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Three major pathogenic states of the prostate, including benign prostatic hyperplasia, prostate cancer, and prostatitis, are related to the local inflammation. However, the mechanisms underlying the initiation of prostate inflammation remain largely unknown. Given that the innate immune responses of the tissue-specific cells to microbial infection or autoantigens contribute to local inflammation, this study focused on pattern recognition receptor (PRR)-initiated innate immune responses in mouse prostatic epithelial cells (PECs). Primary mouse PECs abundantly expressed Toll-like receptor 3 (TLR3), TLR4, TLR5, melanoma differentiation-associated protein 5 (MDA5), and IFN-inducible protein 16 (p204 in mouse). These PRRs can be activated by their respective ligands: lipopolysaccharide (LPS) and flagellin of Gram-negative bacteria for TLR4 and TLR5, polyinosinicpolycytidylic acid (poly(I:C)) for TLR3 and MDA5, and herpes simplex virus DNA analog (HSV60) for p204. LPS and flagellin predominantly induced the expression of inflammatory cytokines, including tumor necrosis factor alpha (TNFA), interleukin 6 (IL6), chemokines monocyte chemoattractant protein-1 (MCP1), and C-X-C motif chemokine 10 (CXCL10). Poly(I:C) and HSV60 predominantly induced the expression of type 1 interferons (IFNA and IFNB) and antiviral proteins: Mx GTPase 1, 2′,5′-oligoadenylate synthetase 1, and IFN-stimulated gene 15. The replication of mumps virus in PECs was inhibited by type 1 IFN signaling. These findings provide insights into the mechanisms underlying innate immune response in the prostate.Summary sentenceToll-like receptors 4, 5 and nucleic acid sensors initiate innate immune responses in prostatic epithelial cells after challenge with their respective ligands, which may be associated with inflammation in the prostate.

Mammalian spermatogenesis is a carefully orchestrated male germ cell differentiation process by which spermatogonia differentiate to spermatozoa in the testis. A highly organized testicular microenvironment is therefore necessary to support spermatogenesis. Regarding immunologic aspects, the testis adapts a specialized immune environment for the protection of male germ cells and testicular functions. The mammalian testis possesses two immunologic features: (1) it is an immunoprivileged organ where immunogenic germ cells do not induce deleterious immune responses under physiologic conditions; and (2) it creates its own effective innate defense system against microbial infection. Various pathologic conditions may disrupt testicular immune homeostasis, thereby resulting in a detrimental immune response and perturbing testicular functions, one of the etiologic factors of male infertility. Understanding the mechanisms underlying immunoregulation in the testis can aid in establishing strategies for the prevention and therapy of immunologic testicular dysfunction and male infertility. This chapter focuses on the mechanisms underlying immune privilege, local innate immunity, and immunologic diseases of the testis.

The mammalian testis requires a highly organized microenvironment for the execution of its main functions, sperm production and testosterone synthesis. The testis possesses an immunoprivileged status essential for the protection of immunogenic male germ cells from detrimental immune responses. To counteract microbial infections, the tissue-specific cells are well equipped with innate defense machineries, termed pattern recognition receptors (PRRs), that can initiate innate immune responses. However, PRR-initiated innate immune responses in testicular cells may disrupt the immunoprivileged status and result in orchitis, thereby impairing spermatogenesis and testosterone synthesis. The mechanisms underlying the innate defense system and immunological pathophysiology in the testis have been intensively studied. Testicular immune homeostasis and disorders are regulated by a coordination of the tissue structure and cellular functions. This article focuses on the pivotal roles that Sertoli cells play in regulating testicular immune homeostasis by facilitating the blood-testis barrier, phagocytosis, paracrine, and innate immune responses.

