Regulation of Astrocyte TIMP-1 in HIV-Associated Dementia

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DESCRIPTION (provided by applicant): The goal of this proposal is to characterize regulation of primary human astrocyte tissue inhibitors of metalloproteases (TIMP) expression during Human Immunodeficiency Virus (HIV)-associated dementia (HAD). HIV-associated neurocognitive disorders are a group of disorders of which HAD is the most severe. HAD affects 10-27% of HIV-infected individuals in the United States. The disease is characterized by an accumulation of activated and infected macrophages/microglia in the central nervous system, which secrete cytokines, infectious virions and viral proteins that activate surrounding astrocytes leading to astrogliosis and altered gene expression. Neural dysfunction is believed to be the key component causing dementia. Astrocytes are essential in maintaining neural homeostasis in the CNS during injury and repair, in part through the production of neuroprotective TIMPs. An imbalance between matrix metalloproteases and TIMPs is thought to contribute to several neurodegenerative disorders, including HAD. TIMP-1, the inducible form of a family of 4 TIMPs, is a multifunctional protein that displays neuroprotective properties via homeostatic maintenance and possibly by inhibiting apoptosis. TIMP-1 levels are decreased in HAD patients compared to HIV-1 seronegative controls and primary human astrocytes acutely stimulated with HAD-relevant stimuli increase TIMP-1 expression, but levels fall after 3 days of exposure. Additionally, deleting the CAAT site at -310 in the TIMP-1 promoter increases transcription. Thus, we propose that down regulation of astrocyte TIMP-1 during HAD is mediated through differential transcriptional regulation via a CAAT site in the TIMP-1 promoter leading to reduced astrocyte TIMP-1 expression and ultimately exacerbating neurodegeneration. Two specific aims have been designed to address regulation of astrocyte TIMP-1 expression during HAD: Aim 1 determine the expression profile of CAAT enhancer binding protein 2 in activated primary human astrocytes and their influence on TIMP-1 transcription and Aim 2 identify signal transduction pathways leading to TIMP-1 down regulation in primary human astrocytes. To complete Aim 1 basic cell culture of primary human astrocytes and HAD-relevant stimuli like cytokines and viral proteins will be used along with analysis tools such as western blot, real-time polymerase chain reaction, immunocytochemistry and enzyme linked immunosorbent assay. Aim 2 will be completed by combining pathway-specific inhibitors with a panel of luciferase expression plasmids, CAAT-binding factor overexpression plasmids and pathway-indicating plasmids with and without HAD-relevant stimuli. Completion of these studies will shed light on the regulation of human astrocyte TIMP-1 expression in HAD. PUBLIC HEALTH RELEVANCE: HIV-1-associated dementia (HAD), the most severe manifestation of HIV-1-associated neurocognitive disorders, is an important neurological complication of HIV-1 infection and is characterized by cognitive, behavioral and motor dysfunction. An estimated 10-27% of HIV-seropositive patients progress to develop HAD in developed worlds such as the United States, despite the availability of highly active antiretroviral therapy. Our studies will provide a better understanding of the specific mechanistic contributions of activated astrocytes to HIV-1-neuropathogensis and neuroinflammation.