Avaliação da atividade biológica do peptídeo 3 semelhante à insulina (INSL3) nos testículos de zebrafish (Danio rerio) adultos

• Ano de defesa: 2015
• Autor(a) principal: Luiz Henrique de Castro Assis
• Tipo de documento: Tese
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Minas Gerais; UFMG
• Palavras-chave em Português: Biologia Celular; Citologia
• Link de acesso: http://hdl.handle.net/1843/BUBD-A2GGBL

Spermatogenesis is a cyclical and organized process that leads to the production of male gamete, the spermatozoa, from undifferentiated spermatogonia. Undifferentiated spermatogonia have the capacity to produce, by cell division, either more undifferentiated spermatogonia (self-renewal) or germ cells committed to the development of spermatogenesis (differentiation). Therefore, the balance between self-renewal and differentiation is crucial for the maintenance of spermatogenesis during the reproductive life. Nevertheless, the regulation of this balance in adult fish still needs to be elucidated, particularly the aspects involving paracrine regulation. INSL3 (insulin-like peptide 3) is a relaxin peptide family member produced by Leydig cells in vertebrate testes. In mammals, it is known that INSL3 has a crucial role on testicular descent during development and also on germ cell survival during adult life. In teleosts, although the testes remain inside the body cavity, the gene insl3 is highly expressed during development and also during the adult life, potentially indicating evolutionary older functions yet undiscovered. Previous observations showed that Amh (anti-müllerian hormone), in addition to inhibiting spermatogonial differentiation and androgen release, inhibited Fsh (follicle-stimulating hormone)-induced increase in insl3 transcript levels in zebrafish testis. Therefore, we investigated if the two growth factors might have antagonistic effects. Ex vivo studies indicated that hINSL3 increased the proliferation of type A undifferentiated (Aund) spermatogonia, while reducing proliferation of Sertoli cells associated with proliferating Aund. Moreover, ex vivo experiments using zebrafish testes with in vivo pre-labeled type Aund spermatogonia showed that hINSL3 decreased the number and area of testicular parenchyma occupied by cysts of type Aund spermatogonia, and increased the area occupied by cysts of type A differentiating (Adiff) spermatogonia. Corroborating the morphometric data, quantitative PCR analyses showed that hINSL3 decreased the transcription of nanos2 gene, a marker for type Aund spermatogonia in zebrafish and other vertebrates. Hence, we conclude that hINSL3 promoted differentiation of type Aund spermatogonia in zebrafish testes culture. However, our investigations did not indicate effects of hINSL3 on androgen release. Finally, the investigations using animals with androgens and gonadotropins deficiency, due to an in vivo exposure to 17-estradiol, showed that mRNA levels for insl3 gene were drastically reduced in this pro self-renewal condition. Additionally, the partial recovery of spermatogenesis observed after the introduction of 11-ketoandrostenedione on the in vivo treatment was not related to the recovery of insl3 transcription. In this context, our data corroborate the hypothesis that hINSL3 recruited Aund spermatogonia into differentiation without androgen participation, potentially mediating an Fsh effect on spermatogenesis.