The human testis can be infected by a large number of RNA and DNA viruses. While various RNA virus infections may induce orchitis and impair testicular functions, DNA virus infection rarely affects the testis. Mechanisms underlying the differential effects of RNA and DNA viral infections on the testis remain unclear. In the current study, we therefore examined the effects of viral RNA and DNA sensor signaling pathways on mouse Sertoli cells (SC) and Leydig cells (LC). The local injection of viral RNA analogue polyinosinic-polycytidylic acid [poly(I:C)] into the testis markedly disrupted spermatogenesis, whereas the injection of the herpes simplex virus (HSV) DNA analogue HSV60 did not affect spermatogenesis. Poly(I:C) dramatically induced the expression of the proinflammatory cytokines tumor necrosis factor α (TNF-α) and interleukin 6 in SC and LC through Toll-like receptor 3 and interferon β promoter stimulator 1 signaling pathways, impairing the integrity of the blood-testis barrier and testosterone synthesis. Poly(I:C)-induced TNF-α production thus plays a critical role in the impairment of cell functions. In contrast, HSV60 predominantly induced the expression of type 1 interferons and antiviral proteins via the DNA sensor signaling pathway, which did not affect testicular cell functions. Accordingly, the Zika virus induced high levels of TNF-α in SC and LC and impaired their respective cellular functions, whereas Herpes simplex virus type 2 principally induced antiviral responses and did not impair such functions. These results provide insights into the mechanisms by which RNA viral infections impair testicular functions.

Numerous types of viruses have been found in human semen, which raises concerns about the sexual transmission of these viruses. The overall effect of semen on viral infection and transmission have yet to be fully investigated. In the present study, we aimed at the effect of seminal plasma (SP) on viral infection by focusing on the mumps viral (MuV) infection of HeLa cells. MuV efficiently infected HeLa cells in vitro. MuV infection was strongly inhibited by the pre-treatment of viruses with SP. SP inhibited MuV infection through the impairment of the virus’s attachment to cells. The antiviral activity of SP was resistant to the treatment of SP with boiling water, Proteinase K, RNase A, and DNase I, suggesting that the antiviral factor would not be proteins and nucleic acids. PNGase or PLA2 treatments did not abrogate the antiviral effect of SP against MuV. Further, we showed that the prostatic fluid (PF) showed similar inhibition as SP, whereas the epididymal fluid and seminal vesicle extract did not inhibit MuV infection. Both SP and PF also inhibited MuV infection of other cell types, including another human cervical carcinoma cell line C33a, mouse primary epididymal epithelial cells, and Sertoli cell line 15P1. Moreover, this inhibitory effect was not specific to MuV, as the herpes simplex virus 1, dengue virus 2, and adenovirus 5 infections were also inhibited by SP and PF. Our findings suggest that SP contains a prostate-derived pan-antiviral factor that may limit the sexual transmission of various viruses.

Mammalian spermatogenesis is a carefully orchestrated male germ cell differentiation process by which spermatogonia differentiate to spermatozoa in the testis. A highly organized testicular microenvironment is therefore necessary to support spermatogenesis. Regarding immunologic aspects, the testis adapts a specialized immune environment for the protection of male germ cells and testicular functions. The mammalian testis possesses two immunologic features: (1) it is an immunoprivileged organ where immunogenic germ cells do not induce deleterious immune responses under physiologic conditions; and (2) it creates its own effective innate defense system against microbial infection. Various pathologic conditions may disrupt testicular immune homeostasis, thereby resulting in a detrimental immune response and perturbing testicular functions, one of the etiologic factors of male infertility. Understanding the mechanisms underlying immunoregulation in the testis can aid in establishing strategies for the prevention and therapy of immunologic testicular dysfunction and male infertility. This chapter focuses on the mechanisms underlying immune privilege, local innate immunity, and immunologic diseases of the testis.

The mumps virus (MuV) causes epidemic parotitis. MuV also frequently infects the testis and induces orchitis, an important etiological factor contributing to male infertility. However, mechanisms underlying MuV infection of the testis remain unknown. Here, we describe that sialic acid, AXL, and MER receptor tyrosine kinases regulate MuV entry and replication in mouse major testicular cells, including Sertoli and Leydig cells. Sialic acid, AXL, and MER were present in Sertoli and Leydig cells. Sialic acid specifically mediated MuV entry into Sertoli and Leydig cells, whereas both AXL and MER facilitated MuV replication within cells through the inhibition of cellular innate antiviral responses. Mechanistically, the inhibition of type 1 interferon signaling by AXL and MER is essential for MuV replication in Sertoli and Leydig cells. Our findings provide novel insights into the mechanisms behind MuV infection and replication in the testis.

