Role of microRNAs in opioid drug abuse induced persistent Inflammation and HIV di

描述（由申请人提供）：慢性药物使用与艾滋病毒感染易感性增加之间存在很强的相关性。慢性药物使用者约占美国所有艾滋病毒病例的三分之一，阿片类药物滥用者向艾滋病的进展明显加快。慢性免疫激活伴随促炎细胞因子血清水平升高是疾病进展的标志。最近的研究表明，血浆内毒素（LPS）升高，肠道细菌移位的结果，可能是免疫激活的HIV感染的可能原因。已经发现免疫活化和T细胞活化/耗竭之间的相关性。有趣的是，研究表明，使用静脉注射海洛因的HIV患者与未使用药物的HIV感染患者相比，显示出更高的血清LPS水平。类似地，我们在药物滥用的鼠模型中显示，与安慰剂治疗的动物相比，内毒素的循环水平更高。这些作用在TATtg小鼠中进一步增强。如在人类患者中所观察到的，这些动物模型中的免疫激活也被延长和持续。这些结果暗示LPS在驱动药物滥用HIV感染患者的疾病进展中的作用。然而，奇怪的是，有充分的证据表明，长时间的LPS暴露导致LPS耐受性和LPS诱导的巨噬细胞刺激减少。然而，持续的免疫激活是HIV患者和HIV动物模型中疾病进展的标志。这种不一致的观察背后的机制是知识上的巨大差距。在LPS耐受中，转录和翻译抑制事件联合收割机紧密调节促炎基因。最近，microRNA被认为是在转录后调节水平上控制多种生物过程的负调节因子。我们在初步数据中显示，在长期接受LPS治疗的动物中，两种关键miRNA（miR-155和miR-146 a）显着诱导。相比之下，miR-155和miR-146 a在吗啡或达特处理的动物中均显著下调。LPS激活Toll样受体4以诱导促炎细胞因子和趋化因子的转录（32-34）。LPS诱导miR-155和miR-146 a可作为TLR 4表达和信号传导的负反馈调节剂，从而防止促炎细胞因子的过度活化。我们假设，通过吗啡和达特抑制LPS诱导的miR-155和miR-146 a的诱导，使抑制性反馈环失调，导致持续的TLR 4表达和信号传导（图1）。具体目标1：我们将检验LPS诱导miR-155抑制TLR 4表达的假设，并确定吗啡和TAT介导的miR-155衰减导致更大和持续的TLR 4表达。具体目标2：我们将检验以下假设：LPS诱导miR-155和miR-146 a作为TLR 4信号传导的负反馈调节剂，并且吗啡和达特调节miR-155和146 a导致持续的TLR 4信号传导。具体目的3：确定吗啡和达特调节LPS诱导的miR-155和146 a表达的机制。具体目标4：使用WT和转基因动物模型确定吗啡和达特调节LPS诱导的miR-155和miR-146 a的体内机制和结果。这些研究的结果将有助于开发新的治疗策略，以减弱HIV感染患者和HIV感染药物滥用人群的免疫激活并逆转HIV疾病进展。

DESCRIPTION (provided by applicant): There is a strong correlation between chronic drug use and increased susceptibility to HIV infection. Chronic drug users account for approximately one third of all cases of HIV in the USA and progression to AIDS is markedly accelerated in opiate drug abusers. Chronic immune activation with increased serum levels of proinflammatory cytokines is a hallmark of progressive disease. Recent studies show that elevated plasma endotoxin (LPS), a consequence of gut bacterial translocation, may be the likely cause of immune activation in HIV infection. A correlation has been found between immune activation and T cell activation/depletion. Interestingly, studies show that HIV patients that use intravenous heroin display higher serum levels of LPS when compared to non-drug using HIV infected patients. Similarly, we show in a murine model of drug abuse, higher circulating levels of endotoxin when compared to placebo treated animals. These effects are further potentiated in the TATtg mice. As observed in human patients, immune activation in these animal models were also prolonged and sustained. These results implicate a role for LPS in driving the disease progression in drug abusing HIV infected patients. Paradoxically, however, it is well documented that, prolonged LPS exposure leads to LPS tolerance with decreased LPS induced macrophage stimulation. Yet, sustained immune activation is a hallmark of disease progression in both HIV patients and in animal models of HIV. The mechanism underlying this discordant observation is a significant gap in knowledge. In LPS tolerance, transcriptional and translational repressive events combine to tightly regulate proinflammatory genes. More recently, microRNAs have been implicated as negative regulators controlling diverse biological processes at the level of post-transcriptional regulation. We show in preliminary data a significant induction of two key miRNAs, miR-155 and miR-146a in animals that are chronically treated with LPS. In contrast, both miR-155 and miR-146a are significantly down regulated in animals that are morphine or TAT treated. LPS activates Toll like receptor 4 to induce transcription of proinflammatory cytokines and chemokines (32-34). LPS induction of miR-155 and miR-146a may act as negative feedback regulators of TLR4 expression and signaling thereby preventing excessive activation of pro-inflammatory cytokines. We hypothesize, that suppression of LPS induced induction of miR-155 and miR-146a, by Morphine and TAT, deregulates the inhibitory feedback loop resulting in sustained TLR4 expression and signaling (Fig. 1). In Specific aim 1: We will test the hypothesis that LPS induction of miR-155 inhibits TLR4 expression and establish that morphine and TAT-mediated attenuation of miR-155 results in greater and sustained TLR4 expression. Specific aim 2: We will test the hypothesis that LPS induction of miR-155 and miR-146a acts as negative feedback regulators of TLR4 signaling and morphine and TAT modulation of miR-155 and 146a results in persistent TLR4 signaling. Specific Aim 3: Determine the mechanism by which Morphine and TAT modulate LPS induced miR-155 and 146a expression. Specific Aim 4: Determine the mechanism and consequence of morphine and TAT modulation of LPS induced miR-155 and miR-146a in vivo using WT and Transgenic animal models. The results from these studies will allow for the development of new therapeutic strategies to attenuate immune activation and reverse HIV disease progression both in HIV infected patients and in HIV infected drug abusing population.