Browsing by Subject "Astrocytes"
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Item Activated human astrocyte-derived extracellular vesicles modulate neuronal uptake, differentiation and firing(Informa UK Limited, trading as Taylor & Francis Group, 2019-12-26) You, Yang; Borgmann, Kathleen; Edara, Venkata Viswanadh; Stacy, Satomi; Ghorpade, Anuja; Ikezu, TsuneyaAstrocytes in the central nervous system (CNS) provide supportive neural functions and mediate inflammatory responses from microglia. Increasing evidence supports their critical roles in regulating brain homoeostasis in response to pro-inflammatory factors such as cytokines and pathogen/damage-associated molecular pattern molecules in infectious and neurodegenerative diseases. However, the underlying mechanisms of the trans-cellular communication are still unclear. Extracellular vesicles (EVs) can transfer a large diversity of molecules such as lipids, nucleic acids and proteins for cellular communications. The purpose of this study is to characterize the EVs cargo proteins derived from human primary astrocytes (ADEVs) under both physiological and pathophysiological conditions. ADEVs were isolated from human primary astrocytes after vehicle (CTL) or interleukin-1beta (IL-1beta) pre-treatment. Label-free quantitative proteomic profiling revealed a notable up-regulation of proteins including actin-associated molecules, integrins and major histocompatibility complex in IL-1beta-ADEVs compared to CTL-ADEVs, which were involved in cellular metabolism and organization, cellular communication and inflammatory response. When fluorescently labelled ADEVs were added into primary cultured mouse cortical neurons, we found a significantly increased neuronal uptake of IL-1beta-ADEVs compared to CTL-ADEVs. We further confirmed it is likely due to the enrichment of surface proteins in IL-1beta-ADEVs, as IL-1beta-ADEVs uptake by neurons was partially suppressed by a specific integrin inhibitor. Additionally, treatment of neurons with IL-1beta-ADEVs also reduced neurite outgrowth, branching and neuronal firing. These findings provide insight for the molecular mechanism of the ADEVs' effects on neural uptake, neural differentiation and maturation, and its alteration in inflammatory conditions.Item Astrocyte TIMP-1: Regulation and Gene Delivery in HAND(2018-12) Joshi, Chaitanya R.; Ghorpade, Anuja; Clark, Abbot F.; He, Johnny J.; Sumien, Nathalie; Rickards, Caroline A.Despite antiretroviral therapy, HIV-associated neurocognitive disorders (HAND) persist in 30-70% of patients. During HAND, elevated matrix metalloproteinases (MMPs) in the brain exacerbate the disease by blood-brain barrier breakdown, neuroinflammation and direct neurotoxicity. Tissue inhibitors of metalloproteinases (TIMPs) counter MMP activity. In the brain, TIMP-1 is primarily produced by astrocytes in response to injury or inflammation. However, TIMP-1 is downregulated during chronic inflammation in astrocytes and in HIV encephalitis brain tissues. We propose that restoring astrocyte TIMP-1 levels could mitigate neurodegeneration due to its MMP-inhibitory and -independent neuroprotective functions. HIV-1 non-productively infects astrocytes, which express viral proteins such as transactivator of transcription (Tat). As Tat mimics aspects of HAND by direct and indirect mechanisms, glial fibrillary acidic protein (GFAP) promoter-restricted Tat expressing (GT-Tg) mice were used to model HAND in our studies. Prolonged astrocyte Tat expression in GT-Tg mice resulted in HAND-relevant behavioral impairments characterized by higher anxiety, lower ambulation, impaired spatial learning, and memory. Importantly, behavioral deficits were accompanied by altered brain MMP/TIMP balance. Our data from GT-Tg mouse model confirmed neurocognitive decline and TIMP-1 dysregulation in the context of HAND. As TIMP-1 was downregulated with prolonged Tat expression in mice, we focused on replenishing TIMP-1 via gene delivery to the brain using cationic polymers. Polyethylenimine (PEI) is a highly efficient polymer for transfecting mammalian cells, however, high cytotoxicity restricts its