Systemic inflammation may impair male fertility, and its underlying mechanisms remain poorly understood. The present study investigates the effect of lipopolysaccharide (LPS)-induced systemic inflammation on the testis and epididymis in mice. Intraperitoneal injection of LPS significantly impaired testicular functions, including testosterone production, spermatogenesis, and blood-testis barrier permeability. The epididymitis characterized by leukocyte infiltration and fibrosis was observed in the cauda epididymis after LPS injection. LPS-induced testicular dysfunction and epididymitis were abolished in tumor necrosis factor alpha (Tnfa) knockout mice. Pomalidomide, a TNFA inhibitor, blocked the detrimental effects of LPS on the testis and epididymis. The results indicate that LPS-induced systemic inflammation impairs male fertility through TNFA production, suggesting that the intervention on TNFA production would be considered for the prevention and treatment of inflammatory impairment of male fertility.

Viral infections of the ovary may perturb ovarian functions. However, the mechanisms underlying innate immune responses in the ovary are poorly understood. The present study demonstrates that cytosolic viral DNA sensor signaling initiates the innate immune response in mouse ovarian granulosa cells and affects endocrine function. The cytosolic DNA sensors p204 and cGAS and their common signaling adaptor stimulator of interferon (IFN) genes (STING) were constitutively expressed in granulosa cells. Transfection with VACV70, a synthetic vaccinia virus (VACV) DNA analog, induced the expression of type I interferons (IFNA/B) and major inflammatory cytokines (TNFA and IL6) through IRF3 and NF-κB activation respectively. Moreover, several IFN-inducible antiviral proteins, including 2′,5′-oligoadenylate synthetase, IFN-stimulating gene 15 and Mx GTPase 1, were also induced by VACV70 transfection. The innate immune responses in granulosa cells were significantly reduced by the transfection of specific small-interfering RNAs targeting p204, cGas or Sting. Notably, the VACV70-triggered innate immune responses affected steroidogenesis in vivo and in vitro. The data presented in this study describe the mechanism underlying ovarian immune responses to viral infection.

Microbial infection and immunological disorder in the male genital tract are among the etiological factors of male infertility. The testis and epididymis possess special immune environments for protecting the sperm from detrimental immune responses and counteracting invading microbial pathogens. However, specific pathological conditions, such as infection, chemical toxins, and tissue injury, may lead to infectious or autoimmune orchitis and epididymitis, resulting in male infertility. Moreover, infectious prostatitis, seminal vesiculitis, and urethritis may impair sperm quality and male fertility. Understanding the mechanisms underlying infectious and immunological infertility can aid in the development of novel preventive and therapeutic strategies. This review describes the mechanisms underlying male infertility caused by infectious and immunological diseases in the male genital tract.

To protect the male germ cells from adverse immune reaction, the male reproductive system adopts special immune environment such as immunoprivileged status. The male genital organs can be infected by various microorganisms via hematogenous dissemination and ascending genitourinary tracts. To overcome the immunoprivileged status, the male genital organs also adopt their own innate defense against microbial infection. The tissue-specific cells in the male reproductive system are well equipped with innate immune machineries, including pattern recognition receptors (PRRs) and their negatively regulatory system. PRR-initiated immune responses must be tightly regulated by the negative regulatory system for the maintenance of immune homeostasis. The immune homeostasis can be disrupted by unrestrictive innate immune response, which may lead to inflammatory conditions in the male genital tracts, an important etiological factor contributing to male infertility. This chapter describes the current understanding of the innate immune responses in the male reproductive system and their effects on male fertility.

Mumps virus (MuV) has high tropism to the testis and may lead to male infertility. Sertoli cells are the major targets of MuV infection. However, the mechanisms by which MuV infection impairs male fertility and Sertoli cell function remain unclear. The present study elucidated the effect of MuV infection on the blood-testis barrier (BTB). The transepithelial electrical resistance of MuV-infected mouse Sertoli cells was monitored, and the expression of major proteins of the BTB was examined. We demonstrated that MuV infection disrupted the BTB by reducing the levels of occludin and zonula occludens 1. Sertoli cells derived from Tlr2(−/−) and Tnfa(−/−) mice were analyzed for mediating MuV-induced impairment. TLR2-mediated TNF-α production by Sertoli cells in response to MuV infection impaired BTB integrity. MuV-impaired BTB was not observed in Tlr2(−/−) and Tnfa(−/−) Sertoli cells. Moreover, an inhibitor of TNF-α, pomalidomide, prevents the disruption of BTB in response to MuV infection. FITC-labeled biotin tracing assay confirmed that BTB permeability and spermatogenesis were transiently impaired by MuV infection in vivo. These findings suggest that the disruption of the BTB could be one of the mechanisms underlying MuV-impaired male fertility, in which TNF-α could play a critical role.—Wu, H., Jiang, X., Gao, Y., Liu, W., Wang, F., Gong, M., Chen, R., Yu, X., Zhang, W., Gao, B., Song, C., Han, D. Mumps virus infection disrupts blood-testis barrier through the induction of TNF-α in Sertoli cells.

