SOX9 : Immunohistochemical study of normal and neoplastic canine Sertoli cells

SOX9 protein plays a pivotal role for male sexual development. Among its numerous functions, it regulates the transcription of the anti-Müllerian hormone (AMH) gene and interacts with other genes to promote the development of testis cords, the multiplication and maturation of Sertoli cells (SCs) and the maintenance of spermatogenesis in the adult testis. The expression of SOX9 in normal testes has been demonstrated in humans, mice and rats. In canine species SOX9 expression in normal SCs has never been investigated and no data are available about neoplastic canine SCs (Sertoli Cell Tumours, SCTs). The present study aimed to investigate the expression of SOX9 in canine SCs during testicular maturation and neoplastic transformation. Testicular samples derived from 1 foetus, 4 newborns, 2 prepuberal puppies, 3 adult dogs, 28 SCTs (1 of them metastasizing) and 3 Leydig cell tumors (LCTs) were selected from the archive and tested immunohistochemically with a polyclonal antibody against SOX9 (1:150). Histologically, 18/28 SCTs were typical SCTs, 10/28 were classified as “lipid rich” SCTs, and 6/28, including the metastasizing one, characterized by capsular invasion, solid growth, severe anisocytosis and anisocytosis, large areas of necrosis and hemorrhages were considered undifferentiated SCTs. Immunohistochemically, all SCs from foetal, neonatal and adult testes had a strong, diffuse and exclusively nuclear labelling for SOX9. In prepuberal testes, SOX9 stained exclusively SCs nucleus in one puppy and both nucleus and cytoplasm in the other one. Leydig cells (LCs) were constantly negative in all samples. Similarly negative were all the LCTs. Concerning the 28 Sertoli cell tumors, 2 were negative for SOX9 and were reclassified as LCTs, while in the remaining 26 SCTs, SOX9 was diffusely expressed confirming the diagnosis of SCTs. In these tumours, the expression of SOX 9 was nuclear, nuclear and cytoplasmic or exclusively cytoplasmic in 14/26, 10/26 and 2/26 SCTs respectively. Moreover, all the undifferentiated SCTs and “lipid rich” cases were characterized by less intense staining. This is the first report on immuhistochemical SOX9 expression in canine testes and demonstrates that in normal SCs from foetal, neonatal and adult testes, SOX9 labelled the nucleus as in human species. On the other hand, the cytoplasmic labelling observed in one puppy might parallel with prepuberal rat testes staining. Considering this datum, the cytoplasmic expression of SOX9 described in numerous canine SCTs could reflect cellular immaturity/dedifferentiation. In addition, the expression of SOX9 in SCTs and its absence in LCTs suggests that SOX9 is a reliable marker in the differential diagnosis between canine SCTs and LCTs.