摘要
目的观察Toll样受体4(Toll-like receptor4,TLR4)及myeloid differentiation protein2(MD2)反义基因对内毒素(lipopolysaccharide,LPS)诱导的肺泡Ⅱ型细胞NF-κB活化的影响。方法培养的肺泡Ⅱ型细胞分为:正常细胞(对照)组、LPS组、LPS+转染空载体组、LPS+转染TLR4反义基因组、LPS+转染MD2反义基因组、LPS+转染TLR4-MD2反义基因组(n=8)。Northern blot检测转染细胞的TLR4及MD2 mRNA表达,Western blot检测转染细胞TLR4及MD2蛋白表达,电泳迁移率改变法检测NF-κΒ的活性,ELISA测定细胞培养上清中TNF-α和IL-6含量。结果与对照组比较,LPS刺激后的细胞TLR4及MD2mRNA和蛋白表达、NF-κB活性、TNF-α和IL-6的生成均显著增加(P<0.01);而转染反义基因组细胞与其他LPS刺激组比较,TLR4及MD2mRNA和蛋白的表达、NF-κB活性及TNF-α和IL-6的含量均明显降低(P<0.01)。结论 TLR4及MD2反义基因能有效抑制LPS诱导的肺泡Ⅱ型细胞的活化。
Objective To observe effects of the antisense genes of Toll-like receptor 4 (TLR4 ) and myeloid differentiation protein-2 (MD2) on the activation of NF-κB in alveolar typeⅡepithelial cells (ATⅡcells) induced by lipopolysaccharide (LPS). Methods Cultured ATⅡcells were randomized to normal cells (control) group,LPS group,LPS+empty vector group,LPS+TLR4 antisense gene group,LPS+MD2 antisense gene group,LPS+TLR4-MD2 antisense gene group. The expressions of TLR4 and MD2 mRNA and protein in ATⅡcells were detected by Northern blotting and Western blotting respectively. The activity of NF-κB in ATⅡcells was measured with electrophoretic mobility shift assay (EMSA). The TNF-α and IL-6 concentrations in the supernatant of cultured ATⅡcells were tested with ELISA.Results Compared with control groups,the expressions of TLR4 and MD2 mRNA and protein,the NF-κΒ activity,the levels of TNF-α and IL-6 were increased remarkably in LPS group and LPS+empty vector group (P0.01). After stimulating with the antisense genes,the expressions of TLR4 and MD2 mRNA and protein,the NF-κΒ activity,the levels of TNF-α and IL-6 were decreased remarkably than in LPS and LPS+empty vector group (P0.01). Conclusion The antisense gene of TLR4 and MD2 can inhibit activation of ATⅡ cells induced by LPS.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2010年第12期1271-1274,共4页
Journal of Third Military Medical University
基金
国家自然科学基金重点项目(39730210)~~
关键词
内毒素类
肺泡
上皮细胞
细胞
培养的
受体
细胞表面
信号转导
TOLL样受体4
MD2
lipopolysaccharides
pulmonary alveoli
epithelial cells
cells
cultured
receptors
cell surface
signal transduction
Toll-like receptor 4
myeloid differentiation protein-2