The seminal vesicles can be infected by microorganisms, thereby resulting in vesiculitis and impairment in male fertility. Innate immune responses in seminal vesicles cells to microbial infections, which facilitate vesiculitis, have yet to be investigated. The present study aims to elucidate pattern recognition receptor–mediated innate immune responses in seminal vesicles epithelial cells. Various pattern recognition receptors, including Toll-like receptor 3, Toll-like receptor 4, cytosolic ribonucleic acid, and deoxyribonucleic acid sensors, are abundantly expressed in seminal vesicles epithelial cells. These pattern recognition receptors can recognize their respective ligands, thus activating nuclear factor kappa B and interferon regulatory factor 3. The pattern recognition receptor signaling induces expression of pro-inflammatory cytokines, such as tumor necrosis factor alpha (Tnfa) and interleukin 6 (Il6), chemokines monocyte chemoattractant protein-1 (Mcp1) and C–X–C motif chemokine 10 (Cxcl10), and type 1 interferons Ifna and Ifnb. Moreover, pattern recognition receptor-mediated innate immune responses up-regulated the expression of microsomal prostaglandin E synthase and cyclooxygenase 2, but they down-regulated semenogelin-1 expression. These results provide novel insights into the mechanism underlying vesiculitis and its impact on the functions of the seminal vesicles.

Human infertility affects approximately 15% of couples at reproductive age worldwide. Male factors contribute up to 45% of infertility cases. Inflammatory conditions in the male genital system are responsible for about 10% of male infertility in developed countries, and the incidence of immunological infertility can be considerably higher in developing countries where medical cares and environment conditions are poor. In particular, immunological disorders in the testis freguently cause infertility because it is a organ where spermatogenesis carries out. The testis possesses a special immune environment because of its immunoprivileged status and innate immune system. Testicular immune privilege is essential for the protection of male germ cells from detrimental immune responses, whereas the local innate immune system plays a crucial role in the testicular defense against microbial infections. Most of testicular cells are involved in the regulation of testicular immune homeostasis. Disruption of testicular immune homeostasis may result in orchitis, one aetiological factor of male infertility. Multiple mechanisms cooperatively regulate testicular immune homeostasis. Recent studies have revealed important roles of Tyro3/Axl/Mer (TAM) receptor tyrosine kinases and pattern recognition receptors (PRRs) in regulating testicular immune environment. TAM receptors belong to one subfamily of receptor tyrosine kinases. The product of growth arrest-specific gene 6 (Gas6) is common ligand of TAM receptors. TAM and Gas6 are expressed in the testis. TAM triple knockout mice are male sterile and develop autoimmune orchitis. TAM receptors regulate testicular immune homeostasis via three mechanisms: (i) PRR-initiated innate immune responses in Sertoli and Leydig cells are inhibited by TAM/Gas6 signaling, which reduce inflammatory cytokine production in the testis; (ii) TAM receptors promote phagocytosis of apoptotic germ cells by Sertoli cells. The phagocytic clearance of apoptotic germ cells by Sertoli cells can remove germ cell antigens, thereby preventing endogenous immune response; (iii) TAM receptors favor systemic immune tolerance to male germ cells. Experimental autoimmune orchitis (EAO) is a testis-specific autoimmune inflammation in rodent animals after immunization with male germ cell antigens. Axl and Mer double knockout mice are susceptible to EAO induction, suggesting Axl and Mer cooperatively regulate the systemic immune tolerance to male germ cell antigens. Although the testis is an immunoprivileged organ, it can be infected by a broad spectrum of microbial pathogens. However, the testis usually recovers from microbial infections without evident inflammatory response, suggesting that the testis adopts an effective local innate defense system against invading microorganisms. Recognition of microbial pathogens by PRRs initiates innate immune responses, which is the first line of the host defense against invading microbes. Several subfamilies of PRRs are expressed in the testis. In particularly, major testicular cells, including Sertoli, Leydig, and male germ cells, express various PRRs. PRRs can be activated in testicular cells by pathogen molecular patterns, thereby initiating innate immune responses. These observations indicate that testicular cells are equipped with innate immune machinery and may be involved in the testicular defense against microbial infections. This article describes role of TAM receptors and PRRs in regulating immune environment in the testis, and speculates aspects of which need priorizing in future investigation.

