Our research is to understand the cellular mechanisms of inflammatory responses within the central nervous system (CNS), particularly as they relate to HIV-1 Associated Dementia (HAD). Chemokines are small molecular weight (7-14 kDa) proteins that mediate the recruitment and activation of leukocytes to sites of inflammation. Aberrant expression of various chemokines contributes to the pathogenesis of HAD, and astrocytes, the major glial cell in the CNS, are a prominent source of chemokines within the brain. Astrocytes express receptors for numerous chemokines, including CXCR4. CXCR4 is the exclusive receptor for the chemokine stromal cell-derived factor-I (SDF-l). Importantly, CXCR4 functions as a co-receptor with CD4 in the binding and fusion of T-cell tropic HIV1 strains, specifically by interacting with the viral protein gp120. Productive HIV-l infection in the CNS is restricted to infiltrating macrophages and resident microglia. Astrocyte infection by HIV-l is rare, thus, CXCR4 receptor expression on astrocytes may have other important functions. We have recently demonstrated that astrocytes express low levels of CXCR4, which are enhanced by the proinflammatory cytokines TNF-a and IL-1b. SDF-1a stimulation of astrocytes results in activation of the mitogen-activated protein kinase (MAPK) pathway, specifically that of ERK. Of most importance, SDF-1a treatment induced expression of three chemokines, MCP-1, IL-8 and IP-lO. Based on these findings, we hypothesize that SDF-1/CXCR4-mediated signaling pathways in astrocytes elicit production of chemokines, which can then further activate inflammatory responses in the CNS. The objectives of Aim 1 are to determine how TNF-alpha and IL-Ibeta enhance CXCR4 expression in human astrocytes by examining transcription of the CXCR4 gene and CXCR4 mRNA and protein expression. Functional analysis of the CXCR4 promoter has not been performed in astrocytes.
In Aim 2, the cis-acting regulatory elements responsive to TNF-alpha and IL-Ibetawill be identified using 5' deletion and site-specific mutants of the CXCR4 promoter.
In Aim 3, we will further our analysis in CXCR4 transfectants to delineate the signal cascades involved in SDF-la induction of chemokines, emphasizing the ERK MAPK pathway and the transcription factor NF-KB. Chemokine receptors are important targets for both anti-inflammatory and anti-viral agents, which will be of great benefit for HAD patients. The findings forthcoming from these studies will provide the first biological assessment of CXCR4 expression in the brain, thereby setting the foundation for future therapeutic manipulation of this critical chemokine receptor.
