基于miR-1297/PTEN/PI3K/AKT通路探讨芪蓝方影响前列腺癌DU145细胞增殖及凋亡机制  

Mechanism of Qilan Decoction's Effect on the Proliferation and Apoptosis of Prostate Cancer DU145 Cells Based on the miR-1297/PTEN/PI3K/AKT Signaling Pathway

作　　者：原凡 景明夷 周静 王家灏 常德贵[1,2] 尤耀东 YUAN Fan;JING Ming-yi;ZHOU Jing;WANG Jia-hao;CHANG De-gui;YOU Yao-dong(Urology and Andrology of Hospital of Chengdu University of Traditional Chinese Medicine,Chengdu,610072;TCM Regulating Metabolic Diseases Key Laboratory of Sichuan Province,Chengdu,610072;Clinical Medicine School of Chengdu University of Traditional Chinese Medicine,Chengdu,610075)

机构地区：[1]成都中医药大学附属医院泌尿男科,成都610072 [2]代谢性疾病中医药调控四川省重点实验室,成都610072 [3]成都中医药大学临床医学院,成都610075

出　　处：《中国中西医结合杂志》2024年第7期847-855,共9页Chinese Journal of Integrated Traditional and Western Medicine

基　　金：国家自然科学基金面上项目(No.81973866);四川省科技厅自然科学基金项目(No.2022NSFSC0684);四川省中医药管理局科学技术研究专项(No.2021ZD016);成都中医药大学“杏林学者”学科人才科研提升计划研究专项(No.QNXZ2019020)。

摘　　要：目的 探究芪蓝方影响前列腺癌细胞增殖及凋亡的可能作用机制。方法 将DU145细胞分为空白组(NC mimics)、芪蓝方组(NC mimics+芪蓝方)、miR-1297过表达组(miR-1297 mimics)、miR-1297过表达+芪蓝方组(miR-1297 mimics+芪蓝方)、AKT激动剂组(SC79)、AKT激动剂+芪蓝方组(SC79+芪蓝方)。CCK-8法、EdU法检测各组DU145细胞增殖;流式细胞术检测各组细胞周期及凋亡情况;RT-PCR检测miR-1297、人第10号染色体缺失的磷酸酶及张力蛋白同源的基因(PTEN)、蛋白激酶B (AKT) mRNA表达;Western Blot检测PTEN、磷脂酰肌醇-3激酶(PI3K)、p-AKT、Cyclin D1、B细胞淋巴瘤/白血病-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、Cleaved-含半胱氨酸的天冬氨酸蛋白水解酶(caspase)3蛋白表达。结果 与空白组比较,芪蓝方组DU145细胞活性、增殖率、EdU阳性细胞率、G1期细胞比例、miRNA 1297 mRNA表达、PI3K、p-AKT、Cyclin D1、Bcl-2蛋白表达降低,G2期细胞比例、Q1-LR(早期凋亡)+Q1-UR(晚期凋亡)、PTEN mRNA表达、PTEN、Bax、Cleaved-caspase 3蛋白表达升高(P<0.01);miR-1297过表达组和AKT激动剂组DU145细胞活性、增殖率、PI3K、p-AKT、Cyclin D1、Bcl-2蛋白表达升高,Q1-LR+Q1-UR、PTEN蛋白表达降低(P<0.01)。与miR-1297过表达组比较,miR-1297过表达+芪蓝方组DU145细胞活性、增殖率、G1期细胞比例、miRNA 1297 mRNA表达、PI3K、p-AKT、Cyclin D1、Bcl-2蛋白下降,G2期细胞比例、Q1-LR+Q1-UR、PTEN mRNA表达、PTEN、Bax、Cleaved-caspase 3蛋白表达升高(P<0.01)。与AKT激动剂组比较,AKT激动剂+芪蓝方组DU145细胞活性、增殖率、EdU阳性细胞率、S期细胞比例、PI3K、p-AKT、Cyclin D1、Bcl-2蛋白表达下降,G1、G2细胞比例、Q1-LR+Q1-UR、PTEN mRNA表达、PTEN、Bax、Cleaved-caspase 3蛋白表达升高(P<0.01)。结论 芪蓝方可通过调节miR-1297/PTEN/PI3K/AKT通路抑制DU145细胞增殖和促进细胞凋亡。Objective To explore the possible mechanism of Qilan Decoction's effect on the proliferation and apoptosis of prostate cancer cells.Methods DU145 cells were divided into negative control group(NC mimics),Qilan Decoction group(NC mimics+QLF),miR-1297 over-expression group(miR-1297 mimics),miR-1297 over-expression+Qilan Decoction group(miR-1297 mimics+QLF),AKT activator group(SC79),AKT activator+Qilan Decoction group(SC79+QLF).CCK-8 method and EdU experiment were used to detect cell proliferation in each group.Flow cytometry were used to detect cell cycle and apoptosis in each group.RTPCR was used to detect the mRNA expression of miR-1297,phosphatase and tensin homologue deleted on chromosome 10(PTEN),and protein kinase B(AKT).Western Blot was used to detect the expression of PTEN,phosphatidylinositol-3 kinase(PI3K),p-AKT,Cyclin D1,B-cell lymphoma/leukemia 2(Bcl-2),Bcl-2 related X protein(Bax),and Cleaved-caspase 3.Results Compared with negative control group,the activity,proliferation rate,EdU positive cell rate,G1 phase cell proportion,miRNA 1297 mRNA protein expression,PI3K,p-AKT,Cyclin D1,and Bcl-2 protein expression of DU145 cells in the Qilan Decoction group decreased,while the G2 phase cell proportion,Q1-LR(early apoptosis)+Q1-UR(late apoptosis),PTEN mRNA expression,PTEN,Bax,and Cleaved-caspase 3 protein expression increased(P<0.01).The miR-1297 over-expression group and AKT activator group showed an increase in DU145 cell activity,proliferation rate,PI3K,p-AKT,Cyclin D1,and Bcl-2 protein expression,while the expression of Q1-LR+Q1-UR,PTEN proteins decreased(P<0.01).Compared with the miR-1297 over-expression group,miR-1297 over-expression group+Qilan Decoction showed a decrease in DU145 cell activity,proliferation rate,G1 phase cell proportion,miRNA 1297 mRNA expression,PI3K,p-AKT,Cyclin D1,and Bcl-2 proteins,while the G2 phase cell proportion,Q1-LR+Q1-UR,PTEN mRNA expression,PTEN,Bax,and Cleaved-caspase 3 protein expression increased(P<0.01).Compared with the AKT activator group,AKT activator+Qilan Decoctio

关 键 词：芪蓝方 前列腺癌 DU145细胞 增殖 凋亡 miR-1297 PTEN/PI3K/AKT通路 中药复方 

分 类 号：R285[医药卫生—中药学]