脂多糖对大鼠膀胱上皮细胞中Toll样受体4与核因子-κB表达的影响

Effect of lipopolysaccharide on Toll-like receptors 4 and nuclear factor-κB expression in bladder epithelial cells of rats

目的观察体外培养的大鼠膀胱上皮细胞中Toll样受体4(TLR4)与核因子-κB(NF-κB)在脂多糖(LPS)刺激下的表达变化。方法分离幼龄Sprague-Dawley大鼠膀胱黏膜,培养上皮细胞。将分离纯化的上皮细胞分为对照组和LPS刺激1、2、3组;对照组不给予任何干预措施,LPS刺激1、2、3组给予LPS(0.01 g·L-1)分别作用1、5、24 h。采用实时定量荧光酶联免疫反应及Western blot技术检测TLR4与NF-κB p65表达量的变化。结果对照组和LPS刺激1、2、3组的TLR4 mRNA相对表达量分别为1.00±0.10、1.32±0.27、1.81±0.10、1.61±0.35;TLR4蛋白相对表达量分别为1.00±0.18、1.36±0.12、1.95±0.08、1.79±0.21。与对照组比较,LPS刺激1、2、3组TLR4 mRNA和蛋白的相对表达量均增多(P<0.05);LPS刺激2、3组TLR4 mRNA和蛋白相对表达量较LPS刺激1组增多(P<0.05);LPS刺激2、3组TLR4 mRNA和蛋白相对表达量比较差异无统计学意义(P>0.05)。对照组和LPS刺激1、2、3组NF-κB p65 mRNA相对表达量分别为1.00±0.08、1.60±0.05、1.51±0.06、1.39±0.04;NF-κB p65蛋白相对表达量分别为1.00±0.08、1.87±0.11、1.69±0.10、1.94±0.18。与对照组比较,LPS刺激1、2、3组NF-κB p65 mRNA和蛋白相对表达量均增多(P<0.05);LPS刺激1、2、3组间NF-κB p65 mRNA和蛋白相对表达量比较差异均无统计学意义(P>0.05)。结论 LPS可上调膀胱上皮细胞中TLR4与NF-κB p65的表达。

Objective To observe the changes of Toll-like receptors 4（ TLR4） and nuclear factor-κB（ NF-κB） expression in bladder epithelial cells of rat cultured in vitro under the stimulating of lipopolysaccharide（ LPS）. Methods The bladder mucosa were separated from infant rats and then the bladder epithelial cells were cultured. Then the isolated and purified bladder epithelial cells were divided into control group,LPS stimulation group 1,LPS stimulation group 2 and LPS stimulation group 3; the control group did not give any intervention measures,the LPS stimulation 1,2,3 group was given LPS（ 0. 01 g·L- 1） for 1,5,24 h respetively. Real time polymerase chain reaction and Western blot technologies were used to determine the expression of TLR4 and NF-κB. Results The relative expression of TLR4 mRNA in control group,LPS stimulation1,2,3 group was 1. 00 ± 0. 10,1. 32 ± 0. 27,1. 81 ± 0. 10,1. 61 ± 0. 35; the relative expression of TLR4 protein in the four groups was 1. 00 ± 0. 18,1. 36 ± 0. 12,1. 95 ± 0. 08,1. 79 ± 0. 21; the relative expression of TLR4 mRNA and protein in LPS stimulation group 1,2,3 was significantly higher than those in control group（ P 〈0. 05）; the relative expression of TLR4 mRNA and protein in LPS stimulation group 2,3 was significantly higher than those in LPS stimulation group 1（ P 〈0. 05）; there was no statistic difference of the relative expressions of TLR4 mRNA and protein between the LPS stimulation group 2 and LPS stimulation group 3（ P 〉0. 05）. The relative expression of NF-κB p65 mRNA in control group,LPS stimulation group 1,2,3was 1. 00 ± 0. 08,1. 60 ± 0. 05,1. 51 ± 0. 06,1. 39 ± 0. 04; the relative expression of NF-κB p65 protein in the four groups was1. 00 ± 0. 08,1. 87 ± 0. 11,1. 69 ± 0. 10,1. 94 ± 0. 18; the relative expression of NF-κB p65 mRNA and protein in LPS stimulation group 1,2,3 was significantly higher than those in control group（ P 〈0. 05）; there was no statistic difference of the relative expressions of NF-κB p65 mRNA and protein among LPS stimulation group 1,2,3（ P 〉0. 05）. Conclusion LPS can raise the expression of TLR4 and NF-κB in bladder epithelial cells of rats.