Entada africana fraction CH2Cl2/MEOH 5% inhibits inducible nitric oxide synthase and pro-inflammatory cytokines gene expression induced by lipopolysaccharide in microglia

BACKGROUND Inflammatory response in the CNS mediated by microglia cells play an important role in host defense and is implicated in the pathology of neurodegenerative diseases. We investigated the capacity of Entada africana to protect microglia from inflammatory insults by exploring the effect of the CH₂Cl₂/MEOH 5% fraction (Ea5) on pro-inflammatory cytokines mRNA expression. Finally, we studied the effect of Ea5 on the inhibition of p38 MAPK Kinase. The results were compared to those obtained with Baicalin, a well reported anti-inflammatory flavonoid. METHODS Barks from E. africana were harvested in 2010, in the west region of Cameroon. A crude extract was prepared using CH₂Cl₂/MEOH 1:1 V/V. The crude extract obtained was further fractionated by flash chromatography. A mouse microglia cell line (N9) was stimulated by LPS with or without different concentrations of Baicalin and Ea5. The release of NO was evaluated using the Griess method. The expression of pro-inflammatory cytokines mRNA (TNFα, IL-1β, IL-6) and iNOS/NO were measured by RT- PCR. The inhibition of p38 MAPK Kinase was assessed using ELISA. RESULTS We found that Ea5, as well as Baicalin inhibited LPS-induced NO production in a dose dependent manner. Ea5 was most active in term of NO inhibition (87.07%), in comparison to Baicalin (70.85%). The expression of TNFα, IL-1β, IL-6 and iNOS was strongly suppressed by Ea5 in microglia. Ea5 also inhibited the activity of p38MAPK Kinase, up to 30% for the concentrations tested, whereas a prominent inhibition was obtained with Baicalin. CONCLUSION These results suggest that E. africana may contain promising compounds useful for the treatment of diseases cause by over-activation of microglia such as Alzheimer disease and other neurological diseases.