use. Hence, PEI was modified using arginine (A) and stabilized with polyethylene glycol (P) to produce multiple AnPn analogues. AnPn analogues were biocompatible and successfully delivered reporter genes to primary neural cells. Select AnPn led to sustained reporter gene expression in human astrocytes and in mouse brains. In order to restrict gene expression to astrocytes, truncated GFAP promoters were used to drive gene expression. Subsequently, GFAP promoters were modified enhancing their activity and increasing gene expression. Lastly, successful polymer-mediated GFAP promoter-driven TIMP-1 gene delivery was demonstrated in human astrocytes. Overall, these findings enhance our understanding HIV-1 Tat-mediated TIMP-1 regulation, provide a novel therapeutic TIMP-1 gene delivery system, and pave the way for future investigations geared towards preclinical translation of TIMP-1-based HAND therapy.Item C1q propagates microglial activation and neurodegeneration in the visual axis following retinal ischemia/reperfusion injury(BioMed Central Ltd., 2016-03-24) Silverman, Sean M.; Kim, Byung-Jin; Howell, Garreth R.; Miller, Joselyn; John, Simon W. M.; Wordinger, Robert J.; Clark, Abbot F.BACKGROUND: C1q represents the initiating protein of the classical complement cascade, however recent findings indicate pathway independent roles such as developmental pruning of retinal ganglion cell (RGC) axons. Furthermore, chronic neuroinflammation, including increased expression of C1q and activation of microglia and astrocytes, appears to be a common finding among many neurodegenerative disease models. Here we compare the effects of a retinal ischemia/reperfusion (I/R) injury on glial activation and neurodegeneration in wild type (WT) and C1qa-deficient mice in the retina and superior colliculus (SC). Retinal I/R was induced in mice through elevation of intraocular pressure to 120 mmHg for 60 min followed by reperfusion. Glial cell activation and population changes were assessed using immunofluorescence. Neuroprotection was determined using histological measurements of retinal layer thickness, RGC counts, and visual function by flash electroretinography (ERG). RESULTS: Retinal I/R injury significantly upregulated C1q expression in the retina as early as 72 h and within 7 days in the superficial SC, and was sustained as long as 28 days. Accompanying increased C1q expression was activation of microglia and astrocytes as well as a significantly increased glial population density observed in the retina and SC. Microglial activation and changes in density were completely ablated in C1qa-deficient mice, interestingly however there was no effect on astrocytes. Furthermore, loss of C1qa significantly rescued I/R-induced loss of RGCs and protected against retinal layer thinning in comparison to WT mice. ERG assessment revealed early preservation of b-wave amplitude deficits from retinal I/R injury due to C1qa-deficiency that was lost by day 28. CONCLUSIONS: Our results for the first time demonstrate the spatiotemporal changes in the neuroinflammatory response following retinal I/R injury at both local and distal sites of injury. In addition, we have shown a role for C1q as a primary mediator of microglial activation and pathological damage. This suggests developmental mechanisms of C1q may be re-engaged during injury response, modulation of which may be beneficial for neuroprotection.Item Endothelin-1-Induced Signaling Involved in Extracellular Matrix Remodeling(2006-12-01) He, Shaoqing; Thomas Yorio; Neeraj Agarwal; Peter KoulenET-1-Induced Signaling in ECM Remodeling in Astrocytes. Shaoqing He, Department of Pharmacology & Neuroscience, University of North Texas Health Science Center, Fort Worth, TX 76107. ET-1 levels are elevated under pathophysiological conditions, including glaucoma, however, ET-1’s ocular functions are not fully documented. Therefore, ET-1-induced signaling and ECM remodeling in astrocytes and at the optic nerve head were determined in this study. Three signaling pathways, including ERK1/2, PKC, and P13 kinase, were involved in ET-1-medicated cell proliferation of U373MG astrocytoma cells. Blocking