Raghava Potula

Methamphetamine disrupts blood-brain barrier function by induction of oxidative stress in brain endothelial cells.

Methamphetamine (METH), a potent stimulant with strong euphoric properties, has a high abuse liability and long-lasting neurotoxic effects. Recent studies in animal models have indicated that METH can induce impairment of the blood–brain barrier (BBB), thus suggesting that some of the neurotoxic effects resulting from METH abuse could be the outcome of barrier disruption. In this study, we provide evidence that METH alters BBB function through direct effects on endothelial cells and explore possible underlying mechanisms leading to endothelial injury. We report that METH increases BBB permeability in vivo, and exposure of primary human brain microvascular endothelial cells (BMVEC) to METH diminishes the tightness of BMVEC monolayers in a dose- and time-dependent manner by decreasing the expression of cell membrane-associated tight junction (TJ) proteins. These changes were accompanied by the enhanced production of reactive oxygen species, increased monocyte migration across METH-treated endothelial monolayers, and activation of myosin light chain kinase (MLCK) in BMVEC. Antioxidant treatment attenuated or completely reversed all tested aspects of METH-induced BBB dysfunction. Our data suggest that BBB injury is caused by METH-mediated oxidative stress, which activates MLCK and negatively affects the TJ complex. These observations provide a basis for antioxidant protection against brain endothelial injury caused by METH exposure.

HIV-1 infection and alcohol abuse: neurocognitive impairment, mechanisms of neurodegeneration and therapeutic interventions.

Clinical studies indicate that alcohol dependence has an additive effect on cognitive deficits associated with HIV-1 infection. Findings in humans and animal models suggest that alcohol, similar to HIV-1, induces inflammatory processes in the brain leading to neurodegeneration. The causes of HIV-1-associated neurotoxicity are comparable to those mediating alcohol-induced neuronal injury. This review aims to present the mechanisms of the combined effects of HIV-1 and alcohol abuse in the brain and to discuss neuroprotective therapies. Oxidative stress, overproduction of pro-inflammatory factors, impairment of blood-brain barrier and glutamate associated neurotoxicity appear to play important roles in alcohol driven neurodegeneration. Diminution of neuroinflammation constitutes a logical approach for prevention of HIV-1 and alcohol mediated neurodegeneration. Agonists of cannabinoid receptor 2 (CB₂) possess potent anti-inflammatory and neuroprotective properties. We address multifaceted beneficial effects of CB₂ activation in the setting of HIV-1 brain infection and alcohol abuse.

Dysfunction of brain pericytes in chronic neuroinflammation.

Brain pericytes are uniquely positioned within the neurovascular unit to provide support to blood brain barrier (BBB) maintenance. Neurologic conditions, such as HIV-1-associated neurocognitive disorder, are associated with BBB compromise due to chronic inflammation. Little is known about pericyte dysfunction during HIV-1 infection. We found decreased expression of pericyte markers in human brains from HIV-1-infected patients (even those on antiretroviral therapy). Using primary human brain pericytes, we assessed expression of pericyte markers (α1-integrin, α-smooth muscle actin, platelet-derived growth factor-B receptor β, CX-43) and found their downregulation after treatment with tumor necrosis factor-α (TNFα) or interleukin-1 β (IL-1β). Pericyte exposure to virus or cytokines resulted in decreased secretion of factors promoting BBB formation (angiopoietin-1, transforming growth factor-β1) and mRNA for basement membrane components. TNFα and IL-1β enhanced expression of adhesion molecules in pericytes paralleling increased monocyte adhesion to pericytes. Monocyte migration across BBB models composed of human brain endothelial cells and pericytes demonstrated a diminished rate in baseline migration compared to constructs composed only of brain endothelial cells. However, exposure to the relevant chemokine, CCL2, enhanced the magnitude of monocyte migration when compared to BBB models composed of brain endothelial cells only. These data suggest an important role of pericytes in BBB regulation in neuroinflammation.

