[Show abstract][Hide abstract] ABSTRACT: The mammalian testis is an immunoprivileged site where male germ cell antigens are immunologically tolerated under physiological conditions. However, some pathological conditions can disrupt the immunoprivileged status and induce autoimmune orchitis, an etiological factor of male infertility. Mechanisms underlying autoimmune orchitis induction are largely unknown. The present study investigated the roles of Toll-like receptor 2 (TLR2) and TLR4 in mediating the induction of experimental autoimmune orchitis (EAO) in mice after immunization with male germ cell antigens emulsified with complete Freund adjuvant. Wild-type (WT) mice developed severe EAO after three immunizations, which was characterized by leukocyte infiltration, autoantibody production, and impaired spermatogenesis. Tlr2 or Tlr4 deficient mice showed relatively low susceptibility to EAO induction compared with WT mice. Notably, Tlr2 and Tlr4 double knockout mice were almost completely protected from EAO induction. Moreover, we demonstrated that TLR2 was crucial in mediating autoantibody production in response to immunization. The results imply that TLR2 and TLR4 cooperatively mediate EAO induction.
Copyright 2015 by The Society for the Study of Reproduction.
Biology of Reproduction 01/2015; 92(3). DOI:10.1095/biolreprod.114.123901 · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The mammalian testis is an immunoprivileged organ where male germ cell autoantigens are immunologically ignored. Both systemic immune tolerance to autoantigens and local immunosuppressive milieu contribute to the testicular immune privilege. Testicular immunosuppression has been intensively studied, but information on systemic immune tolerance to autoantigens is lacking. In the present study, we aimed to determine the role of Axl and Mer receptor tyrosine kinases in maintaining the systemic tolerance to male germ cell antigens using the experimental autoimmune orchitis (EAO) model. Axl and Mer double-knockout (Axl(-/-)Mer(-/-)) mice developed evident EAO after a single immunization with germ cell homogenates emulsified with complete Freund's adjuvant. EAO was characterized by the accumulation of macrophages and T lymphocytes in the testis. Damage to the seminiferous epithelium was also observed. EAO induction was associated with pro-inflammatory cytokine upregulation in the testes, impaired permeability of the blood-testis barrier and generation of autoantibodies against germ cell antigens in Axl(-/-)Mer(-/-) mice. Immunization also induced mild EAO in Axl or Mer single-gene-knockout mice. By contrast, a single immunization failed to induce EAO in wild-type mice. The results indicate that Axl and Mer receptors cooperatively regulate the systemic immune tolerance to male germ cell antigens.Immunology and Cell Biology advance online publication, 18 November 2014; doi:10.1038/icb.2014.97.
[Show abstract][Hide abstract] ABSTRACT: Viruses can infect adipose tissues. However, innate antiviral responses in adipose cells and their effects on adipocyte function have not yet been intensively investigated. In this study, p204-initiated innate antiviral responses in mouse adipose cells were examined. Cytosolic DNA sensor p204 and its signaling adaptor stimulator of interferon (IFN) genes (STING) were constitutively expressed in primary preadipocytes. Synthetic herpes simplex viral DNA (HSV60), a p204 ligand, induced type I IFN expression by activating IFN regulatory factor 3. Major antiviral proteins, including IFN-stimulating gene 15, 2',5'-oligoadenylate synthetase and Mx GTPase 1, in preadipocytes were upregulated by HSV60. HSV60-triggered innate antiviral responses were significantly reduced by inhibition of p204 signaling with specific small interfering RNA targeting p204 or STING. HSV60 inhibited the differentiation of preadipocytes to mature adipocytes and enhanced the proliferation of adipose cells. Moreover, HSV60 induced innate antiviral responses in mature adipocytes and inhibited expressions of several adipokines, including leptin, adiponectin and resistin. These results indicated that p204 initiated innate antiviral responses in adipose cells, thereby modulating adipocyte function.Immunology and Cell Biology advance online publication, 7 October 2014; doi:10.1038/icb.2014.83.
