Extracellular vesicles in semen and genital HIV infection and immunity during heroin addiction and methadone or buprenorphine substitution therapy

 描述(申请人提供)：我们最近发现人类精液含有数万亿个细胞外小泡(SEV)，携带可能调节免疫系统的小细胞外RNA(ExRNA)分子(核酸研究，42：7290-7304,2014)。在未发表的研究中，我们发现SEV抑制适应性T细胞反应，很可能是通过影响抗原提呈。SEV在体外也与艾滋病毒病毒粒子有关，并在我们手中增强艾滋病毒的易感性，尽管另一个小组最近报告了相反的情况。最终，SEV可能通过直接与HIV及其靶细胞相互作用以及通过 危害对HIV的保护性T细胞免疫。阿片类药物也会影响免疫系统，尽管确切的机制和功能结果尚不清楚。海洛因滥用会导致免疫功能受损，增加艾滋病毒的易感性，但造成这种情况的行为和生物原因很难解决。海洛因成瘾通常使用长期的美沙酮或丁丙诺啡药物治疗，这两种阿片类药物的免疫学特性和对艾滋病毒感染的影响也尚不清楚。因此，精液和阿片类药物使用中细胞外小泡的相互作用可能会对艾滋病毒感染和免疫系统产生重要影响，本提案将调查这些联系。我们将在阴道移植模型中确定SEV对HIV感染的影响，并剖析SEV介导的适应性T细胞反应抑制的性质和程度，包括对HIV疫苗的抑制。我们将在开始辅助美沙酮或丁丙诺啡治疗前和三个月后，对正常捐赠者以及海洛因使用者的血液和精液样本中的艾滋病毒易感性、适应性T细胞反应和SEV生物学(exRNA组成和功能)进行比较研究。这三种阿片类药物也将在培养中进行测试，因为阿片受体1受体在阴道白细胞上的表达(我们的数据)，以及其他非阿片受体的报告，如Toll样受体4，表明阿片类药物可能直接影响白细胞生物学。SEV和阿片类药物的相互作用将在组合实验中讨论。这些研究将加深我们对精液中的阿片类药物和细胞外小泡如何单独或联合影响艾滋病毒性传播的理解。了解这些影响将有助于改善艾滋病毒疫苗接种结果，重点关注其他艾滋病毒预防战略，并就美沙酮与丁丙诺啡替代疗法的临床益处提供新的信息。

 DESCRIPTION (provided by applicant): We have recently shown that human semen contains trillions of extracellular vesicles (SEV) carrying small extracellular RNA (exRNA) molecules that may regulate the immune system (Nucleic Acids Research, 42:7290-7304, 2014). In unpublished studies, we have found that SEV suppress adaptive T cell responses, most likely by affecting antigen presentation. SEV also associate with HIV virions in vitro and in our hands enhance HIV susceptibility, although another group recently reported the opposite. Ultimately, SEV likely impact HIV infection, both by directly interacting with HIV and its target cells, and by compromising protective T cell immunity against HIV. Opioids too affect the immune system, although the exact mechanisms and functional outcomes are not well- understood. Heroin abuse leads to an immunocompromised state and increases HIV susceptibility, but the behavioral and biological causes for this are difficult to untangle. Heroin addiction is often treated with long- term methadone or buprenorphine medication, two opioids whose immunological properties and effects on HIV infection also remain unclear. Thus, the interplay of extracellular vesicles in semen and opioid use may have important consequences for HIV infection and the immune system, and this proposal will investigate these connections. We will determine the effects of SEV on HIV infection in a vaginal explant model and dissect the nature and extent of SEV- mediated inhibition of adaptive T cell responses, including to HIV vaccination. We will perform comparative studies of HIV susceptibility, adaptive T cell responses and SEV biology (exRNA composition and function) in blood and semen samples from normal donors, as well as heroin users before and three months after starting assisted methadone or buprenorphine treatment. The three opioids will also be tested in culture, because expression of the Opioid Receptor-Like 1 receptor on vaginal leukocytes (our data), and reports of other non- opioid receptors such as Toll-like receptor 4, indicate that opioids could directly affect leukocyt biology. SEV and opioid interactions will be addressed in combinatorial experiments. These studies will deepen our understanding of how opioids and extracellular vesicles in semen, alone or in conjunction, impact sexual HIV transmission. Knowledge of these effects will help to improve HIV vaccination outcomes, focus other HIV prevention strategies, and provide new information on the clinical benefits of methadone versus buprenorphine substitution therapy.