Activin and follistatin interactions in the male reproductive tract: activin expression and morphological abnormalities in mice lacking follistatin 288.

Activin A is an important regulator of testicular and epididymal development and function, as well as inflammation and immunity. In the adult murine reproductive tract, activin A mRNA (Inhba) expression levels are highest in the caput epididymis and decrease progressively towards the distal vas deferens. The activin-binding protein, follistatin (FST), shows the opposite expression pattern, with exceptionally high levels of the Fst288 mRNA variant in the vas deferens. This unique pattern of expression suggests that activin A and follistatin, in particular FST288, play region-specific roles in regulating the epididymis and vas deferens. The cellular distribution of activin and follistatin and structural organization of the male reproductive tract was examined in wild-type and transgenic (TghFST315) mice lacking FST288. Compared to wild-type littermates, TghFST315 mice showed a 50% reduction in serum follistatin and a significant elevation of both activin A and B. Testicular, epididymal and seminal vesicle weights were reduced, but intra-testicular testosterone was normal. A decrease in the epididymal duct diameter in the corpus and thickening of the peritubular smooth muscle in the cauda, together with increased coiling of the proximal vas deferens, were observed in TghFST315 mice. No immune cell infiltrates were detected. Immunohistochemistry indicated that epithelial cells are the main source of activins and follistatin in the epididymis and vas deferens. Activin A, but not activin B, was also localized to sperm heads in the lumen of the epididymis and vas deferens. Expression of Inhba and another immunoregulatory gene, indoleamine-2,3-dioxygenase (Ido-1), was increased approximately twofold in the TghFST315 caput epididymis, but several other genes associated with immunoregulation, inflammation or fibrosis were unaffected. Our novel data indicate that disruption of follistatin expression has significant effects on the testis and epididymis, and suggest an association between activin A and indoleamine-2,3-dioxygenase in the caput epididymis, with implications for the epididymal immunoenvironment.

GGN1 in the testis and ovary and its variance within the Australian fertile and infertile male population.

Mouse gametogenetin (Ggn) is a testis-enriched gene that encodes multiple spliced transcripts giving rise to three predicted protein isoforms: GGN1, GGN2 and GGN3. Of these, GGN1 has been linked to germ cell development. Based on the spatial and temporal expression pattern of GGN1 during mouse spermatogenesis, it has been proposed as a candidate human infertility gene. Here, we report the localization of GGN1 in the human testis and ovary compared with the mouse orthologue. Within the testis, GGN1 was confined to pachytene spermatocytes and spermatids. During mid-prophase GGN1 redistributes from a solely cytoplasmic localization to both cytoplasmic and nuclear in late prophase spermatocytes and round spermatids, and is ultimately incorporated into the sperm tail. Within both mouse and human ovaries, GGN1 was localized within granulosa cells. Lower levels of expression were observed in mouse oocytes and the cumulus cells. Furthermore, to define the level of sequence variation in the fertile population and to assess the potential for an association with male infertility, we sequenced the coding region of human GGN in 100 idiopathic oligospermic infertile and 100 control men. Fifteen genetic variants were identified, of which 10 had not previously been reported. No significant associations with fertility status were observed, suggesting that variance in the GGN gene are not a common cause of oligospermic infertility in Australian men.

Expression of Wsb2 in the developing and adult mouse testis.

We present a detailed study of the expression pattern of WD repeat and SOCS box-containing 2 (Wsb2) in mouse embryonic and adult gonads. Wsb2 was previously identified in a differential screen aimed at identifying the genes involved in male- and female-specific gonadal development. Wsb2 expression was analysed during mouse gonadogenesis by real-time PCR, whole-mount and section in situ hybridisation and immunofluorescence. Wsb2 mRNA expression was initially detected in gonads of both sexes from 11.5 days post coitum (dpc) until 12.0 dpc. By 12.5 dpc and thereafter, Wsb2 expression rapidly decreased in the female, while persisting in the male gonads. In foetal, newborn and juvenile testes, Wsb2 mRNA and protein were readily detected in the seminiferous cords within both Sertoli and germ cells. Wsb2 mRNA was present in spermatogonia, spermatocytes and in Sertoli cells of the adult mouse testis. The differential expression of Wsb2 in male versus female embryonic gonads suggests some male-specific role in gonad development, and its expression in the first wave of spermatogenesis indicates a role in germ cells. Real-time analysis of adult mouse testis tubules cultured in the presence of the Hedgehog signalling inhibitor, cyclopamine, showed a downregulation of Wsb2 mRNA after treatment which suggests that Wsb2 may be a target of Hedgehog signalling.

Novel scavenger receptor gene is differentially expressed in the embryonic and adult mouse testis.

In an effort to understand the mechanisms that underpin gonadal differentiation at the time of sex determination, we identified a cDNA encoding a putative novel testis expressed scavenger receptor, Tesr. Based on its domain structure, we hypothesize that the function of Tesr is similar to that of other scavenger receptors that play roles in phagocytosis of apoptotic cells, cell-cell adhesion, and defense. Tesr mRNA was detected in fetal mouse gonads of both sexes at 11.5 days post coitum (dpc). From 12.0 dpc, Tesr expression rapidly decreased in the female and was maintained in the male. Expression was seen in embryonic mouse sites other than the testis, such as in brain, eye, head, heart, neural arch, and cartilage primordium. Tesr expression in the newborn testis was faint to undetectable, but it increased from 2 days postpartum (dpp) until 15 dpp and was found in a subset of interstitial cells and in germ and Sertoli cells. Tesr mRNA in the adult mouse testis was observed in Sertoli cells, spermatogonia, spermatocytes, round spermatids, and in a subset of interstitial cells. We conclude that Tesr is differentially expressed in the male vs. female embryonic gonad and is expressed in both the ovary and the testes postnatally after 2 dpp.

Subtractive hybridisation screen identifies sexually dimorphic gene expression in the embryonic mouse gonad.

The sex of most mammals is determined by the action of SRY. Its presence initiates testis formation resulting in male differentiation, its absence results in ovary formation and female differentiation. We have used suppression subtraction hybridisation between 12.0-12.5 days postcoitum (dpc) mouse testes and ovaries to identify genes that potentially lie within the Sry pathway. Normalised urogenital ridge libraries comprising 8,352 clones were differentially screened with subtracted probes. A total of 272 candidate cDNAs were tested for qualitative differential expression and localisation by whole mount in situ hybridisation; germ cell-dependent or -independent expression was further resolved using busulfan. Fifty-four genes were identified that showed higher expression in the testis than the ovary. One novel gene may be a candidate for interactions with WT1, based on its localisation to Sertoli cells and map position (16q24.3).