[Show abstract][Hide abstract] ABSTRACT: The mammalian testis is an immunoprivileged organ where local tissue-specific cells acquire an effective innate immune function against invading microbial pathogens. The current study demonstrated that mouse Leydig cells had innate antiviral activities in response to viral DNA challenge through p204 activation. The DNA sensor p204 and its signaling adaptor stimulator of interferon (IFN) genes (STING) were constitutively expressed in Leydig cells. Synthetic herpes simplex virus DNA analog (HSV60), a p204 agonist, induced the expression of type-I IFNs and various antiviral proteins, including IFN-stimulating gene 15, 2'5'-oligoadenylate synthetase and Mx GTPase 1, in Leydig cells. The HSV60-induced innate antiviral response in Leydig cells was significantly reduced by inhibiting p204 signaling using specific siRNAs targeting p204 and Sting. The antiviral response did not affect steroidogenesis in Leydig cells. These results indicated a novel mechanism underlying the testicular innate antiviral response.
Biology of Reproduction 05/2014; 91(1). DOI:10.1095/biolreprod.114.119396 · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Toxoplasma gondii and uropathogenic Escherichia coli (UPEC) may infect the testis and impair testicular function. Mechanisms underlying testicular innate immune response to these two pathogens remain to be clarified. The present study examined the function of TLR11, which can be recognized by T. gondii-derived profilin and UPEC, in initiating innate immune response in male mouse germ cells in vitro. TLR11 is predominantly expressed in spermatids. Profilin and UPEC induced the expressions of different inflammatory cytokine profiles in the germ cells. In particular, profilin induced the expressions of macrophage chemotactic protein 1 (MCP-1), interleukin (IL)-12 and interferon (IFN)-γ through nuclear factor κB (NF-κB) activation. UPEC induced the expressions of MCP-1, IL-12, and IFN-γ, as well as tumor necrosis factor alpha (TNF-α), IL-6, and IFN-β, through the activation of NF-κB, IFN regulatory factor 3 and mitogen-activated protein kinases. Evidence showed that profilin induced the innate response in male germ cells through TLR11 signaling, and UPEC triggered the response through TLR11 and other TLR signaling pathways. We also provided evidence that local injection of profilin or UPEC induces the innate immune response in the germ cells. Data describe TLR11-mediated innate immune function of male germ cells in response to T. gondii profilin and UPEC stimulations in vitro. This system may play a role in testicular defense against T. gondii and UPEC infections in mice.
Biology of Reproduction 01/2014; 90(2). DOI:10.1095/biolreprod.113.114421 · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: The testis is an immune privileged organ where the tissue-specific cells have adopted effective innate immune functions against microbial pathogens. Toll-like receptors (TLRs) mediate innate immune response in the testis. The current study demonstrates that melanoma differentiation-associated protein 5 (MDA5) and retinoic acid-inducible gene I (RIG-I) initiate the testicular innate antiviral response. Both MDA5 and RIG-I are expressed in Leydig cells, and MDA5 is also expressed in spermatids. Polyinosinic-polycytidylic acid [poly(I:C)], a common agonist of MDA5 and RIG-I, significantly induces the expression of type I interferons (IFN-α/β) and antiviral proteins, including IFN-stimulated gene 15, 2'5'-oligoadenylate synthetase 1, and Mx GTPase 1, in primary TLR3-deficient (TLR3(-/-)) Leydig and germ cells. Moreover, major pro-inflammatory cytokines, including tumor necrosis factor alpha and interleukin 6, are significantly up-regulated by poly(I:C) in these testicular cells. The poly(I:C)-induced innate antiviral response in the testicular cells is significantly reduced by knockdown of individual MDA5 and RIG-I using specific small interfering RNA. We also provide evidence that local injection of poly(I:C) induces antiviral response in the testis of TLR3(-/-) mice. These data provide novel insights into the mechanisms underlying testicular antiviral response.
[Show abstract][Hide abstract] ABSTRACT: Tyro3, Axl and Mer (TAM) receptor tyrosine kinases triple knockout (TAM(-/-)) mice are male infertile due to impaired spermatogenesis. However, the mechanism by which TAM receptors regulate spermatogenesis remains unclear. In this study, we demonstrate that the testicular immune homeostasis was impaired in TAM(-/-) mice. As development after the onset of sexual maturity, germ cells were progressively degenerated. Macrophages and lymphocytes infiltrated into the testis as TAM(-/-) mice aged. Moreover, the integrity of blood-testis barrier was impaired, and the autoantibodies against germ cell antigens were produced. Major inflammatory cytokines, including tumor necrosis factor-α, interleukin-6 and monocyte chemotactic protein 1 were upregulated in the testis of TAM(-/-) mice, and predominantly located in Sertoli cells (SCs). In vitro assays showed that TAM(-/-) SCs secrete significantly high levels of inflammatory cytokines compared with wild-type SCs after coculture with apoptotic germ cells. These results suggest that TAM receptors are important in the maintenance of the immune homeostasis in the testis.Immunology and Cell Biology advance online publication, 21 May 2013; doi:10.1038/icb.2013.22.
