Roles Of Autophagy And Pattern Recognition Receptors In Regulating The Innate Antiviral Responses In Mouse Sertoli Cells And Epididymal Epithelial Cells

Background and objectives:Various microbial pathogens, including bacteria, fungi, viruses, and parasites, may infect testis and epididymis, which impair male fertility. The tissue-specific cells adopt effective innate immune defense mechanisms against microbial infections. Toll-like receptor 3 (TLR3) can initiate innate immune responses in Sertoli cells. Autophagy can directly eliminate invading pathogens and regulate innate immune responses against microbal infections. This thesis focused on the innate immune responses and autophagy in mouse Sertoli cells and epididymal epithelial cells (EECs).Materials and methods:TLR3-/- mice and siRNA approaches were used to investigate gene function. Gene expression at mRNA level was examined using Real-time qRT-PCR. Immunohistochemistry and Western blot were used for the determination of protein distribution and quantification. The cytokine level was measured using ELISA. Poly(I:C) treatment was used to mimic viral infection. Rapamycin and Bafilomycin A1 were used to regulate autophagy.Results:LC3 and beclin-1, two key autophagy-related genes, were highly expressed in mouse Sertoli cells. IFN-β and antiviral proteins, including IFN-stimulated gene 15 (ISG15),2’-5’-oligoadenylate synthetase (OAS1), and myxovirus resistence protein 1 (MX1), were significantly upregulated in WT mouse Sertoli cells after poly(I:C) transfection. However, the expression of these genes was significantly reduced in TLR3-Sertoli cells. Poly(I:C)-induced innate antiviral responses in Sertoli cells depended on IFN regulatory factor-3 activation. Poly(I:C) promoted autophagy. Autophagy facilitated the innate antiviral responses. Moreover, we found that EECs express TLR3 and RIG-I, which initiate the epididymal antiviral responses to poly(I:C).Conclusions:TLR3 in Sertoli cells can be activated by poly(I:C) and then initiates innate antiviral responses. Autophagy enhanced the innate antiviral responses in Sertoli cells. TLR3 and RIG-I cooperatively initiate innate antiviral responses in EECs.