one of these pathways completely abolished cell proliferation. It appeared that ERK1/2 activation was involved, but was independent of PKC and P13 kinase activation by ET-1. It was also determined that the ETB receptor was the dominant receptor involved in ERK1/2 phosphorylation and cell proliferation. In addition, ERK1/2 phosphorylation was not transactivated by the EGF receptor by ET-1. The studies also indicated that there was no activation of c/nPKC, although PKC was involved in cell proliferation. In U373MG astrocytoma cells, MAPK-ERK, PKC and P13K pathways appear to exert their roles in parallel without a direct, apparent “cross-talk”. Based on the signaling pathways obtained from U373MG astrocytoma cells, the regulation of MMPs/TIMPs and fibronectin in ET-1-activated human optic nerve head astroctyes (hONAs) was also determined. ET-1 not only induced rapid phosphorylation of ERK1/2 and PKC βI/ βII/δ but also increased the activity of MMP-2 and the expression of TIMP=1 and 2. The activity of MMP-2 was enhanced in the presence of inhibitors of MAPK or PKC in hONAs, whereas the expression of TIMP-1 and 2 was abolished. ET-1 increased the soluble fibronectin (FN) expression as well as FN matrix formation, however, the expression and deposition of FN were MAPK- and PKC-independent, whereas expression and activity of MMps and TIMPs were MAPK- and PKC-dependent. Therefore, ET-1 shifted the balance of MMPs/TIMPs and substrates that altered the ECM composition and subsequently let to ECM remodeling in activated hONA cells. ET-1’s effects on ECM remodeling at the optic nerve head were also examined following intravitreal administration of ET-1 in rats. The increased expression of MMP-9 and collagen VI was detected in both ETB deficient rats and wildtype Wistar rats post ET-1 intravitreal injection for 2 and 14 days, whereas the deposition of FN and collagen IV was unchanged. There was no significant difference in staining of MMP-9 and collagen VI between ETB deficient rats and wildtype Wistar rats. In this study, ECM remodeling was demonstrated in rats injected with ET-1 into the vitreous. Such changes in the ECM seen in the current study provide additional insight into the mechanisms that might explain the glaucomatous changes observed in ET-1-injection or perfusion models. In summary, ET-1 not only activated several signaling pathways in cell proliferation of astrocytes, but also modulated the expression of ECM molecules in vitro and in vivo, indicating that ET-1 plays a regulatory role in ECM remodeling. These effects coupled with observations that ET-1 levels are elevated in glaucoma patients, suggests that ET-1 may be involved in glaucomatous optic neuropathy.Item HIV-1 TAT EXPRESSION ALTERS HUMAN ASTROCYTE INFLAMMATORY BIOMARKER PROFILES AND GLUTAMATE METABOLISM(2014-03) Joshi, Chaitanya R.; Ghorpade, AnujaPurpose (a): More than 50% of the human immunodeficiency virus type 1 (HIV-1) infected individuals exhibit some form of HIV-associated neurocognitive disorders (HAND). Several studies reported that HIV-1 transactivator of transcription (Tat) protein was associated with HAND pathophysiology. HIV-1 Tat induces apoptosis and dysregulates cytokine/chemokine profiles leading to neurotoxicity. Previous studies have studied the in vivo HIV-1 Tat regulation using transgenic animal models. Although animal models have helped determine the in vivo disease pathology, application of in vitro neural cell models will be critical to decipher cellular and molecular mechanisms associated with HAND. Here, we report an in vitro model system developed by transfecting human astrocytes with a full-length (101 AA) HIV-1 Tat protein expressing plasmid (pTat). HIV-1 Tat expressing in vitro system was used to evaluate HIV-1 Tat regulation of astrocyte inflammatory responses and altered neuroprotective function i.e. glutamate uptake from synapse. Methods (b): Primary human astrocytes were transfected with pTat by nucleofection and HIV-1 Tat expression was evaluated by immunocytochemistry. Effects of HIV-1 Tat on cell viability and replication were determined with metabolic activity and cell proliferation