SIRT1对脂多糖活化BV-2细胞分泌IL-6和TNF-α的调节作用

Role of silence information regulator 1 in regulation of lipopolysaccharide-induced secretion of interleukin-6 and tumor necrosis factor-a in activated BV-2 cells

目的观察脂多糖（LPS）对BV-2细胞形状变化及白介素-6（1L-6）和肿瘤坏死因子-α（TNF-α）分泌情况的影响，探讨沉默信息调控因子1（SIRT1）在LPS诱导BV-2细胞释放前炎症因子过程中的作用。方法BV-2细胞分为调零组、对照组及LPS组，调零组不含细胞，只加培养基；对照组常规培养：LPS组分别加入不同浓度的LPS；MTT法检测各组存活率。另取BV-2细胞分为调零组、对照组、DOSO药物载体组及白藜芦醇组、Sirtinol组，调零组、对照组处理方法同前，DOSO药物载体组加人体积分数为0．3％的DMSO；白藜芦醇组加入10、25、50、100μmol／L白藜芦醇；Sirtinol组加入1、2．5、5、10、25、50μmol／LSirtinl；MTT法检测各组细胞存活率。根据实验结果，选取适当浓度的LPS、白藜芦醇及Sirtinol，将BV-2细胞分为对照组、LPS组、白藜芦醇＋LPS组、Sirtinol＋LPS组，分别用ELISA方法检测各组细胞相应处理12、24h后上清液中的IL-6和TNF-α的含量．Westemblotting检测SIRT1在上述各组细胞加药处理24h后的表达水平。结果与对照组相比，LPS组BV-2细胞数目增多，胞体肿胀，突起变短、增多。随着LPS浓度的增加，BV一2细胞存活率明显增加，差异有统计学意义（P〈0．05）。与对照组比较，LPS组BV-2细胞分泌的IL-6、TNF-α增多，SIRT1的表达量下降，差异有统计学意义（P〈0．05）。与LPS组比较，经白藜芦醇处理后，细胞内SIRTl的表达量上升，而IL-6、TNF-α的分泌水平下降，差异有统计学意义（P〈0．05）；相反，细胞经Sirtinol处理后，胞内SIRT1的表达量下降，IL-6、TNF-OL的分泌水平上升，差异有统计学意义（P〈0．05）。结论LPS可显著改变BV-2细胞形状并诱导细胞前炎症因子的表达，干预SIRT1的表达量对上述现象有明显的调节作用。

Objective To observe the effect oflipopolysaccharide （LPS） on the cell form of BV-2 cells and the expressions ofinterleukin-6 （IL-6） and tumor necrosis factor-a （TNF-a） so as to detect the role of silence information regulator 1 （SIRT1） in regulation of LPS-induced proinflammatory cytokines production in activated BV-2 cells. Methods BV-2 cells were divided into control group （normal culture medium） and treatment groups; BV-2 cells in the treatment groups were subdivided into LPS treatment groups, Resveratrol＋LPS treatment groups and Sirtinol＋LPS treatment groups （cultured with different concentrations of LPS, SIRT1 activator Resveratrol or S1RT1 inhibitor Sirtinol, respectively）. MTT assay was employed to identify the cell survival after the inducement. Based on the above MTT results, the cells were then grouped into the control group, LPS treatment group, Resveratrol＋LPS treatment group and Sirtinol＋LPS treatment group having suitable concentrations of LPS, Resveratrol and Sirtinol; then, the levels of IL-6 and TNF-a were measured with enzyme-linked immuno sorbent assay （ELISA） at 12 and 24 h after the inducement; and the expression of SIRT1 at 24 h after the inducement was detected by Western blotting. Results As compared with those in the controlgroup, the BV-2 cells in the LPS treatment group had increased cell number, hypertrophic cell body, and shorten cell processes. The cell survival rate increased with increased concentrations of LPS. As compared with those in the control group, the levels of IL-6 and TNF-a in LPS treatment group increased and level of SIRT 1 decreased with significant differences （P〈0.05）. Significantly increased levels of IL-6 and TNF-a and obviously decreased expression of SIRT1 in the Resveratrol＋LPS treatment group were noted as compared with those in the LPS treatment group （P〈0.05）; conversely, significantly decreased levels of IL-6 and TNF-a and obviously increased expression of SIRT1 in the Sirtinol＋LPS treatment group were noted as compared with those in the LPS treatment group （P〈0.05）. Conclusion LPS can change the morphology of BV-2 cells, and induce the levels ofproinflammatory cytokines; impairment of SIRT 1 may contribute to such progress obviously.