[Show abstract][Hide abstract] ABSTRACT: Viral infections of the ovary can cause pathological conditions. However, innate antiviral responses in the ovary are poorly understood. In this study, we demonstrate that Toll-like receptor 3 (TLR3), retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA5) are constitutively expressed in the mouse ovary and predominantly located in granulosa cells. Polyinosinic-polycytidylic acid [poly(I:C)], a common agonist of TLR3, MDA5 and RIG-I, induced innate antiviral responses in ovarian granulosa cells. Poly(I:C) up-regulated pro-inflammatory cytokines, including TNF-α and IL-6, and type I interferons (IFN-α/β). Moreover, poly(I:C) induced the expression of antiviral proteins, including 2'-5'-oligoadenylate synthetase, Mx GTPase 1 and IFN-stimulating gene 15, in granulosa cells. In contrast, P450 aromatase expression was inhibited by poly(I:C). The poly(I:C)-induced antiviral responses in TLR3 knockout (TLR3(-/-)) ovarian granulosa cells were reduced, and completely abolished by blocking of MDA5/RIG-I signaling. Further, the poly(I:C)-induced cytokine expression in TLR3(-/-) cells was reduced by knockdown of MDA5 or RIG-I. Data suggest that TLR3, MDA5 and RIG-I cooperate in mediating innate antiviral responses in granulosa cells, which may contribute to the defense of the ovary against viral infections.
[Show abstract][Hide abstract] ABSTRACT: The testis is an immunoprivileged site where local cell-initiated innate immunity plays a crucial role in antimicrobial responses. Toll-like receptors (TLRs) mediate innate immune responses in testicular somatic cells. Although several TLRs are expressed in some stages of male germ cells, the potential role of TLRs in triggering antimicrobial responses in the germ cells has yet to be exclusively studied. The current study demonstrates that TLR3 is constitutively expressed in spermatogonia and spermatocytes and can be activated by a synthetic double-strained RNA analog, polyinosinic-polycytidylic acid. TLR3 activation in these male germ cells up-regulates the expression of proinflammatory cytokines, such as interleukin IL1B, IL6, and tumor necrosis factor alpha, through activation of nuclear factor kappa B; it also induces production of type 1 interferons (IFNA and IFNB) through the activation of IFN regulatory factor 3. In addition, TLR3 activation increases the production of two major antiviral proteins, namely, double-stranded RNA-activated protein kinase and MX1 protein, by germ cells. Data in this article describe an antiviral response of male germ cells through the activation of TLR3 in vitro.
Biology of Reproduction 01/2012; 86(4):106. DOI:10.1095/biolreprod.111.096719 · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Activation of Toll-like receptors (TLRs) triggers rapid inflammatory cytokine production in various cell types. The exogenous product of growth-arrest-specific gene 6 (Gas6) and Protein S (ProS) inhibit the TLR-triggered inflammatory responses through the activation of Tyro3, Axl and Mer (TAM) receptors. However, regulation of the Gas6/ProS-TAM system remains largely unknown. In the current study, mouse macrophages are shown to constitutively express Gas6 and ProS, which synergistically suppress the basal and TLR-triggered production of inflammatory cytokines, including those of tumour necrosis factor-α, interleukin-6 and interleukin-1β, by the macrophages in an autocrine manner. Notably, TLR signalling markedly decreases Gas6 and ProS expression in macrophages through the activation of the nuclear factor-κB. Further, the down-regulation of Gas6 and ProS by TLR signalling facilitates the TLR-mediated inflammatory cytokine production in mouse macrophages. These results describe a self-regulatory mechanism of TLR signalling through the suppression of Gas6 and ProS expression.