Three members of a receptor tyrosine kinase family, including Tyro3, Axl, and Mer, are collectively called as TAM receptors. TAM receptors have two common ligands, namely, growth arrest specific gene 6 (Gas6) and protein S (ProS). The TAM-Gas6/ProS system is essential for phagocytic removal of apoptotic cells, and plays critical roles in regulating immune response. Genetic studies have shown that TAM receptors are essential regulators of the tissue homeostasis in immunoprivileged sites, including the testis, retina and brain. The mechanisms by which the TAM-Gas6/ProS system regulates the tissue homeostasis in immunoprivileged sites are emerging. The roles of the TAM-Gas6/ProS system in regulating the immune privilege were intensively investigated in the mouse testis, and several studies were performed in the eye and brain. This review summarizes our current understanding of TAM signaling in the testis and other immunoprivileged tissues, as well as highlights topics that are worthy of further investigation.

Epididymitis can be caused by infectious and noninfectious etiological factors. While microbial infections are responsible for infectious epididymitis, the etiological factors contributing to noninfectious epididymitis remain to be defined. The present study demonstrated that damaged male germ cells (DMGCs) induce epididymitis in mice. Intraperitoneal injection of the alkylating agent busulfan damaged murine male germ cells. Epididymitis was observed in mice 4 weeks after the injection of busulfan and was characterized by massive macrophage infiltration. Epididymitis was coincident with an accumulation of DMGCs in the epididymis. In contrast, busulfan injection into mice lacking male germ cells did not induce epididymitis. DMGCs induced innate immune responses in epididymal epithelial cells (EECs), thereby upregulating the pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β), as well as the chemokines such as monocyte chemotactic protein-1 (MCP-1), monocyte chemotactic protein-5 (MCP-5), and chemokine ligand-10 (CXCL10). These results suggest that male germ cell damage may induce noninfectious epididymitis through the induction of innate immune responses in EECs. These findings provide novel insights into the mechanisms underlying noninfectious epididymitis, which might aid in the diagnosis and treatment of the disease.

The mammalian testis is a remarkable immunoprivileged site that protects male germ cells from detrimental immune responses. However, a large spectrum of microbial pathogens, including viruses, bacteria, and parasites, may infect the testis. To overcome its immunoprivileged status and enable local defense against microbial infection, the testis has established its own innate immune system. In addition to having immune cells residing in the interstitial spaces, the testis has tissue-specific cells well equipped with innate immune machinery. The testicular immune environment must be tightly regulated to maintain a homeostasis and disruption of testicular immune homeostasis may impair spermatogenesis and male fertility. Understanding the mechanisms underlying testicular immunoregulation aids the development of strategies for the prevention and treatment of immunological impairment of spermatogenesis. Recent studies have revealed that Tyro3, Axl, and Mer (TAM) receptor tyrosine kinases and pattern recognition receptors (PRRs) play important roles in the maintenance of the testicular immunoprivileged status and local innate immune responses against microbial infections. This chapter focuses on the mechanisms by which TAM receptors and PRRs regulate testicular immune homeostasis.

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Mumps virus (MuV) infection has high tropism to the testis and usually leads to orchitis, an etiological factor in male infertility. However, MuV replication in testicular cells and the cellular antiviral responses against MuV are not fully understood. The present study showed that MuV infected the majority of testicular cells, including Leydig cells, testicular macrophages, Sertoli cells, and male germ cells. MuV was replicated at relatively high efficiencies in Sertoli cells compared with Leydig cells and testicular macrophages. In contrast, MuV did not replicate in male germ cells. Notably, testicular cells exhibited different innate antiviral responses against MuV replication. We showed that interferon β (IFN-β) inhibited MuV replication in Leydig cells, macrophages, and Sertoli cells, which were associated with the up-regulation of major antiviral proteins. Autophagy played a critical role in blocking MuV replication in male germ cells. Autophagy was also involved in limiting MuV replication in testicular macrophages but not in Leydig and Sertoli cells. These findings indicate the involvement of the innate defense against MuV replication in testicular cells.

