Development of a novel mouse model with progesterone-independent susceptibility to genital HSV infection

生殖器疱疹是一种由单纯疱疹病毒(HSV)引起的不可治愈的性传播感染(STI)，对妇女的影响不成比例，可能导致特定性别的不良事件，如感染其他STI的风险增加以及在分娩期间垂直传播。小鼠通常被用作研究免疫和阴道HSV感染发病机制的临床前模型。然而，小鼠模型的一个主要缺点是，与人类不同，小鼠只在发情周期的某些阶段容易感染。因此，雌性小鼠通常会用以孕酮为基础的激素避孕药Depo-provera(DMPA，Depo-Provera)进行预处理，以保持小鼠处于发情间期，并确保对HSV感染的一致易感性。然而，DMPA的使用将宿主-病原体相互作用的研究限制在发情周期的单个阶段，尽管有大量证据表明，女性性激素对女性生殖道的抗病毒防御和免疫监控具有重要影响。此外，DMPA的使用排除了与共生细菌或真菌的联合接种研究，因为这些微生物通常需要高雌激素环境才能在小鼠阴道强劲定植。小鼠和人类阴道的一个主要区别是，病毒进入受体Nectin-1在小鼠上皮细胞上的表达是激素依赖性的，但在人类中是结构性表达的。为了克服这一挑战，该提案将开发一种新的小鼠模型，在该模型中，Nectin-1通过转录和翻译后调节机制在小鼠阴道上皮中结构性表达。Nectin-1的表达将被置于两个不同的构成启动子的控制之下，并将引入两个突变来防止胞外结构域的切割。Nectin-1的上皮特异性表达将通过包含一个停滞盒并繁殖到Cre重组酶由人角蛋白14启动子控制的小鼠来实现。最后，将在发情周期的不同阶段测试这些新小鼠对HSV的敏感性。HSV感染新的、更具翻译性的小鼠模型的成功开发将有助于实现我们的长期目标，即探索性激素在调节阴道屏障宿主-病原体相互作用中的作用，以及研究阴道微生物失调或STI联合感染对HSV发病机制的分子影响。我们预计未来的研究将揭示针对生殖器疱疹的保护性免疫的未知方面，包括由疫苗诱导的免疫，并可能导致发现旨在降低对HSV感染易感性的治疗的新靶点。

Genital herpes, an incurable sexually transmitted infection (STI) caused by herpes simplex virus (HSV), disproportionately affects women and can cause sex-specific adverse events such as increased risk of acquiring other STIs as well as vertical transmission during birth. The mouse is commonly used as a preclinical model to study immunity and pathogenesis of vaginal HSV infection. However, one major drawback of the murine model is that unlike humans, mice are susceptible to infection only at certain stages of the estrus cycle. Thus, female mice are typically pre-treated with depot medroxyprogesterone acetate (DMPA, Depo-Provera), a progesterone-based hormonal contraceptive, to hold the mice in diestrus and ensure uniform susceptibility to HSV infection. However, the use of DMPA limits the study of host-pathogen interactions a single stage of the estrus cycle, despite abundant evidence that female sex hormones have a major impact on antiviral defense and immunosurveillance of the female reproductive tract. Furthermore, use of DMPA precludes co-inoculation studies with commensal bacteria or fungi, as these microbes often require an estrogen-high environment to robustly colonize the murine vagina. One major difference between the mouse and human vagina is that expression of the viral entry receptor nectin-1 on epithelial cells is hormone dependent in mice, but constitutively expressed in humans. To overcome this challenge, this proposal will develop a novel mouse model in which nectin-1 is constitutively expressed in the murine vaginal epithelium by addressing transcriptional and post-translational regulatory mechanisms. Nectin-1 expression will be placed under the control two different constitutive promoters, and two mutations will be introduced to prevent ectodomain cleavage. Epithelium-specific expression of nectin-1 will be achieved through the inclusion of a stop-flox cassette and breeding to mice in which Cre recombinase expression is controlled by the human keratin 14 promoter. Finally, the HSV susceptibility of these novel mice will be tested at different stages of the estrus cycle. The successful development of a novel, more translational mouse model of HSV infection would help achieve our long-term goal of exploring the role of sex hormones in modulating host-pathogen interactions at the vaginal barrier as well as investigating the molecular impact of vaginal microbial dysbiosis or STI co- infection on HSV pathogenesis. We expect future studies to reveal unknown aspects of protective immunity against genital herpes, including that induced by vaccines, and potentially lead to the discovery of novel targets for therapeutics aimed at reducing susceptibility against HSV infection.