Project 4: Intracellular mechanism of virus persistence in astrocytes/macrophages

大脑可能是人类免疫缺陷病毒(HIV)的重要储存库。很明显，HIV感染了入侵的巨噬细胞、常驻的小胶质细胞和大脑中的星形胶质细胞。然而，病毒长时间驻留在这些细胞中的基本机制尚不清楚。目前尚不清楚抗逆转录病毒治疗是否能够控制这些细胞中的病毒复制，或者是否对大脑中的病毒库产生影响。因此，我们建议确定病毒进入这些细胞后维持病毒的机制，并确定抗逆转录病毒治疗对这些细胞中病毒生命周期的影响。我们假设在大脑中建立病毒库涉及特定的抗病毒基因 在巨噬细胞和星形胶质细胞中有不同的调节，一旦储存库建立，抗逆转录病毒治疗对病毒在这些细胞中的持久性几乎没有影响。我们将使用体外人脑培养和血液来源的巨噬细胞的组合来解决以下目标。 这些研究将是对项目1和项目2和项目3的补充，项目1将研究病毒在体内和体外巨噬细胞中的持久性，项目2和项目3将确定体内抗逆转录病毒治疗对大脑的影响。我们实验室的初步数据表明，HIV的Tat蛋白调节着几个干扰素诱导的抗病毒基因的表达。我们已经创造了几个新的细胞系和病毒结构，这将使我们能够解决这些与大脑中的病毒库有关的问题。因此，我们提出了以下具体目标：1.确定抗病毒蛋白、早幼粒细胞白血病蛋白(PML)在HIV中的作用 星形胶质细胞和巨噬细胞的复制。2.检测HIV对星形胶质细胞的影响 巨噬细胞的抗病毒基因、干扰素刺激基因-20(ISG-20)和寡腺苷合成酶(OAS-1)。3：研究CART对HIV在星形胶质细胞和巨噬细胞中复制和持续时间的影响。

The brain is likely an important reservoir for the human immunodeficiency virus (HIV). It is clear that HIV infects the invading macrophages, resident microglia and astrocytes in the brain. However, the fundamental mechanisms by which the virus resides in these cells for prolonged periods of time is not understood. It also remains unknown if antiretroviral therapy can control viral replication in these cells or have an impact on the viral reservoirs in brain. We thus propose to determine mechanisms involved in maintaining the virus in these cell types post viral entry and determine the effect of antiretroviral therapy on the viral lifecycle in these cells. We hypothesize that establishment of a viral reservoir in brain involves specific antiviral genes which are differentially regulated in macrophages and astrocytes and once the reservoir is established, antiretroviral therapy will have little or no effect on the persistence of the virus in these cells. We will use a combination of in vitro human brain cultures and blood derived macrophages to address the aims below. These studies will compliment those in project #1 which will study viral persistence in macrophages in vivo and ex vivo and projects #2 and 3 that will determine the effect of antiretroviral therapy on brain in vivo. Preliminary data from our laboratory suggests that the Tat protein of HIV regulates the expression of several interferon inducible antiviral genes. We have generated several novel cell lines and viral constructs that will allow us to address these issues related to viral reservoirs in the brain. We thus propose the following specific aims: 1: To determine the role of antiviral protein, promelyocytic leukemia protein (PML) in HIV replication in astrocytes and macrophages. 2. To determine the effect of HIV in astrocytes and macrophages on antiviral genes, interferon stimulated gene-20 (ISG 20) and oligoadenylate synthetase (OAS-1). 3: To determine the effect of CART on HIV replication and persistence in astrocytes and macrophages.