Peroxisome proliferator-activated receptor-γ activation suppresses Hiv-1 replication in an animal model of encephalitis

Objective:Poor penetration of antiretroviral therapy across the blood–brain barrier poses an impediment on control of HIV-1 infection in brain macrophages. Peroxisome proliferator-activated receptor (PPAR)-γ, a member of the nuclear receptors family, regulates important physiological functions (including anti-inflammatory effects) in response to ligand-mediated activation. As PPARγ agonists are rapidly absorbed by oral administration and efficiently permeate the blood–brain barrier, we hypothesized that PPARγ stimulation may suppress HIV-1 replication. Design and methods:We investigated the effect of PPARγ ligand (rosiglitazone) on HIV-1 replication in human monocyte-derived macrophages and in vivo using a murine model (immunodeficient mice reconstituted with human lymphocytes and intracerebrally inoculated with HIV-1 infected macrophages) of HIV-1 encephalitis. Results:Treatment with rosiglitazone caused a significant decrease of virus infection in macrophages. PPARγ stimulation inhibited virus replication by modulating NF-κB activation in a receptor-dependent manner, leading to downregulation of HIV-1 long terminal repeat (LTR) promoter activity and suppression of HIV-1 replication. These effects were PPARγ specific as PPARγ silencing or addition of PPARγ antagonist abolished effects of PPARγ stimulation on HIV-1 LTR and virus replication. Using a murine model for HIV-1 encephalitis, we demonstrated that PPARγ ligand suppressed HIV-1 replication in macrophages in brain tissue and reduced viremia by 50%. Conclusion:In vitro data delineated the novel mechanism by which PPARγ activation suppresses HIV-1 replication, and in vivo findings underscored the ability of PPARγ agonists to reduce HIV-1 replication in lymphocytes and brain macrophages, thus offering a new therapeutic intervention in brain and systemic infection.

Methamphetamine induces trace amine- associated receptor 1 (TAAR1) expression in human T lymphocytes: role in immunomodulation.

The novel transmembrane G protein‐coupled receptor, trace amine‐associated receptor 1 (TAAR1), represents a potential, direct target for drugs of abuse and monoaminergic compounds, including amphetamines. For the first time, our studies have illustrated that there is an induction of TAAR1 mRNA expression in resting T lymphocytes in response to methamphetamine. Methamphetamine treatment for 6 h significantly increased TAAR1 mRNA expression (P < 0.001) and protein expression (P < 0.01) at 24 h. With the use of TAAR1 gene silencing, we demonstrate that methamphetamine‐induced cAMP, a classic response to methamphetamine stimulation, is regulated via TAAR1. We also show by TAAR1 knockdown that the down‐regulation of IL‐2 in T cells by methamphetamine, which we reported earlier, is indeed regulated by TAAR1. Our results also show the presence of TAAR1 in human lymph nodes from HIV‐1‐infected patients, with or without a history of methamphetamine abuse. TAAR1 expression on lymphocytes was largely in the paracortical lymphoid area of the lymph nodes with enhanced expression in lymph nodes of HIV‐1‐infected methamphetamine abusers rather than infected‐only subjects. In vitro analysis of HIV‐1 infection of human PBMCs revealed increased TAAR1 expression in the presence of methamphetamine. In summary, the ability of methamphetamine to activate trace TAAR1 in vitro and to regulate important T cell functions, such as cAMP activation and IL‐2 production; the expression of TAAR1 in T lymphocytes in peripheral lymphoid organs, such as lymph nodes; and our in vitro HIV‐1 infection model in PBMCs suggests that TAAR1 may play an important role in methamphetamine ‐mediated immune‐modulatory responses.

Dysregulation of Claudin-5 in HIV-induced Interstitial Pneumonitis and Lung Vascular Injury. Protective Role of Peroxisome Proliferator–activated Receptor-γ

