Pattern Recognition Receptor-initiated Innate Immune Responses In The Mouse Epididymis

Background and Objectives:Epididymis can be infected by a broad spectrum of microbial pathogens, including bacteria, viruses, and parasites, which may cause epididymitis, an aetiological factor of male infertility. Uropathogenic E. coli (UPEC), Mumps virus (MuV), and Herpes simplex virus 2 (HSV-2) predominantly infect the epididymis and induce epididymitis. The mechanisms underlying the innate immune responses to these pathogens in the epididymis are not fully understood. This thesis investigated pattern recognition receptor (PRR)-initiated innate immune responses in epididymal epithelial cells (EECs).Materials and Methods:TLR2-/-、TLR4-/-、TLR5-/- mice and RNA interference (RNAi) approaches were used to investigate gene functions. Primary epididymal epithelial cells (EECs) were isolated and stimulated with UPEC, MuV, and HSV-2 for in vitro study. Gene expression at mRNA level was examined by real-time quantitative RT-PCR. Western blot and ELISA were used for analyzing protein levels. Immunohistochemistry was used to determine protein distribution. The innate immune responses in vivo were investigated after injection of UPEC, MuV, and HSV-2 into the epididymis.Results:Several PRRs are expressed in EEC. UPEC triggers innate immune responses through the activation of TLR4 and TLR5 in EEC. TLR2 and retinoic acid-inducible gene I (RIG-1) can be activated by MuV, HSV-2 activates DNA-dependent activator of interferon (DAI), p204, cyclic GMP-AMP synthase (cGAS). Both UPEC and MuV can induce the expression of proinflammatory cytokines, chemokines and type 1 interferons (IFN-a and IFN-β), while HSV-2 only induces the expression of type 1 interferons. UPEC-triggered epididymal innate immune responses are significantly reduced in TLR4--/-and TLR5-/- EEC, suggesting that TLR4 and TLR5 cooperatively initiate the innate immune responses to UPEC infection. MuV-triggered innate immune responses in EEC are remarkably reduced by the knockout of TLR2 and the knockdown of individual RIG-I, suggesting that MuV induces innate immune responses through TLR2 and RIG-I. HSV-2-induced innate antiviral responses in EEC were reduced by silencing p204, DAI, cGAS, which indicate that these DNA sensors initiate innate antiviral responses to HSV-2 stimulation.Conclusions:The data of this thesis have revealed the mechanisms by which UPEC, MuV, and HSV-2 induce innate immune responses in EEC, which indicate that EEC would be involved in the regulation of inflammatory responses in the epididymis and the epididymal defenses against these pathogens.