姜黄素对脂多糖激活的小胶质细胞iNOS表达的抑制及抗氧化作用

Effect of Curcumin on iNOS Expression in LPS-activated Microglia Cells and Anti-oxidation

用姜黄素预处理小胶质细胞株BV,1h后加用脂多糖(200ng/ml)进行刺激,通过MTT检测细胞活性;硝酸还原酶法检测细胞上清液中一氧化氮(NO)的含量;Western印迹、免疫细胞化学染色检测细胞活化后形态及诱导型一氧化氮合酶(iNOS)蛋白的表达;瞬时转染和荧光素酶报告基因鉴定iNOS和NF-κB基因表达活性;SOD和GSH-Px检测姜黄素的抗氧化能力.结果证明,脂多糖可促使小胶质细胞活化,使iNOS和NF-κB基因表达活性显著增强;iNOS蛋白表达明显上调,NO释放增多;细胞内SOD和GSH-Px活性明显下降.而姜黄素(10μmol/L)可以显著抑制活化后小胶质细胞NO的产生、iNOS蛋白的表达及iNOS-Luc、NF-κB-Luc的表达活性,其机制可能通过NF-κB的信号转导途径抑制iNOS的表达.姜黄素可通过提高细胞内SOD和GSH-Px的活性发挥抗氧化作用.

Microglia BV cells were stimulated with lipopolysaccharide（ LPS, 200 mg/ml） after pretreatment with curcumin for 1 hour. The cytotoxicity of curcumin was measured by MTT assay. The production of nitric oxide （NO） was determined with nitrate reductase. The expression of inducible nitric oxide synthase（iNOS） was detected with Western blotting and immunocytochemical stain. Transcriptional activities of iNOS and NF- kB were detected by transient transfection. Anti-oxidant activities of curcumin were determined by SOD and GSH-Px assays. The results showed that LPS was able to activate microglia cell and elevate the expression of iNOS and NF-kB gene. The activities of SOD and GSH-Px in cells were apparently suppressed. Whereas, 10 μmol/L curcumin was significantly able to inhibit the production of NO in activated microglia cells, the expression of iNOS protein, and activities of iNOS-Luc and NF-kB-Luc. The mechanism might be revolved in iNOS repression induced by NF-kB signaling pathway. Anti-oxidant effect of curcumin could be achieved through activating the activities of SOD and GSH-Px in LPS activated microglia cells.