The chemokine receptors CCR5 and CXCR4 serve, in addition to CD4, as coreceptors for human immunodeficiency virus-1 (HIV-1), and infection with HIV-1 can cause dementia. In brain-derived cells, HIV-1 envelope glycoprotein gp120 initiates a signaling cascade that involves p38 mitogen-activated protein kinase and leads to neuronal cell death. Using mixed neuronal/glial cultures from rats and mice genetically deficient in one or both HIV coreceptors, we show here that CCR5, CXCR4 or both can mediate HIV/ gp120 neurotoxicity depending on the viral strain. Paradoxically, we also found evidence for a CCR5-mediated neuroprotective pathway. We identify protein kinase Akt/PKB as an essential component of this pathway, which can be triggered by the CCR5 agonists macrophage inflammatory protein-1b and regulated-and-normal-T-cell-expressed-and-secreted. Moreover, these CCR5 ligands prevent neuronal cell death induced by stromal cell-derived factor-1, a CXCR4 agonist. Both neurons and glia coexpress CXCR4 and CCR5. Ca 2 þ imaging experiments demonstrate that engagement of CCR5 prevents CXCR4-triggered increases in intracellular free Ca 2 þ . This finding suggests that CCR5 ligands can protect neurons at least, in part, by modulating CXCR4-mediated toxicity through heterologous desensitization.

HIV-1 envelope protein gp120 has been implicated in neurotoxin production by monocytic cells, namely macrophages and microglia, and the pathogenesis of HIV-1 associated neurocognitive disorders (HAND). We previously showed in cerebrocortical cell cultures from rodents containing microglia, astrocytes and neurons, that overall inhibition of p38 MAPK signaling abrogated the neurotoxic effect of HIV-1 gp120. However, the time course of p38 MAPK activation and the contribution of this kinase in the various cell types remained unknown. In this study, we found that for HIV gp120-induced neurotoxicity to occur, active p38 MAPK is required in monocytic lineage cells, namely macrophages and microglia, and neuronal cells. In cerebrocortical cell cultures HIV-1 gp120 stimulated a time-dependent overall increase of active p38 MAPK and the activated kinase was primarily detected in microglia and neurons. Interestingly, both increased activation of p38 MAPK and neuronal death in response to gp120 were prevented by prior depletion of microglia, or in the presence of CCR5 ligand CCL4 or of p38 MAPK inhibitors. In human monocytic THP-1 cells and primary monocyte-derived macrophages (MDM), HIV gp120 stimulated production of neurotoxins was abrogated by prior introduction into the cells of a dominant-negative p38 MAPK mutant or p38 MAPK siRNA. In addition, the neurotoxic effects of cell-free supernatants from gp120-stimulated monocytic THP-1 cells were prevented in microglia-depleted cerebrocortical cells pretreated with a pharmacological inhibitor of p38 MAPK. Thus, p38 MAPK signaling was critical upon exposure to HIV gp120 for both the neurotoxic phenotype of monocytic cells and subsequent toxin-initiated neuronal apoptosis.

The innate immune system has been implicated in several neurodegenerative diseases, including human immunodeficiency virus (HIV)-1 associated dementia. Here we show that genetic ablation of CCR5 prevents microglial activation and neuronal damage in a transgenic model of HIV-associated brain injury induced by a CXCR4-utilizing viral envelope gp120. The CCR5 knockout (KO) also rescues spatial learning and memory in gp120-transgenic (tg) mice. However, the CCR5KO does not abrogate astrocytosis, indicating it can occur independently from neuronal injury and behavioral impairment. To further characterize the neuroprotective effect of CCR5-deficiency we performed a genome –wide gene expression analysis of brains from HIVgp120tg mice expressing or lacking CCR5 and non-transgenic controls. Comparison with a human brain microarray study reveals that brains of HIVgp120tg mice and HIV patients with neurocognitive impairment share numerous differentially regulated genes. Furthermore, brains of CCR5 wild-type (WT) and CCR5KO gp120tg mice express markers of an innate immune response. One of the most significantly up-regulated factors is the acute phase protein lipocalin-2 (LCN2). Using cerebrocortical cell cultures, we find that LCN2 is neurotoxic in a CCR5-dependent fashion while inhibition of CCR5 alone is not sufficient to abrogate neurotoxicity of a CXCR4-utilizing gp120. However, the combination of pharmacological CCR5 blockade and LCN2 protects neurons from toxicity of a CXCR4-utilizing gp120 thus recapitulating the finding in CCR5-deficient gp120tg mouse brain. Altogether, our study provides evidence for an indirect pathological role of CCR5 and a novel protective effect of LCN2 in combination with inhibition of CCR5 in HIV-associated brain injury.

Infection with human immunodeficiency virus-1 (HIV-1) often leads to HIV-associated neurocognitive disorders (HAND) prior to the progression to acquired immunodeficiency syndrome (AIDS). At the cellular level, mitogen-activated protein kinases (MAPK) provide a family of signal transducers that regulate many processes in response to extracellular stimuli and environmental stress, such as viral infection. Recently, evidence has accumulated suggesting that p38 MAPK plays crucial roles in various pathological processes associated with HIV infection, ranging from macrophage activation to neurotoxicity and impairment of neurogenesis to lymphocyte apoptosis. Thus, p38 MAPK, which has generally been linked to stress-related signal transduction, may be an important mediator in the development of AIDS and HAND.