assays. Proinflammatory cytokines and chemokines were assayed using ELISAs. HIV-1 Tat regulation of glutamate clearing ability of astrocytes was determined using a modified amplex red glutamic acid/glutamate oxidase assay. Additionally, mRNA and protein expression of excitatory amino acid transporter-2 (EAAT-2), the major glutamate transporter in astrocytes, was measured by RT-PCR and western blot analysis respectively. Results (c): The immunostaining confirmed HIV-1 Tat expression in transfected astrocytes, while glial fibrillary acidic protein (GFAP) staining indicated morphological alterations. The pTat transfection did not significantly change cell metabolism as compared to controls. However, HIV-1 Tat expression altered chemokine and cytokine levels; specifically HIV-1 Tat increased CCL2 levels significantly (P. Conclusions (d): HIV-1 Tat expression upregulated inflammatory biomarkers and altered glutamate clearing ability of astrocytes, implicating a direct role for astrocyte-expressed HIV-1 Tat in HAND neuropathogenesis.Item METHAMPHETAMINE (METH) REGULATES ASTROCYTE EXCITATORY AMINO ACID TRANSPORTER-2 (EAAT-2) VIA ACTIVATION OF TRACE AMINE ASSOCIATED RECEPTOR (TAAR1) AND DOWNSTREAM CAMP SIGNALING(2014-03) Cisneros, Irma E.; Ghorpade, AnujaMethamphetamine (METH) abuse has prevailed as a drug epidemic within the United States and worldwide with an estimated 27.4 million users.Short-term side effect of METH abuse include a heightened libido and impaired judgment increasing the users chances of contracting Human immunodeficiency virus-1 (HIV-1). HIV-1 associated dementia (HAD), is the most severe manifestation of HIV-1-associated neurocognitive disorders and is an important neurological complication of HIV-1 infection characterized by cognitive, behavioral and motor dysfunction. Long-term METH users share characterized neurocognitive defects and disorders as HIV-1 infected individuals.Molecular outcomes of METH/HIV-1-induced neurotoxicity include Excitotoxicity, oxidative stress, glial cell activation, inflammation, and hyperthermia. Astrogliosis is a key pathological feature of METH exposure and HAD, however, the molecular mechanisms remain unclear. The current studies investigate METH-induced TAAR1 activation and will uncover molecular mechanisms associated with glutamate transporter (EAAT-2) dysregulation. Purpose (a): Glutamate is an excitatory neurotransmitter that is highly regulated in the central nervous system (CNS). High concentrations of extracellular glutamate result in excitotoxicity and can exacerbate neurodegenerative disorders, including human immunodeficiency virus-1 (HIV-1)-associated neurocognitive disorders (HAND). Additionally, drugs of abuse such as methamphetamine (METH) can increase the severity of excitotoxicity and can accelerate HAND. Excitatory amino acid transporter-2 (EAAT-2) is responsible for approximately 90% of extracellular glutamate uptake from the synapse and is primarily localized in astrocytes. Dysregulation of EAAT-2 leads to astrocytes decreased ability to clear glutamate. Methods (b): It is established that METH leads to excitotoxicity in neurons, however, in astrocytes the molecular mechanisms resulting in METH-mediated EAAT-2 dysregulation are unclear. Previously we showed that HIV-1ADA, METH and transient hyperthermia regulates localization and expression of astrocyte trace amine associated receptor 1 (TAAR1). Results (c): Our data shows METH-induced activation of astrocyte TAAR1 increases intracellular cAMP levels in astrocytes that is significantly decreased in siTAAR1-transfected astrocytes. Further, METH treatment downregulates EAAT-2 mRNA levels. We propose downstream cAMP signaling pathways of METH-induced astrocyte TAAR1 activation result in EAAT-2 dysregulation. Conclusions (d): The results of this study will uncover novel molecular mechanism of METH-induced astrocyte TAAR1 activation and the downstream effects of cAMP signaling on astrocyte EAAT-2 levels in the context of HAND.Item METHYLENE BLUE PROTECTS ASTROCYTES FROM HYPOXIA-REOXYGENATION INJURY BY IMPROVING CELLULAR BIOENERGETICS(2014-03) Roy