Although wide range of viruses can infect adipose tissues, innate antiviral response of adipose cells has not been investigated. This study focused on innate antiviral system in mouse adipose cells. Major virus sensors including Toll-like receptor 3 (TLR3), melanoma differentiation-associated antigen 5 (MDA5) and retinoic acid-inducible gene I (RIG-I) are constitutively expressed in preadipocytes and adipocytes. Poly(I:C), a common agonist of TLR3, MDA5 and RIG-I, induced the expression of type I interferons (IFN-α/β) in the two types of adipose cells through the activation of IFN-regulatory factor 3 and upregulated pro-inflammatory factors such as TNF-α and IL-6 through the activation nuclear factor kappa B. Moreover, poly(I:C) induced multiple antiviral proteins including IFN-stimulating gene 15, 2'5'-oligoadenylate synthetase and Mx GTPase 1 in preadipocytes and adipocytes. The poly(I:C)-induced innate antiviral response was reduced by TLR3 deficiency and knockdown of MDA5 or RIG-I. Poly(I:C) also inhibited the differentiation of preadipocytes to adipocytes and suppressed the expression of leptin, adiponectin and resistin in mature adipocytes. The results demonstrated that adipose cells are equipped with innate antiviral system, which may modulate the function of adipocytes.

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.

Testicular inflammation, including noninfectious inflammatory responses in the testis, may impair male fertility. Mechanisms underlying the initiation of noninfectious testicular inflammation are poorly understood. In the current study, we demonstrate that damaged spermatogenic cell products (DSCPs) induce expression of various inflammatory mediators, including TNF-α, IL-1β, IL-6, and macrophage chemotactic protein 1 (MCP-1), in Sertoli cells. Notably, the DSCP-induced inflammatory gene expression was significantly reduced by knockout Toll-like receptor (TLR)2 or TLR4, and abolished by double knockout TLR2 and TLR4 (TLR2−/−TLR4−/−). MCP-1 secreted by Sertoli cells after stimulation with DSCPs promotes macrophage migration. We also provide evidence that busulfan-induced spermatogenic cell damages in vivo upregulate TNF-α and MCP-1 expression in Sertoli cells, and facilitate macrophage infiltration into the testis in wild-type mice. These phenomena were not observed in TLR2−/−TLR4−/− mice. Data indicate that DSCPs induce inflammatory gene expression in Sertoli cells via the activation of TLR2 and TLR4, which may initiate noninfectious inflammatory responses in the testis. The results provide novel insights into the mechanisms underlying damaged spermatogenic cell-induced testicular inflammation.

Mumps virus (MuV) infection may lead to oophoritis and perturb ovarian function. However, the mechanisms underlying the activation of innate immune responses to MuV infection in the ovary have not been investigated. This study showed that Toll-like receptor 2 (TLR2) and retinoic acid-inducible gene I (RIG-I) cooperatively initiate innate immune responses to MuV infection in mouse ovarian granulosa cells. Ovarian granulosa cells infected with MuV significantly produced pro-inflammatory cytokines and chemokines, including interleukin-1β (IL-1β), tumor necrosis factor α (TNF-α), monocyte chemotactic protein 1 (MCP-1), and type 1 interferons (IFN-α and IFN-β). Knockdown of RIG-I significantly decreased MuV-induced cytokine expression. TLR2 deficiency reduced the expression of IL-1β, TNF-α, and MCP-1 but did not affect the expression of IFN-α and IFN-β in granulosa cells after infection with MuV. Intraperitoneal injection of MuV induced the ovarian innate immune responses in vivo, which suppressed estradiol synthesis and induced granulosa cell apoptosis. The results provide novel insights into the mechanisms underlying MuV-induced innate immune responses in the mouse ovary.