RATIONALE HIV-1-induced interstitial pneumonitis (IP) is a serious complication of HIV-1 infection, characterized by inflammation and cellular infiltration in lungs, often leading to respiratory failure and death. The barrier function of the pulmonary endothelium is caused in part by tight junction (TJ) proteins, such as claudin-5. Peroxisome proliferator-activated receptor (PPAR)-γ is expressed in lung tissues and regulates inflammation. We hypothesize that HIV-1 induces vascular lung injury, and HIV-1-mediated damage of the pulmonary endothelium and IP is associated with dysregulation of PPAR-γ. OBJECTIVES Investigate the effects of HIV-1 infection on the pulmonary microvasculature and the modulatory effects of the PPAR-γ ligands. METHODS Using human lung tissues, we demonstrated down-regulation of claudin-5 (marker of pulmonary barrier integrity), down-regulation of PPAR-γ transcription, and expression in lung tissues of HIV-1-infected humans with IP. MEASUREMENTS AND MAIN RESULTS Human lung microvascular endothelial cells expressed the TJ proteins claudin-5, ZO-1, and ZO-2; HIV-1 decreased TJ proteins expression and induced nuclear factor-κB promoter activity, which was reversed by PPAR-γ agonist. Using two murine HIV/AIDS models, we demonstrated decreased claudin-5 expression and increased macrophage infiltration in the lungs of HIV-1-infected animals. Activation of PPAR-γ prevented HIV-1-induced claudin-5 down-regulation and significantly reduced viremia and pulmonary macrophage infiltration. CONCLUSIONS HIV-induced IP is associated with injury to the lung vascular endothelium, with decreased TJ and PPAR-γ expression, and increased pulmonary macrophage infiltration. PPAR-γ ligands abrogated these effects. Thus, regulation of PPAR-γ can be a therapeutic approach against HIV-1-induced vascular damage and IP in infected humans. Removal of Expression of Concern: Issues leading to the previous expression of concern for this article have been resolved after further revisions and editorial review. No further concerns exist.

Comparative Evaluation of the BD Phoenix Yeast ID Panel and Remel RapID Yeast Plus System for Yeast Identification

Becton Dickinson Phoenix Yeast ID Panel was compared to the Remel RapID Yeast Plus System using 150 recent clinical yeast isolates and the API 20C AUX system to resolve discrepant results. The concordance rate between the Yeast ID Panel and the RapID Yeast Plus System (without arbitration) was 93.3% with 97.3% (146/150) and 95.3% (143/150) of the isolates correctly identified by the Becton Dickinson Phoenix and the Remel RapID, respectively, with arbitration.

Alcohol and Neurodegeneration

Strong association between alcohol abuse and an increase in both systemic and brain levels is evident from both humans and animal models. This chapter comprehensively reviews the interplay between progression of neuroinflammation and neurological disorders initiated by alcohol abuse. Recent observations of excitotoxicity associated with excessive neurotransmitter release, oxidative stress leading to free radical damage, and cell death through an enhanced inflammatory response provide important clues to the mechanisms that could mediate alcohol’s toxic effects on brain cells. Chronic alcoholics have the temporal hallmark of neurocognitive deficits, neuronal injury, and neurodegeneration. Studies suggest that the initiation and progression of alcohol-mediated neurodegeneration is driven in part by release of pro-inflammatory factors from activated microglia, oxidative stress, impairment of blood–brain barrier (BBB), and glutamate-associated neurotoxicity. Recent observation of strong associations between cannabinoid systems within the central nervous system in regulating neuroinflammation via the cannabinoid receptor 2 highlights the importance of this pathway in alcohol-driven neuroinflammation. Regulatory mechanisms that regulate alcohol-induced neuroinflammation, oxidative neuronal injury, and altered BBB are examined, as well as modalities to ameliorate these processes are discussed.

Alcohol Abuse, HIV-1, and Hepatitis C Infection

Alcohol is the most commonly used and abused drug in the United States. Approximately 14 million Americans meet the criteria for alcohol abuse or dependence [1]. Alcohol abuse significantly affects morbidity and mortality from infectious diseases [2] and contributes to 85,000 deaths each year in the United States [3]. Alcohol use is common among HIV-infected individuals, and its impact on HIV disease progression has been suggested. Since alcohol use is widespread and it is often heavy among HIV-infected individuals and among the most sexually active age groups at risk of HIV infection, it has been of a great interest to investigate the role of alcohol abuse in promoting HIV transmission and infection. It was suggested that acute and chronic alcohol use might increase host susceptibility to HIV infection [4, 5]. Heavy alcohol consumption was strongly associated with incidence of HIV infection [6]. Alcohol use and abuse have been identified as potential behavioral risk factors for the transmission of HIV, in the form of drinking before risky sexual events or frequent binge drinking as associated with HIV incidence [7–9]. Alcohol consumption, particularly at high levels, has a negative impact on adherence to medications in general [10] and specifically to combined antiretroviral therapy, cART [11–14]. Chander et al. found that hazardous levels of alcohol use were associated with decreased cART utilization, adherence, and viral suppression, independent of active drug use. Combined alcohol and drug use was associated with lower odds of adherence and viral suppression than either drugs or alcohol alone [15].