Choudhury, Gourav; Winters, Ali; Rich, Ryan; Ryou, Myoung-Gwi; Gryczynski, Zygmunt; Yang, ShaohuaIn ischemic stroke, which is caused by a clot that blocks a brain blood vessel, a portion of the brain is deprived of oxygen and nutrients that are carried in the blood. When this clot is removed either by time or drugs, the affected region is flooded with an overabundance of nutrients and oxygen to a degree that is larger than the cells in the region can process and this unbalances the system and causes further damage. In our current study, we are testing a chemical compound called Methylene Blue to see if it can improve the cells’ ability to handle this excess of nutrients and oxygen available to them at the time of clot dissolution and thereby improve their survival after ischemic stroke. Purpose (a): Ischemic Stroke inflicts a double blow to the affected brain region by characteristically presenting a period of acute ischemia during which the cells are completely deprived of valuable nutrients (Glucose & oxygen) resulting in cell death, however as the blood flow is restored (Spontaneously/surgically) the surviving cells are exposed to an overwhelming levels of glucose and oxygen resulting in reperfusion injury which further aggravates the cellular injury inflicted by ischemia. Methylene Blue (MB) is a heterocyclic aromatic compound shown to function as an alternative electron carrier and improve glucose uptake, cerebral blood flow (CBF), and cerebral metabolic rate of oxygen in the brain. In our current study we aim to delineate if MB is protective in astrocyte against hypoxia-reoxygenation injury and determine its underlying mechanism. Methods (b): Primary astrocytes cultures isolated from day old C57BL6 were used in the current study. Protective role of Methylene blue (MB) in primary astrocyte cultures was evaluated in an in vitro model of cellular hypoxia (0.1% O2, 6h) and re-oxygenation (24h). The effect of MB on glucose uptake was determined by using the 2-NBDG assay. Oxygen sensitive dye Tris (2,2′-bipyridyl) dichloro Ruthenium(II) hexahydrate and Fluorescence Life Time Imaging (FLTI) was used to determine the effect of MB on intracellular oxygen concentration. ATP assay was used to determine the effect of MB on cellular energy status. Results (c): Results from cell viability assay showed that MB treatment significantly protected astrocytes from hypoxia-reoxygenation induced cell death. MB treatment significantly increased cellular glucose uptake in primary astrocyte cultures. FLTI showed that MB significantly increased intracellular oxygen concentration in primary astrocytes. Astrocytes treated with MB also had significantly higher ATP concentration compared to non-treated cells. Conclusions (d): Methylene Blue protects astrocytes against hypoxia-reoxygenation injury by improving astrocyte bioenergetics.Item Modulation of Astrocyte Phenotype in Response to T-cell Interaction(2021-05) Hersh, Jessica M.; Yang, Shaohua; Smith, Michael L.; Jin, Kunlin; Hodge, Lisa M.We determined that T-cell astrocyte interaction modulates interleukin-10 (IL-10) production from both cell types. The impact of IL-10 on astrocytes was compared to IL-10 generated from T-cell-astrocyte interactions in vitro. We demonstrated that T-cells directly interact with astrocytes to upregulate gene expression and secretion of IL-10, confirmed by elevated STAT3p/STAT3 expression in astrocytes. IL-10 increased astrocytes proliferation. In addition, IL-10 treatment and CD4+ co-culture shifts primary astrocytes toward a more energetic phenotype. These findings indicate that direct interaction of CD4+ T-cells with astrocytes, activated the IL-10 anti-inflammatory pathway, altering astrocyte phenotype, metabolism, and proliferation.Item Post-transcriptional Regulation of Astrocyte-Tissue Inhibitor of Metalloproteinases-1 (TIMP-1) In HIV-Associated Neurocognitive Disorders(2015-12-01) Thete, Mayuri V.; Ghorpade, Anuja; Mathew, Porunelloor A.; Clark, Abbot F.HIV-1 leads to several central nervous system (CNS) problems termed as HIV-associated neurocognitive disorders (HAND). Tissue inhibitor of metallopoteinases-1 (TIMP-1) / matrix metalloproteinase (MMP) imbalance has been observed in HAND and several other neuroinflammatory conditions. Astrocytes are major contributors to brain TIMPs, and they regulate TIMP / MMP balance [1, 2]. Differential regulation of TIMP-1 in acute versus chronic neuroinflammation is relevant to HAND neuropathogenesis and long-term neurodegeneration[3]. However, the underlying mechanisms are still being uncovered. Our previous work has shown the neuroprotective role of TIMP-1 via MMPdependent and independent manners [4, 5]. In this study, we investigated regulation of astrocyte TIMP-1 in HAND. First, microarray analyses were performed to analyze micro RNA (miRNA) changes in IL-1β activated astrocytes. For further studies, TIMP-1 3’ untranslated region (TIMP-1 3’UTR) cloned downstream of firefly luciferase and miRNA overexpression constructs were used to investigate miRNA-mediated TIMP-1 3’UTR post-transcriptional regulation. Firefly luciferase activity and endogenous astrocyte TIMP-1 levels were measured in parallel experiments. A total of 12 miRNAs were significantly increased and four were significantly decreased; seven of those were further confirmed by real-time PCR (RT2-PCR). The most increased were miR 155, miR 146b and miR 29b, whereas, the most downregulated one was miR 518e. Overexpression of miR 155, miR 146b, miR 29b altered both firefly luciferase activity via and endogenous astrocyte TIMP-1 levels in IL-1β and/or HIV-1 activated astrocytes. Comparable responses were obtained in luciferase activity changes and de novo TIMP- 1 protein levels corroborating post-transcriptional regulation of astrocyte-TIMP-1. Thus, our data suggests that astrocyte-TIMP-1 is regulated post-transcriptionally by miRNAs during HAND. Given the emerging role of miRNAs in several neuroinflammatory and neurodegenerative diseases, our data uncover a novel mechanism of TIMP-1 regulation in astrocytes that may have significant impact on future studies on MMP/TIMP balance in HIV-associated neuroinflammation.Item TAAR1-dependent astrocyte dysregulation during HAND and METH exposure(2018-12) Mythen, Shannon; Ghorpade, Anuja; Basu, Alakananda; Barber, Robert C.; Gatch, Michael B.; Krishnamoorthy, Raghu R.Excitatory amino-acid transporter (EAAT)-2 is predominantly expressed in astrocytes and clears glutamate from tripartite synapses preventing excitotoxicity. EAAT- 2 dysregulation occurs during human immunodeficiency virus (HIV)-associated neuroinflammation and methamphetamine (METH) abuse, leading to neurotoxic outcomes. Trace amine associated receptor (TAAR) 1, a METH receptor in astrocytes, triggers EAAT-2 dysfunction. Protein kinase C (PKC) signaling promotes ubiquitination of EAAT-2 C-terminal lysine residues, resulting in EAAT-2 internalization. As a G protein coupled receptor, TAAR1's signaling is implicated in PKC activation. In this work, we investigated the role of TAAR1 in PKC-mediated EAAT-2 ubiquitination during HIV- associated neurocognitive disorders (HAND) and METH comorbidities. We evaluated a TAAR1 overexpression model in primary astrocytes to elucidate TAAR1-mediated functional changes. We found that TAAR1-selective inhibitor, EPPTB, reduced EAAT-2 ubiquitination, and a PKC activator decreased glutamate clearance in METH-pretreated human astrocytes. Therapies targeting astrocyte dysfunction may improve outcomes during HAND, METH abuse and other neuroinflammatory disorders.Item Transcriptome and functional profiles of R/G-HIV+ human astrocytes: Implications for shock or lock therapies in the brain(2020-05) Edara, Venkata Viswanadh; Borgmann, Kathleen; Berg, Rance E.; Krishnamoorthy, Raghu R.; Mathew, Porunelloor A.; Yang, Shaohua; Patterson, Rita M.A significant number of people living with human immunodeficiency virus (HIV) suffer from HIV-associated neurocognitive disorders (HAND). Many previous studies investigating HIV in astrocytes as a heterogenous population have established the relevance of astrocytes to HIV-associated neuropathogenesis. However, these studies were unable to differentiate the state of infection, i.e. active or restricted, or to evaluate how this affects astrocyte biology. In