Implicating MicroRNAs as Regulators of Microglia and Astrocyte Responses in Human CNS Inflammatory Disease (P5.018)

OBJECTIVE: To characterize and compare cell-specific microRNA (miRNA) expression in human astrocytes and microglia/macrophages within the inflamed human CNS and demonstrate their capacity to regulate disease-relevant responses. BACKGROUND: The functional role of astrocytes and microglia in the inflamed CNS remains enigmatic, yet both cell types are important and direct contributors to both inflammatory and repair mechanisms following CNS injury. MiRNAs are non-coding RNA molecules involved in regulating post-translational activities of mRNA transcripts. We hypothesize that expression of distinct miRNAs in glial cells will influence the molecular mechanisms responsible for their unique properties under inflammatory conditions. Identifying differential patterns of miRNA expression may provide insight into the capacity for glial cells to influence CNS injury and repair. DESIGN/METHODS: Laser-capture microdissection (LCM) was used to isolate CD68+ and GFAP+ cells from formalin-fixed autopsy control and MS brain samples. MiRNA expression was determined by TaqMan qPCR miRNA expression assays. For in vitro studies, astrocytes and microglia were isolated from fetal/adult CNS tissue and activated using IL-1s and LPS, respectively. Transfection of miRNA mimics and inhibitors was performed using a RNAi lipofectamine reagent. RESULTS: In situ LCM miRNA analysis of microglia/ macrophages and astrocytes in control brain versus MS lesions demonstrated differential expression of a subset of inflammation-related miRNAs. Guided by these results, transfection of either a mir-146a inhibitor or a mir-125b mimic in vitro, using isolated human microglia and macrophages, increased LPS-induced expression of TNF and IL-6. In astrocytes, mimics of both mir-365 and mir-146b-5p significantly attenuated IL-1s-induced IL-6 release. Astrocyte CXCL10 (IP-10) expression was also decreased following transfection with a mir-146b-5p mimic, but was not influenced by mir-365 mimic. CONCLUSIONS: Our results demonstrate that unique miRNAs, implicated in situ within astrocytes and microglia/macrophages of MS lesions, can impact glial responses that may be relevant in mechanisms of injury and repair. Disclosure: Dr. Moore has nothing to disclose. Dr. Rao has nothing to disclose. Dr. Fuh has nothing to disclose. Dr. Bedell has received personal compensation for activities with Biospective, Inc. Dr. Bedell holds stock and/or stock options in Biospectivce, Inc., which sponsored research in which Dr. Bedell was involved as an investigator. Dr. Bedell has received research support from Biospective, Inc. Dr. Ludwin has nothing to disclose. Dr. Bar-Or has received personal compensation for activities with Amplimmune, Aventis, Bayhill Therapeutics, Berlex/Bayer, Biogen Idec, BioMS, Diogenix, Eli Lilly & Company, EMD Serono, Genentech, Inc., Genzyme, GlaxoSmithKline, Inc., Guthy-Jackson/GGF, Medimmune, Mitsubishi Pharma, Novartis, Ono Pharma, Receptos, Roche, Sanofi-Aventis Pharmaceuticals, Inc., Teva Neuroscience, and Wyeth Pharmaceuticals. Dr. Bar-Or has received research support from Amplimmune, EMD Serono, and Novartis. Dr. Antel received personal compensation from BiogenIdec, TEVA, emdSerono, Genzyme, Sanofi Aventis, Novartis Dr. Antel received personal compensation from Multiple Sclerosis Journal for serving as a journal editor. . . . . Dr. Antel received financial support for research activities from Novartis effects of FTY on the immune and nervous systems.