Uropathogenic Escherichia coli (UPEC) may cause epididymitis and impair male fertility. The mechanisms underlying the innate immune responses to UPEC infection in the epididymis are not fully understood. This study showed that UPEC induced innate immune responses in mouse epididymal epithelial cells (EECs) through the activation of Toll-like receptor 4 (TLR4) and TLR5. Infection with UPEC significantly induced the expression of proinflammatory cytokines, including tumor necrosis factor alpha, interleukin 6, and monocyte chemoattractant protein 1, in EECs through the activation of nuclear factor kappa B. Moreover, UPEC induced the production of type 1 interferons by EECs through the activation of interferon regulatory factor 3. The UPEC-induced innate immune responses were significantly reduced in the EECs of Tlr4 or Tlr5 knockout mice. The innate immune responses were further reduced in Tlr4 and Tlr5 double-knockout EECs. Furthermore, we demonstrated that TLR4 and TLR5 cooperatively initiated the epididymal innate immune responses to UPEC infection in vivo. The results provide novel insights into the mechanisms underlying the epididymal innate immune responses to UPEC infection.

Mumps virus (MuV) infection frequently causes orchitis and impairs male fertility. However, the mechanisms underlying the innate immune responses to MuV infection in the testis have yet to be investigated. This study showed that MuV induced innate immune responses in mouse Sertoli and Leydig cells through TLR2 and retinoic acid-inducible gene I (RIG-I) signaling, which result in the production of proinflammatory cytokines and chemokines, including TNF-α, IL-6, MCP-1, CXCL10 and type 1 interferons (IFN-α and IFN-β). By contrast, MuV did not induce the cytokine production in male germ cells. In response to MuV infection, Sertoli cells produced higher levels of proinflammatory cytokines and chemokines but lower levels of type 1 IFNs than Leydig cells did. The MuV-induced cytokine production by Sertoli and Leydig cells was significantly reduced by the knockout of TLR2 or the knockdown of RIG-I signaling. The local injection of MuV into the testis triggered the testicular innate immune responses in vivo. Moreover, MuV infection suppressed testosterone synthesis by Leydig cells. This is the first study examining the innate immune responses to MuV infection in testicular cells. The results provide novel insights into the mechanisms underlying the MuV-induced innate immune responses in the testis.

Sperm production and androgen synthesis are two key functions of the testis. Immune protection of the testicular functions is critical for the maintenance of reproduction and development of a species. The mammalian testis provides a special immune environment because of its two characteristics: (1) the testis is a remarkable immune privilege site where immunogenic germ cells are protected from detrimental immune attack and (2) the testis adopts effective local innate defense mechanisms against microbial infections. In addition to systemic immune cells, testis-specific cells can also defend against invading microbes through innate immune responses. Breakdown of immune homeostasis in the testis may lead to orchitis and impair male fertility. Understanding the mechanisms underlying testicular immunoregulation would aid in establishing preventive and therapeutic strategies for immune-impaired male fertility.

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.

The testis is an immunoprivileged site, where the local cell-initiated testicular innate immune responses play a crucial role in defense against microbial infections. Mechanisms modulating the testicular cell-built defense system remain to be clarified. In this article, we demonstrate that Leydig cells, a major cell population in the testicular interstitium, initiate innate immunity through the activation of Toll-like receptors (TLRs). Several TLRs are expressed in mouse Leydig cells; among these, TLR3 and TLR4 are expressed at relatively high levels compared with other TLR members. Both TLR3 and TLR4 can be activated by their agonists (polyinosinic:polycytidylic acid and lipopolysaccharide) in Leydig cells and subsequently induce the production of inflammatory factors, such as IL-1β, IL-6, TNF-α, and type 1 interferons (IFN) (IFN-α and IFN-β). Notably, the activation of TLR3 and TLR4 suppresses steroidogenesis by Leydig cells. Further, we provide evidence that Axl and Mer receptor tyrosine kinases are expressed in Leydig cells and regulate TLR-mediated innate immune responses negatively. Data presented here describe a novel function of Leydig cells in eliciting testicular innate immune responses that should contribute to the protection of the testis from microbial infections.

The testis exhibits a distinctive form of immune privilege to protect the germ cells from the host immune attack. The property of testicular immune privilege was originally attributed to the blood-testis barrier in the seminiferous epithelium, which sequesters antigens. Recent studies have uncovered several levels of immune control besides the blood-testis barrier involved in the privilege of the testis, including the mechanisms of immune tolerance, reduced immune activation, localized active immunosuppression and antigen-specific immunoregulation. The somatic cells of the testis, especially Sertoli cells, play a key role in regulating the testicular immune privileged status. The constitutive expression of anti-inflammatory factors in the testis by somatic cells is essential for local immunosuppression. Growing evidence shows that androgens orchestrate the inhibition of proinflammatory factors and shift cytokine balance toward a tolerogenic environment. Disruption of these protective mechanisms, which may be caused by trauma, infection and genetic factors, can lead to orchitis and infertility. This review article highlights the unique immune environment of the testis, particularly focuses on the regulation of testicular immune privilege.

