抑制猪支持细胞外泌体对精原干细胞自我更新及分化的影响.

[Objectives] This study investigates the capability of exosome secretion by Sertoli cells and its influence on the self-renewal and differentiation status of porcine spermatogonial stem cells(pSSC) using an in vitro model. [Methods] The two-step enzyme digestion method was employed to isolate Sertoli cells and pSSC, and an in vitro co-culture model of Sertoli cells and pSSC was established. Suppression of exosome secretion by Sertoli cells was achieved to examine the effects on pSSC self-renewal and differentiation markers, thereby studying the impact of Sertoli cell-derived exosome on the fate of pSSC. [Results] The isolated Sertoli cells exhibited a Wilm tumor 1(WT1) positive rate of (87.1±0.02)% and an intorgrin β-1(ITGB1) positive rate of (94.2±0.01)%. The isolated pSSC showed a promyelocytic lenkaemia zine finger protein (PLZF) positive rate of (80.6±0.02) %. Presence of exosome secreted by Sertoli cells was confirmed through Western blot analysis and immunofluorescence assays. Treatment with 20 µmol L-1 GW4869 effectively inhibited exosome secretion without compromising Sertoli cells. Co-culture of pSSC with Sertoli cells treated with the inhibitor revealed significant differences in pSSC status compared to the control group. RT-PCR results showed a significant increase in the expression of the PLZF gene, while expression levels of receptor tyrosine kinase gene (KIT), thymus cell antigen 1 gene (THYI), and dead box 4 gene (DDX4) were significantly reduced. Western blot analysis indicated that levels of protein gene product 9.5(PGP9.5) protein expression elevated and DDX4 protein expression level decreased in pSSC co-cultured with inhibitor-treated Sertoli cells. [Conclusions] Exosomes secreted by Sertoli cells had a regulatory role in both self-renewal and differentiation of pSSC in vitro. This finding implied that exosome represented an important means through which Sertoli cells influence the fate of spermatogonial stem cells, offering novel insights for enhancing boar reproductive performance and fertility improvement. [ABSTRACT FROM AUTHOR]