al was the identification of Hsd17b1 and Hsd17b3 as target genes of Activin A. Indeed, the mRNA levels of both Hsd17b1 and Hsd17b3 were reduced following the loss of Activin A activity, suggesting that Activin A governs testosterone synthesis in fetal and neonatal Sertoli cells. The authors further validated this theory in testes from e17.5 embryos by measuring the intratesticular steroid levels by liquid chromatography-mass spectrometry. Interestingly, androstenedione levels increased significantly in the testes from the KO animals compared with WT littermates, while the testosterone/androstenedione ratio significantly decreased and testosterone levels tended to decrease in the testes from the KO animals. These results, combined with the fact that the mRNA levels of steroidogenic genes in the fetal Leydig cells were not modified in the Inhba KO, confirmed that Activin A only regulates steroidogenesis in the Sertoli cells. Furthermore, the inability of Sertoli cells to convert androstenedione into testosterone leads to the accumulation of androstenedione and its subsequent conversion in 11-ketoandrostenedione (11KA4) and 11-ketotestosterone (11KT), resulting in the modification of the steroid hormone profile in the testes of Inhba KO mice.

The study by Whiley et al illustrates the value of reexploring previously-characterized transgenic mouse models. By doing so with the Inhba KO mouse model, the authors were able to identify a large set of potential Activin A target genes in the developing Sertoli cells, as well as identifying a crucial role for Activin A in the regulation of steroidogenesis in the fetal testis. Though it is important to mention that Inhibin A or Activin AB might also contribute to the regulation of these genes, this represents an exciting first step towards a better understanding of androgen production within the Sertoli cells. Further exploration of the exact mechanisms whereby Activin A regulates Hsd17b1 and Hsd17b3 in the fetal Sertoli cells is clearly warranted.

Abbreviations

Abbreviations
 
• KO

  knockout

 
• TDS

  testicular dysgenesis syndrome

 
• WT

  wild-type

Acknowledgments

Financial Support: This work was supported by Discovery Grants RGPIN-2014-04358 (to AB) from the Natural Sciences and Engineering Research Council of Canada (NSERC).

Additional Information

Disclosure Summary: The authors have nothing to disclose.

Data Availability: Data sharing is not applicable to this article as no datasets were generated or analyzed during the current study.