[P2–185]: THE STUDY OF SUPPRESSION OF HYPOXIA‐INDUCED INFLAMMATION BY TIBETAN MEDICINE RATANASAMPIL IN MICROGLIA CELLS

Ratanasampil (RNSP), a traditional Tibetan medicine, improves the cognitive function of mild-to-moderate AD patients living at high altitude, as well as learning and memory in an AD mouse model (Tg2576); however, mechanism underlying the effects of RNSP is unknown. Most studies suggested hypoxia facilitates the pathogenesis of AD through accelerating the β-Amyloid (Aβ) accumulation. Little is known about the hypoxia induced inflammation in AD pathogenesis. Hypoxia induced microglia activation and animal studies have been shown correlated with neuronal and cognitive decline. To explore the potential protective effect and molecular mechanism of RNSP on hypoxia and reoxygenation induced inflammation using MG6 microglia (MG6 cells). MG6 cells were seeded in 24-well plated for 24h before experiment. The control or 1h pre-treatment of RNSP in MG6 were maintained in hypoxia chamber (1% O2, 5%CO2, 95%N2) for 6h. After 6h hypoxia, MG6 were cultured in normal condition for reoxygenation for 12h(H6/R12). The cell viability assay was conducted by MTT assay; The mRNA level of inflammatory cytokines was measured by real-time PCR. HIF-1a protein expression level was measured by western blotting. The hypoxia significantly reduced the cell viability of MG6 cells, and RNSP significantly inhibited the hypoxia (12h)-reduced the cell viability of MG6 cells, respectively, which can be significantly protected by pre-treatment of RNSP. RNSP significantly inhibited the hypoxia reduced the cell viability of MG6 cells after 6h of hypoxia followed by 12h reoxygenation (H6/R12), respectively. The mRNA level of Interleukin6 (IL-6), Interleukin-1b (IL-1b) and Inducible Nitric Oxide Synthase (iNOS) were significantly up-regulated by MG6 cells during hypoxia-reoxygenation(H6/R12). The up-regulation of inflammatory cytokines were suppressed by pretreatment of RNSP. Furthermore, RNSP was found to significantly reduce the hypoxia-upregulated expression of HIF-1a in MG6 cells. Our data suggest that RNSP may inhibit the microglia-mediated neuroinflammation, thus provide a potential mechanism of the RNSP in improving the cognitive function in AD during hypoxia and H/R. The suppression of inflammation by RNSP may provide therapeutic strategy for AD treatment. The on-going work of HIF-1a expression assay may also represent the potential therapeutic target of RNSP on the inflammation induced by hypoxia.