摘要
目的探讨胃复春抑制幽门螺杆菌(Hp)诱导人胃GES-1细胞的炎症及其与NF-κB信号通道的关系。方法采用Hp国际标准产毒株(CagA+、VacA+)NCTCl 1637感染人胃GES-1细胞,采用胃复春进行干预,根据噻唑蓝(MTT)法实验筛选出胃复春低、中、高浓度组(浓度分别为5、10、20μg/mL),并设置空白组、Hp模型组(模型组)。然后采用MTT测定法测定人胃GES-1细胞生长抑制率,流式细胞仪分析细胞周期,ELISA法检测上清液中白介素-8(IL-8)、白介素-4(IL-4)等的水平,Westen blot检测核因子κB(NF-κB)蛋白表达情况。结果 MTT法显示胃复春对胃GES-1细胞的IC5为10μg/mL,取5、10、20μg/mL为低、中、高浓度组。与Hp共培养24h后的GES-1细胞周围出现碎片,细胞之间连接减少,呈现核质融合状态;予胃复春低、中、高3个浓度药物干预后的GES-1细胞,贴壁细胞逐渐增多,细胞形态逐渐复原。Hp感染后,模型组上清液中IL-8明显高于空白组(P<0.05),此后逐渐回落,而予不同浓度胃复春干预后,IL-8水平在12、24、48、72h均低于模型组,其中以高浓度组下降最为明显,差异有统计学意义(P<0.05)。而在模型组上清液中抑炎因子IL-4明显低于空白组,在高浓度胃复春组IL-4明显升高,差异有统计学意义(P<0.05),而中、低浓度组与空白组比较,差异无统计学意义。Hp感染人胃GES-1细胞后60min,核内P65表达增高,IkBα表达下降,而予胃复春干预后核内P65表达下降,IkBα表达增强。结论胃复春可以有效平衡Hp诱导人GES-1细胞的促炎性因子与抑炎因子表达,其机制可能与胃复春抑制p65进入胞核,抑制Hp所刺激的IkBα的降解有关,胃复春是治疗Hp相关胃炎的有效药物。
Objective To study the effect of Weifuchun on inflammation of Helicobacter pylori （Hp）-infected gastric epithelial cells （GES-1 ） and its correlation with NF-KB signaling pathway. Methods Hp standard home-made strain （CagA ＋, VacA ＋ ） NCTCI 1637 infected GES-1 cells were used. Weifuchun was used as intervention. Weifuchun of different concentrations （5,10, and 20μg/mL） were screened by M-IF assay. A blank group and the model group were set up. Then the growth inhibition rate of drugs on gastric epithelial GES- 1 cells was detected with MTT assay. Cell cycle was detected using flow cytometry. The supernatant liquid was separated to detect the contents of IL-8 and IL-4 by ELISA.The protein expression level of NF-κB was detected by Western blot analysis. Results Ml-r assay indicated significantly inhibitory effect of Weifuchun on GES-1 cells [5% inhibiting concentration （IC5） was 10 μg/ml in the Weifuchun group. After GES-1 and Hp were cultured together,the contents of IL-8 in the supernatant were more obviously higher in the model group than in the blank group （P 〈0.05）, and then gradually decreased. After treatment with different concentrations of Weifuchun, the levels of IL-8 in the supernatant were less when compared with the model group at 12, 24, 48, and 72 h （P 〈0.05）. The decrement was the most significant in the high dose Weifuchun group. The IL-4 level in the supernatant was obviously lower in the model group than in the blank group. It obviously increased in the high concentration Weifuchun group （P 〈0.05）. There was no statistical difference in the IL-4 level between middle, low concentration Weifuchun group and the blank group （P 〉0.05）. The protein expression of intranuclear P65 increased and that of IkBα decreased 60 min after Hp infection. But the protein expression of intranuclear P65 decreased and the protein expression of IkBα increased after intervention of Weifuchun. Conclusions Weifuchun adjusted H. pylori induced IL- 8 and IL-4 production by gastric epithelial cells through blocking NF-κB pathways. Its mechanisms might possibly lie in inhibiting p65 from entry into nucleus and the degradation of IkBoL. Weifuchun was an effective drug for treatment of Hp correlated chronic gastritis.
出处
《中国中西医结合杂志》
CAS
CSCD
北大核心
2014年第4期450-454,共5页
Chinese Journal of Integrated Traditional and Western Medicine
基金
浙江省自然科学基金资助项目(No.LY12H29002)
浙江省中医药管理局基金资助项目(No.2011ZB032,No.2010ZB045)
