扶正化瘀解毒汤调控ITGB3-Ras轴对人前列腺癌细胞生物学行为的影响  

Effect of Fuzheng Huayu Jiedu Decoction on Biological Behaviors of Human Prostate Cancer Cells by Regulating ITGB3-Ras Axis

作　　者：周仕轶 尤耀东[2] 赵涛 龚艳菊 张国臣 邵继春 Zhou Shiyi;You Yaodong;Zhao Tao;Gong Yanju;Zhang Guochen;Shao Jichun(Basic Medical School of Chengdu University of Traditional Chinese Medicine,Chengdu,611137,China;Clinical Medical School of Chengdu University of Traditional Chinese Medicine,Chengdu,610075,China;Department of Urology,Second Affiliated Hospital of Chengdu Medical University/416 Hospital of Nuclear Industry,Chengdu,610051,China)

机构地区：[1]成都中医药大学基础医学院,四川成都611137 [2]成都中医药大学临床医学院,四川成都610075 [3]成都医学院第二附属医院/核工业四一六医院泌尿外科,四川成都610051

出　　处：《成都中医药大学学报》2023年第2期29-38,共10页Journal of Chengdu University of Traditional Chinese Medicine

基　　金：国家自然科学基金面上项目(81973866);四川省科技厅应用基础研究计划项目(2018JY0368);四川省中医药管理局中医药科研专项(2018JC035)。

摘　　要：目的:探讨扶正化瘀解毒汤通过调控ITGB3-Ras轴对前列腺癌(Prostate Cancer,PCa)的肿瘤细胞增殖、侵袭和迁移等肿瘤生物学行为的影响。方法:RM-1细胞建立PCa体内荷瘤模型,该方灌胃观察对移植瘤的生长抑制效应。该方含药血清处理人PCa细胞系LNCaP、DU-145、PC-3细胞以及正常前列腺细胞系WPMY-1,MTT检测细胞增殖,集落形成实验观察细胞集落形成,Transwell小室实验观察细胞侵袭和迁移能力,Western blot和RT-qPCR检测含药血清对ITGB3-Ras轴的调控作用,shRNA和过表达验证含药血清的作用靶点。iTRAQ分析检测该方含药血清对PC-3细胞ITGB3-Ras轴主要蛋白表达调控作用。高效液相色谱法(HPLC)测定该方的有效成分含量。结果:该方体内可明显抑制RM-1细胞小鼠的皮下移植肿瘤体积(P<0.001)和重量(P<0.01)。含药血清抑制了PC-3和DU-145的细胞生长活力和集落形成,并呈剂量依赖性。Transwell小室实验显示,低浓度含药血清可抑制PC-3和DU-145的细胞的侵袭和迁移能力。iTRAQ蛋白组学分析,经过含药血清干预的PC-3细胞,ITGB3-Ras轴表达水平明显与未经处理的PC-3细胞降低。高转移潜能的PC-3细胞与不具有转移潜能的LNCaP细胞相比,ITGB3和Ras蛋白和ITGB3 mRNA表达水平更高,表明ITGB3-Ras对PCa肿瘤进展特别是转移特性获得具有重要作用。shRNA沉默ITGB3后,ITGB3促上皮-间充质转化(EMT)能力降低,PC-3和DU-145细胞转移能力降低,这与含药血清的作用效果相似。该方提取物中有效成分含有姜黄素(0.79 mg/mL)、迷迭香酸(0.16 mg/mL)和人参皂苷R g3(68.4μg/mL)。结论:扶正化瘀解毒汤可抑制具有转移特性的人PCa的PC-3和DU-145细胞肿瘤生物学行为,其可能机制是调控细胞ITGB3-Ras轴表达,抑制EMT转化,从而降低细胞增殖、侵袭、迁移和体内成瘤的能力。Objective:To investigate the effect of Fuzheng Huayu Jiedu Decoction on the proliferation,invasion and migration of prostate cancer(PCa)cells by regulating ITGB3-Ras axis.Methods:RM-1 cells were used to establish the tumor-bearing model of PCa in vivo,and the growth inhibition effect of Fuzheng Huayu Jiedu Decoction on PCa was observed by Gavage.The LNCaP,DU-145,PC-3 cells and normal prostate cell line WPMY-1 were treated with Fuzheng Huayu Jiedu Decoction serum.The proliferation of the cells was detected by MTT and the formation of the cell colonies was observed by colony-forming assay,transwell 24-well chamber assay was used for in vitro cell migration and invasion capacity.Western blot and RT-qPCR were used to detect the regulation of ITGB3-Ras axis,and shRNA or shRNA overexpression were used to identify the targets of the serum.The iTRAQ analysis detected the regulation of major protein expression in PC-3 cell ITGB3-Ras axis by drug-containing serum.The content of the active ingredients was determined by High-performance liquid chromatography(HPLC).Results:Fuzheng Huayu Jiedu Decoction could significantly inhibit the subcutaneous graft tumor volume(P<0.001)and the weight(P<0.01)of transplanted RM-1 cells in mice.Drug-containing serum inhibited the growth viability and colony formation of PC-3 and DU-145 cells in a dose-dependent manner.The transwell chamber assay showed that low concentrations of drug-containing serum inhibited the invasion and migration capacity of PC-3 and DU-145 cells.By iTRAQ proteomics analysis showed that the expression level of ITGB3-Ras axis in PC-3 cells with serum intervention was significantly lower than that in untreated PC-3 cells.The ITGB3 proteins and Ras proteins and ITGB3 mRNA expression levels were higher in PC-3 cells with high metastatic potential than in LNCaP cells without metastatic potential,it is suggested that ITGB3-RAS plays an important role in the progression of PCa,especially in the acquisition of metastatic characteristics.After silencing ITGB3 by shRNA,the abil

关 键 词：扶正化瘀解毒汤 前列腺癌 整合素Β3 RAS基因 增殖 侵袭 迁移 

分 类 号：R737.25[医药卫生—肿瘤] R289[医药卫生—临床医学]