this study a pseudotyped doubly labelled fluorescent reporter R/G-HIV-1 was used to identify and enrich restricted and active populations of HIV+ astrocytes based on the viral promoter activity. Here we report, the majority of human astrocytes restricted R/G-HIV-1 gene expression early during infection and were resistant to reactivation by vorinostat and interleukin-1β. However, actively infected astrocytes were inducible, leading to increased expression of viral proteins upon reactivation. R/G-HIV-1 infection also significantly decreased cell proliferation and glutamate clearance ability of astrocytes, which may contribute to excitotoxicity. Moreover, transcriptome analyses to compare gene expression patterns of astrocytes harboring active vs restricted long terminal repeats revealed that the gene expression patterns were similar, and the active population demonstrated more widespread and robust changes. Our data suggest that harboring the HIV genome profoundly alters astrocyte biology and strategies that keep the virus latent (e.g. Block and Lock), or those that reactivate the latent virus (e.g. Shock and Kill) may be detrimental to astrocyte function and possibly augment their deleterious contributions to HAND.Item Uncovering the Multifaceted HAND of Reactive astrocytes in HIV-Associated Neurocognitive Disorders(2017-08-01) Nooka, Shruthi; Ghorpade, Anuja; Luedtke, Robert; Mathew, Porunelloor A.Despite the advent of antiretroviral therapy (ART), central nervous system (CNS) complications associated with HIV-1 infection, collectively referred to as HIV-associated neurocognitive disorders (HAND), continue to increase. HIV infection promotes cognitive dysfunction and neurodegeneration through persistent inflammation, oxidative stress from infected and/or activated macrophages, astrocytes and neurons. Additionally, recent studies demonstrated that neurotoxic side effects of antiretroviral (ARV) drugs are among several contributing factors to this continued prevalence of HAND. In recent years, a new appreciation of the role of astrocytes in regulating HIV-1 CNS infection has emerged. Thus, investigating the elusive cellular and molecular mechanisms regulated by astrocytes during HIV-1/ART-induced neurotoxicity could provide insight into HAND pathogenesis. The work presented in this thesis contributes to the documentation of astrocyte dysfunction in HIV-1-infection. The accumulated data reveal the multifaceted mechanisms and roles of astrocytes in HIV-1 CNS infection. A neuropathological feature of HIV-1 infection includes reactive astrogliosis, which is a hallmark of many neurodegenerative diseases. Our studies reveal the regulation of β-catenin signaling on major aspects of reactive astrogliosis i.e., proliferation, wound healing and inflammation. HAND-relevant inflammatory stimuli activate β-catenin signaling in astrocytes. Our in vitro studies reveal knockdown of β-catenin impairs astrocyte responses to injury. Further, reduced levels of β-catenin also show less proliferation and inflammatory responses in astrocytes. We also demonstrated that Wnt/ β-catenin and NF-κB crosstalk links with inflammation during HIV-1 CNS infection. We next investigated endoplasmic reticulum (ER) stress associated with HAND-relevant neuroinflammation. Our studies show that ART (abacavir) and interleukin-1β increase cytosolic calcium in astrocytes, which in turn regulates ER stress and mitochondrial depolarization. We also identify astrocyte elevated gene (AEG)-1 as an ER stress inducible gene. In addition, AEG-1 interacts with calnexin, which emphasizes AEG-1 as a scaffolding protein regulating ER calcium signaling. Further, HIV-1-coupled inflammation and oxidative stress significantly increase regulator of ribosome synthesis (RRS1) expression, suggesting inhibition of rRNA transcription in astrocytes. Further, AEG-1 overexpression enhanced oxidative stress-induced RRS1 expression, i.e., nucleolar stress in astrocytes. Taken together, this study identified novel regulatory mechanisms in reactive astrocytes during HIV-1-induced neurodegeneration that might serve as innovative therapeutic targets for HAND.