MACAQUE EXPLANT MODEL FOR MICROBICIDE TESTING

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 背景资料：50 mM MIV-150和14 mM醋酸锌二水合物的组合在施用后至少24小时内提供完全的感染保护。我们假设MIV-150/醋酸锌组合凝胶将保护猕猴阴道粘膜外植体免受SHIV RT感染。研究方法：在体外用SHIV RT感染之前，用PHA/IL-2刺激非极化的阴道外植体（来自未处理和再循环的SHIV RT暴露/感染的动物的活组织检查或尸检组织）。测试在角叉菜胶中配制的MIV-150和乙酸锌的抗病毒活性，并与仅含有乙酸锌或MIV-150的凝胶的活性进行比较。用p27 ELISA和SIVgagPCR监测组织感染。结果：经PHA/IL-2过夜转运刺激的猕猴阴道粘膜对SHIV RT的体外感染敏感。体外未暴露于病毒的PHA/IL-2刺激的组织包括在所有实验中作为阴性对照。在存在MIV-150/乙酸锌组合凝胶以及仅含MIV-150或乙酸锌的凝胶的情况下外植体的感染导致针对感染的完全保护。在基于角叉菜胶的媒介物存在下的感染导致延迟的感染动力学或完全保护。当组织暴露于更高剂量的SHIV RT时，载体的这种保护作用被克服。暴露于含MIV-150/乙酸锌的凝胶后24小时攻击的组织没有被感染（记忆效应），而用角叉菜胶载体预处理的组织像没有凝胶处理的对照一样被感染。比较凝胶记忆效应的实验正在进行中。结论：当在病毒暴露前24小时施用时，新型MIV-150/乙酸锌组合凝胶（但不是角叉菜胶载体）在猕猴阴道外植体中有效对抗SHIV RT。这反映了阴道应用后在体内观察到的情况，支持在体内动物试验之前使用该外植体系统预筛选候选杀微生物剂。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Background: Combination of 50mM MIV-150 and 14mM zinc acetate dihydrate provides complete protection from infection for at least 24h after application. We hypothesized that the MIV-150/zinc acetate combination gel will protect against SHIV RT infection in macaque vaginal mucosal explants. Methods: Non-polarized vaginal explants (biopsies or necropsy tissues derived from naive and recycled SHIV RT exposed/infected animals) were stimulated with PHA/IL-2 before infection in vitro with SHIV RT. Anti-viral activity of MIV-150 and zinc acetate formulated in carrageenan was tested and compared with activity of gels containing only zinc acetate or MIV-150. Tissue infection was monitored using p27 ELISA and SIVgag PCR. Results: Overnight transported PHA/IL-2 stimulated macaque vaginal mucosa is susceptible to infection with SHIV RT in vitro. PHA/IL-2 stimulated tissues not exposed to virus in vitro were included in all experiments as negative controls. Infection of explants in the presence of the MIV-150/zinc acetate combination gel, as well as MIV-150 or zinc acetate only containing gels, resulted in complete protection against infection. Infection in the presence of carrageenan-based vehicle resulted in delayed infection kinetics or complete protection. This protective effect of vehicle was overcome when tissues were exposed to a higher dose of SHIV RT. Tissues challenged 24h after exposure to the MIV-150/zinc acetate containing gel did not become infected (memory effect), while tissues pretreated with the carrageenan vehicle were infected like no gel treated controls. Experiments comparing the memory effect of the gels are ongoing. Conclusion: A novel MIV-150/zinc acetate combination gel (but not the carrageenan vehicle) is effective against SHIV RT in macaque vaginal explants when applied 24h prior to virus exposure. This mirrors what was observed in vivo following vaginal application supporting the use of this explant system to prescreen candidate microbicides prior to in vivo animal testing.