The expression and action of inflammatory cytokine molecules in the brain requires tight regulation since they alter cognitive and motivational systems and if over-expressed can be neurotoxic. Preliminary data presented in this application indicate that microglia cultured from brain of aged mice spontaneously secrete copious amounts of the inflammatory cytokine, interleukin-6 (IL-6). Consequently, a positive correlation between age and IL-6 concentration in brain has been found, even in healthy mice. We believe that chronic over-expression of the IL-6 gene in the brain of the elderly may establish a state that is permissive to the onset of neurodegenerative disease and be causally related to anorexia and impaired memory and learning. Unfortunately, little to nothing is known about how aging affects IL-6 gene expression in the brain, which is the focus of this proposal. Here, several approaches are proposed to (1) define the expression of IL-6 in the brain of aged mice, (2) determine the effects of age on the transcription factors that regulate IL-6 gene expression in brain and in glial cells; (3) explore the ability to regulate IL-6 gene expression in the brain of aged mice with supplemental dehydroepiandrosterone (DHEA), and (4) describe the effects of DHEA on the transcription factors regulating the IL-6 gene. Specifically, we will employ immunohistochemistry and in situ hybridization to map IL-6 expression in aged mouse brain, and two-color flow cytometry to evaluate the proportion of astrocytes and microglia cultured from brain of neonate, adult, and aged mice expressing IL-6. We will determine if the increase in IL-6 production in aged brain is due to an increase in active Nuclear Factor-KB or a decrease in Recombinant Signal Sequence Binding-Protein JK, which has been recently shown to inhibit spontaneous IL-6 production by occupying the IL-6-KB site in the IL-6 promoter/enhancer region. Moreover, the critical possibility that the age-related change(s) in transcription factors regulating IL-6 gene expression in the brain is due to the decline in DHEA, which occurs with age, will be tested. These novel studies will provide needed information to better understand, treat, or prevent the neuropathophysiological manifestations of old age. There are strong preliminary data supporting each of the objectives and all of the techniques needed to successfully complete these studies in mice have been developed.
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