Cannabinoids are established as being immunomodulatory, however, the involvement of the cannabinoid receptors in specific aspects of immune function has been demonstrated in few studies. Many effects of cannabinoids are mediated by two G-protein coupled receptors, designated CB1 & CB2. Most modern studies on immunomodulatory effects of cannabinoids have been focused on CB2 receptor, since it is primarily expressed on immune cells. We have shown that CB1 receptor is expressed in microglia that also express MHC class II antigen. The regulation of class II MHC genes occur primarily at the transcriptional level, and a non-DNA-binding protein, class II transactivator (CIITA), has been shown to be the master activator for class II transcription. Our recent data indicate that cannabinoids affect MHC class II expression through actions on CIITA at the transcriptional level. The CIITA gene is known to be controlled by multiple promoters. Interferon (IFN)- g -inducible expression of CIITA is primarily regulated by promoter IV which contains three binding sites, an IFN-g activation sequence, an E Box and an IFN regulatory factor site that bind the transcription factors STAT-la, USF-1 and IRF-1 & IRF-2 respectively. Synergistic activation of the promoter by STAT-1, USF-1 and IRF-1 and IRF-2 leads to expression of CIITA, which then activates transcription of class II MHC genes. The present proposal will elucidate the mechanism by which the cannabinoids mediate the expression of CIITA in human microglia. To address our goal, we will isolate and study primary cultures of microglial cells from human brain. Cells activated with IFN-g will be exposed to cannabinoid agonists and antagonists to determine the effect of cannabinoid receptor activation on stimulations of CIITA mRNA and protein levels.
The specific aims of the proposal are 1) Determine whether cannabinoid effects on CIITA are receptor mediated and 2) Determine whether the binding of specific transcription factors to CIITA promoter is affected by cannabinoids. Data obtained from our proposed pilot study will provide valuable information for our long-range plan to define the functional significance of cannabinoid receptors in microglia. Microglia form the first line of defense during infection in the CNS and are involved in various immune diseases including AIDS. ? ?
