肿瘤自分泌相关因子TGFβRⅡ对MKN-45人胃癌调控作用的体外及体内研究

Regulation effect of tumor autocrine related factor TGF beta RⅡ on the human gastric cancer MKN-45 cell and solid tumor

目的从肿瘤自分泌相关因子TGFβRⅡ探讨消痰散结方对MKN-45人胃癌调控作用。方法采用体外细胞培养和体内动物实验的方法,Weston blot法检测不同组别TGFβRⅡ的蛋白含量;real-time PCR法检测不同组别TGFβRⅡ的基因表达。结果体外实验中TGFβRⅡ的蛋白含量分别为空白对照组0.342±0.032、空白药物血清组0.359±0.043、消痰散结方血清组0.825±0.143、替加氟血清组0.969±0.077,四组间比较差异有统计学意义(P<0.01),消痰散结方组与空白对照组、空白药物血清组比较差异均有统计学意义(P<0.01);real-time PCR法检测体外实验中不同组别TGFβRⅡ的基因表达:空白药物血清组6.17±0.77、消痰散结方血清组6.64±0.97、替加氟血清组9.13±2.12,虽组间比较差异无统计学意义(P>0.05),但替加氟组与消痰散结方组均有上调其表达的趋势,替加氟的上调趋势更明显。体内实验中各组TGFβRⅡ的蛋白含量分别为空白对照组0.597±0.04、生理盐水组0.692±0.091、消痰散结方组0.639±0.099、替加氟化疗组0.465±0.064,四组间表达差异有统计学意义(P=0.035),进一步两两比较,替加氟组与生理盐水比较差异有统计学意义(P=0.042)。表明替加氟组不能上调体内实验瘤中TGFβRⅡ的蛋白表达,消痰散结方组与空白对照组比较虽然差异未见统计学意义,但有上调趋势。体内实验各组TGFβRⅡ的基因表达分别为空白模型组1.56±0.39、消痰散结方组3.30±1.17、替加氟化疗组6.44±3.50,不同组别TGFβRⅡ的基因表达差异有统计学意义(P<0.01),替加氟组表达高于模型组及消痰散结方组(P<0.05),消痰散结方组与模型组虽然表达差异无统计学意义,但有明显的上调趋势。结论消痰散结方对TGFβRⅡ蛋白和基因表达均有不同程度的上调作用,表明消痰散结方的治疗胃癌的作用可能是通过上调胃癌自分泌因子TGFβRⅡ发挥作用。

Objective To investigate the regulation effect of Xiaotansanjie Recipe on gastric cancer MKN-45 based on tumor autocrine related factor TGF beta RⅡ.Methods Used the method in vitro cell culture and in vivo animal experiment,Weston blot method was used to detect the protein content of TGF beta RⅡ in different groups and detected gene expression of TGF beta RⅡ in different groups by real-time PCR.Results The protein content of in vitro TGF beta RⅡ were control group 0.342±0.032,blank drug serum group 0.359±0.043,Xiaotansanjie Recipe group 0.825±0.143,tegafur group 0.969±0.077,respectively.There was significant difference between the four groups(P<0.01).There was significant difference(P<0.01) between the comparison of Xiaotansanjie Recipe group and blank control group and Xiaotansanjie Recipe group and blank drug serum group.Detected TGF beta RⅡ gene expression by the method of real-time PCR in vitro:blank drug serum group 6.17±0.77,Xiaotansanjie group 6.64±0.97,tegafur group 9.13±2.12 respectively.There was no significant difference between the four groups(P>0.05),but both of tegafur group and Xiaotansanjie group had the trend of increasing TGF beta RⅡexpression.The trend of the increased expression of tegafur serum group was more obvious.In vivo experiment,the protein content of TGF beta RⅡin each group were control group 0.597±0.04,saline group 0.692±0.091,Xiaotansanjie Recipe group 0.639±0.099,tegafur chemotherapy group 0.465±0.064,respectively.There was significant difference among the four groups(P=0.035).There was significant difference between the treatment group and the saline group(P=0.042).The results showed that the tegafur group could not raise the expression of TGF beta RⅡprotein in vivo.There was no significant difference between Xiaotansanjie group and control group,but the upward trend was presented.In vivo experiment the expression levels of TGF beta RⅡgenes:blank model group 1.56±0.39,Xiaotansanjie group3.30±1.17,tegafur chemotherapy group 6.44±3.50,respectively.There was significant difference among the three groups in the expression of TGF beta RⅡ(P<0.01).Tegafur group was higher than that in the model group and Xiaotansanjie group(P<0.05).Although the expression difference was not statistically significant,there was obvious upward trend between Xiaotansanjie Recipe group and model group.Conclusion Xiaotansanjie Recipe may upregulate TGF beta RⅡprotein and gene expression in different degrees,which indicate that the treatment effect of Xiaotansanjie Recipe on gastric carcinoma may play a role through upregulation of autocrine factor TGF beta RⅡ.