基于PTP1B探讨芪术抗癌方调控同、异源黏附串扰抑制结肠癌转移的作用机制  

Exploring the regulation of homo⁃heterologous adhesion crosstalk by Qizhu Kangai Formula based on PTP1B research on the mechanism of inhibiting colon cancer metastasis

作　　者：万林鹭 梁研 孙若岚 梁众擎 殷启航 王旭[1] 赵凡[1] 唐德才[1] WAN Linlu;LIANG Yan;SUN Ruolan;LIANG Zhongqing;YIN Qihang;WANG Xu;ZHAO Fan;TANG Decai(School of Traditional Chinese Medicine and School of Integrated Chinese and Western Medicine,Nanjing University of Chinese Medicine,Nanjing 210023,China)

机构地区：[1]南京中医药大学中医学院·中西医结合学院,南京210023

出　　处：《北京中医药大学学报》2024年第2期249-259,共11页Journal of Beijing University of Traditional Chinese Medicine

基　　金：国家自然科学基金项目(No.82274116);江苏省中医药科技发展专项(No.2020ZX01)。

摘　　要：目的探讨芪术抗癌方调控同、异源黏附串扰关键蛋白蛋白酪氨酸磷酸酶1B(PTP1B)抑制结肠癌转移的作用机制。方法制备空白血清及芪术抗癌方含药血清。采用液质联用技术分析芪术抗癌方含药血清的有效化学成分。通过质粒转染,体外构建HCT116细胞的PTP1B过表达模型(OE⁃PTP1B HCT116)。CCK-8法检测OE⁃PTP1B HCT116存活率后选择20%空白血清及5%、10%、20%芪术抗癌方含药血清。采用划痕实验、transwell实验和黏附实验分别检测细胞迁移、侵袭及黏附情况,采用蛋白质印迹法检测PTP1B、黏着斑蛋白(vinculin)、E-钙黏蛋白(E⁃cadherin)、整合素αν亚单位(integrinανsubunit)、整合素β3亚单位(integrinβ3 subunit)蛋白表达水平。结果液质联用分析得到芪术抗癌方含药血清中12个主要有效化学成分,含量最高的是毛蕊异黄酮苷。OE⁃PTP1B HCT116细胞PTP1B、vinculin蛋白表达升高(均P<0.05),表明模型构建成功。与空白血清组比较,不同体积分数的芪术抗癌方含药血清组存活率降低(均P<0.05)。与空白血清组比较,10%、20%芪术抗癌方含药血清组伤口愈合率下降,各芪术抗癌方含药血清组侵袭细胞数均减少,各芪术抗癌方含药血清组黏附在细胞上的HCT116细胞数量均增加、黏附在基质凝胶上的HCT116细胞数量减少;与空白血清组比较,20%芪术抗癌方含药血清组E⁃cadherin蛋白表达水平升高(均P<0.05),PTP1B、vinculin、integrinανsubunit、integrinβ3 subunit蛋白表达降低。结论芪术抗癌方抑制结肠癌转移的作用机制可能与通过调控以PTP1B为核心介导的同、异源黏附串扰有关。Objective Exploring the mechanism of Qizhu Kangai Formula in regulating the key protein of protein tyrosine phosphatase 1B(PTP1B)of homo⁃heterologous adhesion crosstalk inhibition of colon cancer metastasis.Methods Prepare blank serum and Qizhu Kangai Formula containing serum.Analysis of effective chemical components in serum containing Qizhu Kangai Formula using liquid Journal of Beijing University of Traditional Chinese Medicine chromatography⁃mass spectrometry technology.Constructing PTP1B overexpression model(OE⁃PTP1B HCT116)in vitro through phasmid transfection.After detecting the cell survival rate of OE⁃PTP1B HCT116 using CCK⁃8 method,select 20%medicated serum and 5%、10%、20%Qizhu Kangai Formula containing serum.Scratch assay,transwell assay,and adhesion assay were used to detect the migration,invasion,and adhesion of cells overexpressing PTP1B,respectively.Western blotting was used to detect protein expression level of PTP1B,vinculin,E⁃cadherin,integrinανsubunit and integrinβ3 subunits.Results Liquid chromatography⁃mass spectrometry analysis revealed that the serum of Qizhu Kangai Formula contains 12 main effective chemical components.Among them,the highest content is Calycosin⁃7⁃glucoside.The expression levels of PTP1B and vinculin proteins were increased(P<0.05),indicating the successful construction of the model.Compared with the blank serum group,the survival rate of cells overexpressing PTP1B was decreased in different volume fractions of Qizhu Kangai Formula containing serum groups(P<0.05).Compared with the blank serum group,the wound healing rate of cells in the 10%and 20%Qizhu Kangai Formula containing serum groups decreased,the number of invasive cells in each containing serum group decreased,the number of HCT116 cells adhering to cells in each Qizhu Kangai Formula containing serum group increased,the number of HCT116 cells adhering to the matrigel decreased.Compared with the blank serum group,the expression level of E⁃cadherin protein was increased(P<0.05),the expressio

关 键 词：芪术抗癌方 结肠癌 蛋白酪氨酸磷酸酶1B 同源黏附 异源黏附 大鼠 

分 类 号：R285.5[医药卫生—中药学]