inflammatory mediators and viral products produced by HIV-infected microglia and astrocytes perturb the function and viability of adjacent uninfected neuronal and glial cells and contribute to the pathogenesis of HIV-associated neurocognitive disorders (HAND). We hypothesized that HIV-infected microglia and astrocytes display increased sensitivity to the proinflammatory effects of the increased levels of serum LPS detected in HIV-infected individuals which may contribute to the development of HAND. We tested this hypothesis using our JR-CSF/hu-cycT1 mouse model that is transgenic for an integrated full-length nfectious HIV-1 provirus and the human cyclin T1 gene under the control of a CD4 promoter, and demonstrated that in vivo administered LPS more potently activated JR-CSF/hu-cycT1 mouse microglia and astrocytes as compared to control littermate mouse microglia and astrocytes. The expanding role of buprenorphine for the treatment of opiate addiction in the population of HIV-infected intravenous drug users increases the importance of determining the capacity of buprenorphine to synergize with HIV-induced pathways to promote HIV neurocognitive decline. To delineate the mechanistic basis of our finding about the synergistic effects of HIV and LPS on microglial and astrocyte activation and to address the possible contributory impact of buprenorphine, we propose to analyze in a defined biological context (? HIV, ? inflammatory stimulus (LPS), ? buprenorphine) the effects of HIV, inflammatory stimuli and buprenorphine on the brain proteome profile. Quantitative brain proteomics will be carried out using stable isotope labeling of the animals (SILAM) fed with N15 or natural labeled chow. Targeted analysis of high pH carbonate stripped membranes from the brain will be performed. This will provide insights into the changes in expression or modification of receptors and transporters induced by HIV infection, HIV-associated inflammation and buprenorphine. Changes in tyrosine phosphorylation will be studied using immobilized anti-phosphotyrosine to isolate tyrosine phosphorylated proteins and peptides. Samples will be archived for future analysis of: brain region cytosol, neuropeptides, peptides that are serine/threonine phosphorylated, serum/plasma, and other tissues. Overall, these data will provide a detailed study of the proteome of the HIV-infected brain in response to LPPS and buprenorphine which is not feasible in human subjects. These data will provide correlative insights for biomarkers of neurocognitive changes as well as buprenorphine use and may suggest new pathways by which HIV and opiates interact in the development of HAND.
Showing the most recent 10 out of 17 publications
