摘要
目的通过观察葛根素对肾透明细胞癌786-O细胞增殖与凋亡的影响,探讨其抑制786-O细胞生长的作用机制。方法分别以二甲基亚砜(空白对照)、不同浓度(10、20、40 mg/L)葛根素和70 mg/L 5-氟尿嘧啶干预对数生长期786-O细胞48 h,采用CCK-8法检测各组细胞增殖率,细胞克隆实验观察细胞克隆形成能力,流式细胞术分析细胞周期分布,AnnexinV-FITC/PI双染法检测细胞凋亡水平,Western blot法检测细胞周期和细胞凋亡相关蛋白表达水平。结果与空白对照相比,20、40 mg/L葛根素或5-氟尿嘧啶干预能够降低786-O细胞增殖率和细胞克隆数目(均P<0.01),提高处于G0/G1期细胞百分比并降低S期细胞百分比(均P<0.01),提高细胞凋亡率(均P<0.01),上调第10号染色体缺失的磷酸酶(PTEN)、激活型半胱氨酸天冬氨酸蛋白酶-3(Cleaved Caspase-3)、周期蛋白激酶抑制因子p27表达水平(均P<0.01),下调磷酸化磷脂酰肌醇3-激酶(p-PI3K)、磷酸化蛋白激酶B(p-Akt)、细胞周期蛋白D1(cyclin D1)表达水平(均P<0.05或0.01),降低PI3K、Akt磷酸化比值(均P<0.05或0.01)。与5-氟尿嘧啶干预相比,40 mg/L葛根素干预能够降低786-O细胞增殖率和细胞克隆数目(均P<0.01),提高G0/G1期细胞百分比并降低S期细胞百分比(P<0.05或0.01),提高细胞凋亡率(P<0.05),上调PTEN、Cleaved Caspase-3、p27表达水平并下调p-PI3K、p-Akt、cyclin D1表达水平(P<0.05或0.01),降低PI3K、Akt磷酸化比值(均P<0.05)。结论葛根素能够通过抑制细胞增殖和促进细胞凋亡而抑制肾透明细胞癌786-O细胞生长,可能与诱导PTEN表达而抑制PI3K/Akt通路有关。
Objective To investigate the inhibitory effect of puerarin on human renal clear cell carcinoma 786-O cells.Methods The cultured human renal clear cell carcinoma 786-O cells were treated with puerarin(0,10,20,40 mg/L)and 5-fluorouracil(70 mg/L)for 48 h.The cell proliferation rate was measured by CCK-8 method.Cell clone formation ability was determined by cell cloning assay.Cell cycle distribution was analyzed by flow cytometry.Cell apoptosis level was detected by AnnexinV-FITC/PI double staining method.Cell cycle-related and apoptosis-related protein expression were detected by Western blot.Results Compared with blank control group(0 mg/L puerarin),puerarin 20,40 mg/L or 5-fluorouracil 70 mg/L decreased the proliferation activity of 786-O cells and decreased the number of cell clones(P<0.01),increased the percentage of G0/G1 phase cells and decreased the percentage of S phase cells(P<0.01),increased the apoptosis rate(P<0.01),up-regulated the expression of PTEN,Cleaved Caspase-3 and cyclin kinase inhibitor p27(P<0.01),down-regulated the expression of p-PI3K,p-Akt and cyclin D1(P<0.05 or P<0.01),decreased the phosphorylation ratio of PI3K to Akt(P<0.05).Compared with 5-fluorouracil 70 mg/L group,the proliferation activity of 786-O cells,the number of cell clones and the apoptosis rate in puerarin 40 mg/L were higher(P<0.05 or P<0.01).The percentage of G0/G1 cells was higher and the percentage of S phase cells was lower(P<0.05 or P<0.01).The expression of PTEN,Cleaved Caspase-3,p27 was higher and the expression of p-PI3K,p-Akt,cyclin D1 was lower(P<0.05 or P<0.01),and phosphorylation ratio of PI3K to Akt was also lower(P<0.05).Conclusion Puer arin can inhibit the growth of 786-O cells by inhibiting proliferation and promoting apoptosis,which may be related to the inhibition of PI3K/Akt pathway by inducing PTEN expression.
作者
杨智勇
王晓英
陈永革
周莎莎
魏亚超
李晶
郭志远
祁卫华
YANG Zhiyong;WANG Xiaoying;CHEN Yongge;ZHOU Shasha;WEI Yachao;LI Jing;GUO Zhiyuan;QI Weihua(Second Department of Oncology,Handan Central Hospital,Handan 056001,China)
出处
《浙江医学》
CAS
2021年第9期930-935,I0004,共7页
Zhejiang Medical Journal
基金
河北省医学科学研究课题计划(20191843)。
作者简介
通信作者:杨智勇,E-mail:zhyshine@163.com。