The detrimental effects of Zika virus (ZIKV) infection on mouse testicular functions have reminded a viral threat to male fertility. A broad range of virus families has tropism for male reproductive system, particularly the testes. Certain virus types of these viruses, such as mumps virus and human immunodeficiency virus (HIV), may severely damage the testes and consequently lead to male infertility. ZIKV has been recently found to damage testicular functions and lead to male infertility in mice. Many other viruses also have detrimental effects on host reproduction. Public attention has been paid to sexually transmitted viruses, such as HIV and hepatitis B and C viruses in humans and likewise in economically important farm animals. This article provides an overview on main viruses affecting the male reproductive system and their detrimental effects on fertility, and outlines some important issues for future study.

Removal of apoptotic cells from inflammatory sites by macrophages is an important step in the resolution of inflammation. However, the effect of inflammatory modulators on phagocytic clearance of apoptotic cells remains to be clarified. In this paper, we demonstrate that lipopolysaccharide (LPS), a potent inflammatory agent, inhibits the phagocytosis of apoptotic neutrophils by mouse peritoneal macrophages. This inhibition can be attributed to both LPS-mediated induction of tumour necrosis factor (TNF-α) and suppression of growth arrest-specific gene 6 (Gas6) in macrophages. We found that LPS-induced TNF-α production inhibited phagocytic ability of macrophages in an autocrine manner. In contrast, Gas6 expression in macrophages was blocked by LPS, which also contributes to the inhibition of macrophage phagocytosis by LPS. Our data suggest that phagocytic clearance of apoptotic neutrophils by macrophages can be regulated by local pro- and anti-inflammatory factors in two opposite states.

Mice lacking TYRO3, AXL and MER (TAM) receptor tyrosine kinases (RTKs) are male sterile. The mechanism of TAM RTKs in regulating male fertility remains unknown. In this study, we analyzed in more detail the testicular phenotype of TAM triple mutant (TAM−/−) mice with an effort to understand the mechanism. We demonstrate that the three TAM RTKs cooperatively regulate male fertility, and MER appears to be more important than AXL and TYRO3. TAM−/− testes showed a progressive loss of germ cells from elongated spermatids to spermatogonia. Young adult TAM−/− mice exhibited oligo-astheno-teratozoospermia and various morphological malformations of sperm cells. As the mice aged, the germ cells were eventually depleted from the seminiferous tubules. Furthermore, we found that TAM−/− Sertoli cells have an impaired phagocytic activity and a large number of differentially expressed genes compared to wild-type controls. By contrast, the function of Leydig cells was not apparently affected by the mutation of TAM RTKs. Therefore, we conclude that the suboptimal function of Sertoli cells leads to the impaired spermatogenesis in TAM−/− mice. The results provide novel insight into the mechanism of TAM RTKs in regulating male fertility.

Apoptotic spermatogenic cells and residual bodies are phagocytosed and degraded by Sertoli cells during mammalian spermatogenesis. The meaning of this event remains to be clarified. In this report, we demonstrate that apoptotic spermatogenic cells and residual bodies can be used to produce ATP by Sertoli cells after phagocytosis of them. Sertoli cells produced the highest level of ATP compared with other testicular cells. Phagocytosis assayin vitroshowed that engulfment of apoptotic spermatogenic cells increases ATP production by Sertoli cells. The increased ATP production was detected in seminiferous tubules at the stages where phagocytosis occurs. Induced apoptosis of spermatogenic cellsin vivoincreased ATP production in seminiferous tubules. The augmentation of ATP production bothin vitroandin vivoassociated with the lipid formation in Sertoli cells after phagocytosis of apoptotic spermatogenic cells. The lipid β-oxidation was a predominant pathway to produce ATP in Sertoli cells. We conclude that after phagocytosis by Sertoli cells, apoptotic spermatogenic cells are degraded to form lipids that are then used to produce ATP. The results suggest that apoptotic spermatogenic cells can be energy sources for Sertoli cells that may define a novel meaning of spermatogenic cell death.