Fichorova Raina N, Mendonca Kevin, Yamamoto Hidemi S, Murray Ryan, Chandra Neelima, Doncel Gustavo F
Laboratory of Genital Tract Biology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA.
Laboratory of Genital Tract Biology, Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital and Harvard Medical School, Boston, MA, USA.
Toxicol Appl Pharmacol. 2015 Jun 15;285(3):198-206. doi: 10.1016/j.taap.2015.02.017. Epub 2015 Mar 25.
Any vaginal product that alters the mucosal environment and impairs the immune barrier increases the risk of sexually transmitted infections, especially HIV infection, which thrives on mucosal damage and inflammation. The FDA-recommended rabbit vaginal irritation (RVI) model serves as a first line selection tool for vaginal products; however, for decades it has been limited to histopathology scoring, insufficient to select safe anti-HIV microbicides. In this study we incorporate to the RVI model a novel quantitative nuclease protection assay (qNPA) to quantify mRNA levels of 25 genes representing leukocyte differentiation markers, toll-like receptors (TLR), cytokines, chemokines, epithelial repair, microbicidal and vascular markers, by designing two multiplex arrays. Tissue sections were obtained from 36 rabbits (6 per treatment arm) after 14 daily applications of a placebo gel, saline, 4% nonoxynol-9 (N-9), and three combinations of the anti-HIV microbicides tenofovir (TFV) and UC781 in escalating concentrations (highest: 10% TFV+2.5%UC781). Results showed that increased expression levels of toll-like receptor (TLR)-4, interleukin (IL)-1β, CXCL8, epithelial membrane protein (EMP)-1 (P<0.05), and decreased levels of TLR2 (P<0.05), TLR3 and bactericidal permeability increasing protein (BPI) (P<0.001) were associated with cervicovaginal mucosal alteration (histopathology). Seven markers showed a significant linear trend predicting epithelial damage (up with CD4, IL-1β, CXCL8, CCL2, CCL21, EMP1 and down with BPI). Despite the low tissue damage RVI scores, the high-dose microbicide combination gel caused activation of HIV host cells (SLC and CD4) while N-9 caused proinflammatory gene upregulation (IL-8 and TLR4) suggesting a potential for increasing risk of HIV via different mechanisms depending on the chemical nature of the test product.
任何改变黏膜环境并损害免疫屏障的阴道产品都会增加性传播感染的风险，尤其是艾滋病毒感染，艾滋病毒在黏膜损伤和炎症环境中易于滋生。美国食品药品监督管理局（FDA）推荐的兔阴道刺激（RVI）模型是阴道产品的一线筛选工具；然而，几十年来，它一直局限于组织病理学评分，不足以筛选出安全的抗艾滋病毒杀微生物剂。在本研究中，我们通过设计两个多重阵列，将一种新型的定量核酸酶保护分析（qNPA）纳入RVI模型，以量化代表白细胞分化标志物、Toll样受体（TLR）、细胞因子、趋化因子、上皮修复、杀微生物和血管标志物的25个基因的mRNA水平。在每天应用安慰剂凝胶、生理盐水、4%壬苯醇醚-9（N-9）以及三种浓度递增的抗艾滋病毒杀微生物剂替诺福韦（TFV）和UC781组合（最高浓度：10%TFV + 2.5%UC781）14天后，从36只兔子（每个治疗组6只）获取组织切片。结果显示，Toll样受体（TLR）-4、白细胞介素（IL）-1β、CXCL8、上皮膜蛋白（EMP）-1的表达水平升高（P<0.05），而TLR2（P<0.05）、TLR3和杀菌通透性增加蛋白（BPI）的水平降低（P<0.001），这些与宫颈阴道黏膜改变（组织病理学）相关。七个标志物显示出预测上皮损伤的显著线性趋势（CD4、IL-1β、CXCL8、CCL2、CCL21、EMP1升高，BPI降低）。尽管RVI评分显示组织损伤较低，但高剂量杀微生物剂组合凝胶导致艾滋病毒宿主细胞（SLC和CD4）活化，而N-9导致促炎基因上调（IL-8和TLR4），这表明根据测试产品的化学性质，通过不同机制增加艾滋病毒感染风险的可能性。