BackgroundThe chemokine receptor CXCR4 (CD184) and its natural ligand CXCL12 contribute to many physiological processes, including decisions about cell death and survival in the central nervous system. In addition, CXCR4 is a co-receptor for human immunodeficiency virus (HIV)-1 and mediates the neurotoxicity of the viral envelope protein gp120. However, we previously observed that CXCL12 also causes toxicity in cerebrocortical neurons but the cellular mechanism remained incompletely defined.MethodsPrimary neuronal-glial cerebrocortical cell cultures from rat were exposed to a neurotoxicity-inducing CXCL12 concentration for different times and the activity of the stress-associated mitogen-activated protein kinase p38 (p38 MAPK) was assessed using an in vitro kinase assay. Neurotoxicity of CXCL12 and cellular localization of p38 MAPK was analyzed by immunofluorescence microscopy. Pharmacological inhibition of NMDA-type glutamate receptor-gated ion channels (NMDAR) of l-type Ca2+ channels was employed during 12- and 24-h exposure to neurotoxic amounts of CXCL12 to study the effects on active p38 MAPK and neuronal survival by Western blotting and microscopy, respectively. Neurotoxicity of CXCL12 was also assessed during pharmacological inhibition of p38 MAPK.ResultsHere, we show that a neurotoxic amount of CXCL12 triggers a significant increase of endogenous p38 MAPK activity in cerebrocortical cells. Immunofluorescence and Western blotting experiments with mixed neuronal-glial and neuron-depleted glial cerebrocortical cells revealed that the majority of active/phosphorylated p38 MAPK was located in neurons. Blockade of NMDAR-gated ion channels or l-type Ca2+ channels both abrogated an increase of active p38 MAPK and toxicity of CXCL12 in cerebrocortical neurons. Inhibition of l-type Ca2+ channels with nimodipine kept the active kinase at levels not significantly different from baseline while blocking NMDAR with MK-801 strongly reduced phosphorylated p38 MAPK below baseline. Finally, we confirmed that directly blocking p38 MAPK also abrogated neurotoxicity of CXCL12.ConclusionsOur findings link CXCL12-induced neuronal death to the regulation of NMDAR-gated ion channels and l-type Ca2+ channels upstream of p38 MAPK activation.

Infection with HIV-1 frequently affects the brain and causes NeuroAIDS prior to the development of overt AIDS. The HIV-1 envelope protein gp120 interacts with host CD4 and chemokine co-receptors to initiate infection of macrophages and lymphocytes. In addition, the virus or fragments of it, such as gp120, cause macrophages to produce neurotoxins and trigger neuronal injury and apoptosis. Moreover, the two major HIV co-receptors, the chemokine receptors CCR5 and CXCR4, serve numerous physiological functions and are widely expressed beyond immune cells, including cells in the brain. Therefore, HIV co-receptors are poised to play a direct and indirect part in the development of NeuroAIDS. Although rodents are not permissive to infection with wild type HIV-1, viral coreceptors - more than CD4 - are highly conserved between species, suggesting the animals can be suitable models for mechanistic studies addressing effects of receptor-ligand interaction other than infection. Of note, transgenic mice expressing HIV gp120 in the brain share several pathological hallmarks with NeuroAIDS brains. Against this background, we will discuss recently completed or initiated, ongoing studies that utilize HIV co-receptor knockout and viral gp120-transgenic mice as models for in vitro and in vivo experimentation in order to address the potential roles of HIV gp120 and its co-receptors in the development of NeuroAIDS.

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Pharmacy transitions-of-care services at the time of hospital discharge are helpful in reducing medication errors. Validated risk tools are commonly used by pharmacists to identify patients at greatest benefit of these services. However, current tools lack assessment of medication-related risk factors and predict hospital readmissions rather than medication errors. To address this, a novel medication-focused risk tool (UCSD-Rx risk score) was created to help classify patients at a higher risk for medication errors. This study was split into 2 phases aimed to internally validate the risk score. Phase I of the study compared the predictability of 30-day unplanned readmissions between the UCSD-Rx risk score and a well-validated risk tool, the LACE+ index. To further specify our risk score for pharmacist use, phase II of the study analyzed the predictability of the risk score to medication errors at discharge. Phase I demonstrated similar classification performance of 30-day unplanned readmissions between the UCSD-Rx risk score (C-statistic, 0.66; 95% confidence interval [CI], 0.64-0.68; P < .0001) and the LACE+ index (C-statistic, 0.69; 95%CI, 0.67-0.71; P < .0001). In phase II, logistic regression showed an increasing UCSD-Rx risk score was predictive of individuals who would experience a medication error at discharge (odds ratio, 1.068; 95%CI, 1.005-1.136; P = .035). Results of this study demonstrate that the UCSD-Rx risk score is a promising tool targeted for pharmacist use to identify patients that may benefit most from transitions-of-care services prior to discharge.