半夏泻心汤人含药血清对HP感染GES-1细胞TGF-β/Smad信号通路的影响

Effect of Banxia Xiexin Tang Medicated Serum on HP-infected GES-1 Cells Through TGF-β/Smad Signaling Pathway

目的：探讨半夏泻心汤人含药血清对幽门螺杆菌（Helicobacter pylori,HP）感染人胃黏膜上皮GES-1细胞转化生长因子-β（TGF-β）/信号转导蛋白（Smad）信号通路的影响,以揭示该方治疗消化性溃疡可能的作用机制。方法：GES-1细胞分为空白组,HP感染模型组,健康人血清组,雷贝拉唑血清组,用药前血清组,10%半夏泻心汤血清组,20%半夏泻心汤血清组。除空白组外,其余各组均接种HP造成幽门螺杆菌感染模型,采用雷贝拉唑、半夏泻心汤含药血清及用药前、健康人血清进行干预,24 h后收集细胞及其上清液,噻唑蓝（MTT）比色法检测细胞增殖情况;蛋白质免疫印迹法（Western blot）检测Smad2/3,Smad7,TGF-β1及磷酸化Smad2/3（p-Smad2/3）,磷酸化Smad7（p-Smad7）蛋白的表达。结果：与空白组比较,HP可明显抑制GES-1细胞的增殖,降低GES-1细胞内Smad2/3,TGF-β1及p-Smad2/3蛋白表达,增加Smad7及p-Smad7蛋白表达（P〈0.05,P〈0.01）;与HP感染模型组比较,雷贝拉唑血清组及10%,20%半夏泻心汤血清组可明显降低HP对GES-1细胞增殖的抑制,增加GES-1细胞内Smad2/3,TGF-β1,p-Smad2/3蛋白表达,并明显降低Smad7,p-Smad7蛋白表达（P〈0.05,P〈0.01）;与用药前血清组比较,加入雷贝拉唑、半夏泻心汤含药血清后均可降低HP对GES-1细胞增殖的抑制,其中雷贝拉唑血清组及20%半夏泻心汤血清组可增加GES-1细胞内Smad2/3,TGF-β1,p-Smad2/3蛋白表达,降低Smad7,p-Smad7蛋白表达（P〈0.05,P〈0.01）。结论：半夏泻心汤能够减轻HP诱导GES-1细胞的损伤,促进GES-1细胞增殖,治疗HP相关性消化性溃疡,提高溃疡愈合质量,其机制可能与其调控TGF-β/Smad信号通路有关。

Objective： To investigate the effects of Banxia Xiexin Tang medicated serum on the Helicobacter pylori（ HP）-infected GES-1 cells through transforming growth factor-β（ TGF-β）/Smad signaling pathway to reveal the potential effect and mechanism of this prescription in treatment of heal peptic ulcer. Method：The GES-1 cells were divided into 7 groups,including blank control group,HP-infected model group,healthy human serum group,Rabeprazole serum group,pre-medicated serum group,10% and 20% Banxia Xiexin Tang serum groups. Expect those in blank control group,HP was vaccinated in all the other groups to establish helicobacter pylori model,and then Rabeprazole serum,Banxia Xiexin Tang medicated serum and premedication serum and healthy human serum were adopted for intervention. After 24 hours,cells and their supernatant were collected,and methylthiazolyldiphenyl-tetrazolium bromide（ MTT） assay was used to test the cell proliferation; the protein expression levels of Smad2/3, Smad7, transforming growth factor-β1（ TGF-β1） and phosphorylated Smad2/3（ p-Smad2/3）,phosphorylated Smad7（ p-Smad7） were detected by using Western blot. Result： As compared with the blank control group,HP significantly inhibited proliferation of GES-1 cells,decreased protein expression levels of Smad2/3,TGF-β1 and p-Smad2/3 in GES-1 cells,and increased protein expression levels of Smad7 and p-Smad7（ P〈0. 05,P〈0. 01）. As compared with HP-infected model group,Rabeprazole serum group and 10%,20% Banxia Xiexin Tang serum groups obviously decreased the inhibition on proliferation of GES-1 cells,increased protein expression levels of Smad2/3,TGF-β1 and p-Smad2/3 in GES-1 cells,and significantly decreased Smad7 and p-Smad7 protein expression levels（ P〈0. 05,P〈0. 01）. As compared with pre-medicated serum group,the inhibition on proliferation of GES-1 cells was decreased in Rabeprazole serum group and Banxia Xiexin Tang serum groups; the protein expression levels of Smad2/3,TGF-β1 and p-Smad2/3 in GES-1 cells were increased and the protein expression levels of Smad7 and p-Smad7 were decreased in Rabeprazole serum group and20% Banxia Xiexin Tang medicated serum group（ P〈0. 05,P〈0. 01）. Conclusion： Banxia Xiexin Tang alleviated HP-induced GES-1 cells injury,promoted GES-1 cells proliferation,and improved the curing quality of ulcers in the treatment of HP-associated peptic ulcer. The mechanism may be related to regulating the TGF-β/Smad signaling pathway.