不同浓度茶多酚对脂多糖诱导的内皮细胞中NF-κB p65表达的影响

Effect of tea polyphenols on the expression levels of NF-κB p65 in HUVECs induced by lipopolysaccharide

目的探讨不同浓度茶多酚(TP)对牙龈卟啉单胞菌脂多糖(Pg-LPS)诱导的人脐静脉内皮细胞(HUVECs)中核因子-κB(NF-κB)p65表达影响。方法选取不同浓度的TP和Pg-LPS作用于HUVECs,用CCK-8细胞活性检测法对TP和Pg-LPS药物浓度进行筛选,选取Pg-LPS诱导TP预刺激后最适浓度的HUVECs建立炎症反应实验模型,将实验分为Control组、Pg-LPS组、LPS+TP1组(5 mg/L)和LPS+TP2组(40 mg/L),用Westernblot法检测细胞核及胞浆内NF-κB p65蛋白表达。结果在不同浓度TP作用下(5 mg/L、10 mg/L、20 mg/L、40 mg/L),12 hHUVECs增殖率较24 h和48 h高,差异有统计学意义(P<0.05),其中作用浓度在5~40 mg/L时促进HUVECs增殖,达到80 mg/L后出现抑制作用;与Control组比较,在不同时间、浓度条件下(4 h、8 h、24 h;0.1μg/mL、1μg/mL、10μg/mL),Pg-LPS均会对细胞增殖产生抑制作用,且抑制率与作用时间和浓度相关(P<0.05),其中在相同作用时间下,浓度为10μg/mL的Pg-LPS的细胞抑制率最高,在相同浓度下,Pg-LPS作用24 h后细胞抑制率最高,在Pg-LPS的作用下,细胞核内的p65表达增加而胞浆内的p65表达降低;选取一定浓度的TP预处理HUVECs后,细胞核内NF-κB p65蛋白表达降低,浆内NF-κB p65蛋白表达增加。结论TP对HUVECs的增殖与抑制作用与浓度及时间有密切的关系,具有低浓度促进、高浓度抑制与短时促进、长时抑制的作用;TP抑制Pg-LPS诱导的HUVECs中NF-κB p65由胞浆转位至细胞核,即抑制NF-κB p65活化,对HUVECs具有一定的保护作用。

Objective To explore the effect of tea polyphenols(TP)on porphyromonas gingivalis lipopolysac-charide(Pg-LPS)-induced nuclear factor kppa B(NF-κB)p65 expression in human umbilical vein endothelial cells(HU-VECs).Methods Different concentrations of TP and Pg-LPS were selected to treat HUVECs.The concentrations of TP and Pg-LPS were screened by CCK-8 cell activity assay.The optimal concentration of HUVECs with Pg-LPS-induced TP prestimulation was selected to establish an experimental model of inflammatory response.The experiment was divided in-to four groups:control group,PG-LPS group,LPS+TP1(5 mg/L),LPS+TP2(40 mg/L).The expressed of intracellular NF-κB p65 protein was detected by western blot.Results Under different TP concentration(5 mg/L,10 mg/L,20 mg/L,40 mg/L),the proliferation rate of 12 h HUVECs was significantly higher than that of 24 h and 48 h(P<0.05),and the pro-liferation of HUVECs was promoted at the concentration of 5-40 mg/L and inhibited at the concentration of 80 mg/L.In different time and concentrations(4 h,8 h,24 h;0.1μg/mL,1μg/mL,10μg/mL),Pg-LPS could inhibit cell proliferation,and the inhibition rate was correlated with time and concentration(P<0.05);at the same time,10μg/mL Pg-LPS had the highest inhibition rate on cells,and at the same concentration of Pg-LPS,the inhibition rate was the highest at 24 h.What's more,Pg-LPS increased the expression of p65 in nucleus and decreased the expression of p65 in cytoplasm.TP can de-crease the expression of p65 in nucleus and increase the expression of p65 in cytoplasm.Conclusion TP has a close rela-tionship with concentration and time,and has the functions of low concentration promotion,high concentration inhibition,short-term promotion and long-term inhibition.TP inhibits the transformation of NF-κB p65 from cytoplasm to nucleus,the same as inhibition of activation of NF-κB p65,which has a certain protective effect on HUVECs.