Effects of Antiretroviral Therapy on Telomerase Function in Human Oral Epithelium

描述（由申请人提供）：本项目的长期目标是减少或逆转感染人类免疫缺陷病毒（HIV）的患者接受高效抗逆转录病毒治疗（HAART）后的口腔并发症。虽然在引入HAART后，HIV感染的口腔表现显著减少，但也报告了口腔粘膜的几种不良反应，包括复发性口腔溃疡、上皮萎缩、多形性红斑、上皮脱落、发疹性鳞屑和多发性口腔疣。本申请的目的是阐明逆转录酶抑制剂（RTI）对端粒酶抑制在介导HAART副作用中的作用。端粒酶是一种细胞逆转录酶，具有至少两种不同的生物学功能：（1）合成端粒DNA和（2）维持基因组完整性。本实验室在正常人口腔上皮来源的角质形成细胞（NHOK）中发现了高水平的端粒酶活性。NHOK中的端粒酶活性与活跃的增殖细胞特异性相关，并且在细胞衰老期间完全丧失。重要的是，端粒酶活性可以有效地抑制几个RTIs常用的HAART。该建议的中心假设是，RTIs对NHOK的端粒酶抑制是导致口腔上皮再生能力降低和HIV+患者长期HAART相关的不良口腔粘膜并发症的原因。为了验证我们的假设，我们提出了三个具体的目标：（1）在体外和在有和没有HAART的HIV+患者来源的细胞中，确定RTI暴露于NHOK中的端粒酶活性、端粒状态和细胞表型改变;（2）确定RTI/HAART对NHOK中DNA修复活性、突变频率和遗传完整性的影响;（3）研究AZT对表达外源性端粒酶或获得增强的复制潜能的NHOK表型改变的影响。目的1和2将研究RTI/HAART在口腔上皮中的详细表型和遗传效应。在目标3中，我们将确定是否增加细胞端粒酶活性和/或“引发”细胞具有增强的复制潜力可以防止AZT的不良表型效应。该项目的成果将用于预防和管理艾滋病毒感染和获得性免疫缺陷综合症（艾滋病）的口腔表现，减少高效抗逆转录病毒疗法的负面影响。

DESCRIPTION (provided by applicant): The long-term goal of this project is to reduce or reverse the oral complications of highly active antiretroviral therapy (HAART) in patients infected with the human immunodeficiency virus (HIV). Although the oral manifestations of HIV infection have significantly decreased after the introduction of HAART, several adverse effects have also been reported in the oral mucosa, including recurrent oral ulceration, epithelial atrophy, erythema multiforme, epithelial desquamation, eruptive chielitis, and multiple oral warts. The purpose of the current application is to elucidate the effects of telomerase inhibition by reverse transcriptase inhibitors (RTIs) in mediating the side effects of HAART. Telomerase is a cellular reverse transcriptase with at least two distinct biological functions in (1) synthesis of telomere DNA and (2) maintenance of genome integrity. Our laboratory found remarkably high level of telomerase activity in normal human oral keratinocytes (NHOK) derived from oral epithelium. Telomerase activity in NHOK is specifically associated with actively proliferating cells and is completely lost during cellular senescence. Importantly, telomerase activity can be effectively inhibited by several RTIs commonly used as HAART. The central hypothesis of this proposal is that telomerase inhibition in NHOK by RTIs is responsible for the diminution of regenerative capacity of the oral epithelium and adverse oral mucosal complications associated with long-term administration of HAART in HIV+ patients. To test our hypothesis, we propose three Specific Aims: (1) to determine the telomerase activity, telomeric status, and cellular phenotypic alterations in NHOK exposed to RTIs in vitro and in cells derived from HIV+ patients with and without HAART; (2) to determine the effects of RTI/HAART on the DNA repair activities, mutation frequency, and genetic integrity in NHOK; and (3) to investigate the effects of AZT on phenotypic alterations in NHOK expressing exogenous telomerase or acquiring enhanced replication potential. The aims 1 and 2 will investigate the detailed phenotypic and genetic effects of RTI/HAART in oral epithelium. In aim 3, we will determine whether augmenting cellular telomerase activity and/or "priming" the cells with enhanced replicative potential can prevent the adverse phenotypic effects of AZT. The outcome of this project will be used for prevention and management of oral manifestations of HIV infection and the acquired immunodeficiency syndrome (AIDS) by reducing the negative effects of HAART.