Under attack: Modulation of the blood-testes barrier by Zika virus

摘要 2015-2016年寨卡病毒（ZIKV）的流行导致了严重的意外临床结果，包括 和格林-巴利综合征。ZIKV主要是一种蚊子传播的病毒， 随着艾滋病毒/艾滋病感染者人数的惊人增加， ZIKV感染妇女中的小头畸形病例与精液中病毒的检测一致。 ZIKV与血精症的关联以及当病毒血症时精液中存在传染性ZIKV。 在人类中不可检测的结果表明ZIKV在睾丸中建立感染的能力。缺乏任何 用于对抗ZIKV感染和预防睾丸感染的疫苗或特异性治疗药物， 迫切需要开发体外系统和体内模型，以了解 相关机制。生精小管是睾丸内精子发生的部位，是一个重要的细胞， 免疫特权部位，具有有效的血液-睾丸屏障（BTB），可防止微生物感染， 保护免疫原性生殖细胞免受系统性免疫攻击。BTB完全由以下人员组成： 相邻支持细胞（SC）之间的特化紧密连接蛋白（TJP）提供了物理和 生精小管的免疫屏障。炎症介质的产生增加， 如TNF-α、IL-8、基质金属蛋白酶（MMPs）和细胞粘附分子（CAM）， 建议破坏BTB并促进睾丸感染。虽然，最近的令人信服的数据 研究表明ZIKV具有感染睾丸的能力，这可能使其逃避免疫检测， 关于睾丸的ZIKV感染知之甚少。该提案旨在确定 ZIKV通过利用使用原代人SC的体外BTB模型系统和体内BTB模型系统进入睾丸 非人类灵长类动物模型我们将讨论这样一个假设，即即使在睾丸中保持传染性， 在病毒从外周清除后，ZIKV感染SC和睾丸巨噬细胞， 炎症反应，改变TJP的表达，导致BTB破坏，并允许进入 病毒进入生精小管，从而在睾丸中建立持续感染。我们集团 由于我们之前的研究，我们的研究定位独特，可以描述ZIKV进入睾丸的机制。 利用体外血脑屏障系统和体内模型研究WNV神经侵袭的经验 机制等具体目标1将确定无细胞和巨噬细胞相关的ZIKV如何在细胞内表达。 穿过人体外BTB模型并影响屏障完整性。我们还将评估， ZIKV对包括细胞因子、MMPs和CAM在内的炎性介质的诱导可以影响 多个TJP和BTB完整性的表达。具体目标2将评估ZIKV是否影响BTB 完整性，并确定与病毒持久性的关系 以及睾丸中的炎症和抗病毒反应。预计，来自 体外人体系统和高度相关的NHP模型将填补NHP机制的根本空白， ZIKV进入睾丸并增强我们对睾丸病毒特异性免疫反应的理解 感染这些信息可以共同指导未来的策略，以操纵系统， 开发从睾丸中根除病毒的新方法。

Abstract The 2015-2016 epidemic of Zika virus (ZIKV) has caused severe unexpected clinical outcomes including microcephaly and Guillain-Barre Syndrome. ZIKV is primarily a mosquito-transmitted virus but the concerns regarding sexual transmission has gained urgency as the alarming increase in the number of cases of microcephaly in ZIKV-infected women coincided with the detection of the virus in the semen. Association of ZIKV with hematospermia and presence of infectious ZIKV in the semen when viremia is undetectable in humans suggest the ability of ZIKV to establish infection in the testes. Lack of any vaccine or specific therapeutic drugs to combat ZIKV infection and prevent testicular infection has brought an urgent need for the development of in vitro systems and in vivo models to understand the associated mechanisms. Seminiferous tubules, the site of spermatogenesis within the testes, is an immune privileged site with an effective blood-testes barrier (BTB) that prevents microbial infections and protects immunogenic germ cells from systemic immune attack. The BTB, constituted exclusively by specialized tight junctions proteins (TJP) between the adjacent sertoli cells (SC) provides physical and immunological barrier to the seminiferous tubules. Increased production of inflammatory mediators such as TNF-α, IL-8, matrix metalloproteinases (MMPs) and cell-adhesion molecules (CAM) has been suggested to disrupt BTB and facilitate testicular infections. Although, compelling data from recent studies suggest that ZIKV has the ability to infect testes that may allow escape from immune detection, little is known about ZIKV infection of the testes. This proposal seeks to determine the mechanisms of ZIKV entry into the testes by utilizing an in vitro BTB model system using primary human SC and in vivo nonhuman primate model. We will address the hypothesis that to remain infectious in the testes even after the virus is cleared from the periphery, ZIKV infects SC and testes macrophages to induce inflammatory response that alters the expression of TJP, leading to BTB disruption and allowing entry of the virus into the seminiferous tubules thus establishing persistent infection in the testes. Our group is uniquely positioned to delineate the mechanisms of ZIKV testicular entry because of our previous experience of utilizing in vitro blood-brain barrier system and in vivo models to study WNV-neuroinvasion mechanisms. The specific aim 1 will determine how cell-free and macrophage-associated ZIKV transmigrates across the human in vitro BTB model and affects the barrier integrity. We will also assess if the induction of inflammatory mediators including cytokines, MMPs and CAM by ZIKV can affect expression of multiple TJP and BTB integrity. Specific Aim 2 will evaluate whether ZIKV affects the BTB integrity in the experimentally infected macaques and determine the relationship with persistence of virus and inflammatory and antiviral response in the testes. It is anticipated that the integration of data from the in vitro human system and highly relevant NHP model will fill the fundamental gap of the mechanism of ZIKV-testes entry and enhance our understanding of the specific immune response to testicular virus infection. Together this information may guide future strategies to manipulate the system for the development of novel approaches to eradicate virus from the testes.