柚皮素介导Ras同源基因家族成员A/Rho关联含卷曲螺旋结合蛋白激酶1通路对人结直肠癌HT-29/奥沙利铂耐药细胞株的影响  

Effect of naringin-mediated RhoA/ROCK1 pathway on oxaliplatin resistant human colorectal cancer cell line HT-29/L-OHP

作　　者：郑才勇 于冬 李永春[1] ZHENG Caiyong;YU Dong;LI Yongchun(Department of General Surgery,Mianyang People's Hospital,Mianyang,Sichuan 621053)

机构地区：[1]四川省绵阳市人民医院普外科,四川绵阳621053

出　　处：《河北中医》2024年第9期1501-1505,1511,共6页Hebei Journal of Traditional Chinese Medicine

摘　　要：目的探讨柚皮素介导Ras同源基因家族成员A(RhoA)/Rho关联含卷曲螺旋结合蛋白激酶1(ROCK1)通路对人结直肠癌细胞株HT-29对奥沙利铂(L-OHP)化疗敏感性的作用。方法取人结直肠癌细胞株HT-29,建立稳定的耐药细胞株HT-29/L-OHP,接种至96孔板中,调整细胞浓度至1.0×10^(5)个/mL,设置4组,分别为空白组、L-OHP组、柚皮素组、柚皮素+L-OHP组,每组6个复孔。L-OHP组予以5μmol/L L-OHP 2μL,柚皮素组予以100μmol/L柚皮素2μL,柚皮素+L-OHP组予以100μmol/L柚皮素2μL+5μmol/L L-OHP 2μL,空白组仅予以无菌0.9%氯化钠注射液。噻唑兰(MTT)法检测各组细胞活力和半抑制浓度(IC50);实时荧光定量聚合酶链式反应(RT-qPCR)检测各组细胞RhoA、ROCK1、多药耐药相关蛋白1(MRP1)、抗凋亡因子B细胞淋巴瘤-2(Bcl-2)mRNA表达;蛋白免疫印迹法检测各组细胞RhoA、ROCK1、MRP1、Bcl-2蛋白表达。结果成功构建人结直肠癌耐药细胞株HT-29/L-OHP;与空白组和L-OHP组比,柚皮素组和柚皮素+L-OHP组细胞增殖能力下降,IC50值降低,RhoA、ROCK1、MRP1、Bcl-2 mRNA及蛋白表达降低,比较差异均有统计学意义(P<0.05),且空白组上述指标与L-OHP组比较、柚皮素组上述指标与柚皮素+L-OHP组比较差异均无统计学意义(P>0.05)。结论柚皮素能显著降低人结直肠癌耐药细胞株HT-29/L-OHP的增殖能力,提高其对L-OHP的化疗敏感性,推测与抑制RhoA、ROCK1、MRP1、Bcl-2的表达有关。Objective To investigate the effect of naringin-mediated Ras homologous gene family member A(RhoA)/Rho associated coiled coil binding protein kinase 1(ROCK1)pathway on enhancing the sensitivity of human colorectal cancer cell line HT-29 to oxaliplatin(L-OHP)chemotherapy.Methods Human colorectal cancer cell line HT-29 with a stable resistance to L-OHP(HT-29/L-OHP)was established.HT-29/L-OHP cells were inoculated to 96-well plates,with the cell density of 1.0×10^(5)/mL.Cells were induced with blank control(injection of sterile 0.9%NaCl 2μL),5μmol/L L-OHP 2μL,100μmol/L naringin 2μL and 5μmol/L L-OHP 2μL+100μmol/L naringin 2μL,with 6 replicates.The cell viability and the half maximal inhibitory concentration(IC50)of each group were detected by methyl thiazolyl tetrazolium(MTT)assay.Real-time reverse transcription quantitative polymerase chain reaction(RT-qPCR)was used to detect the messenger RNA(mRNA)expressions of RhoA,Rock1,multidrug resistance associated protein 1(MRP1)and anti apoptotic factor(Bcl-2).The protein expressions of RhoA,Rock1,MRP1 and Bcl-2 were detected by Western blot(WB).Results Human colorectal cancer resistant cell line HT-29/L-OHP was successfully constructed.Compared with those treated with blank control or L-OHP group,cells treated with naringin or naringin+L-OHP had significantly lower cell proliferation ability,mRNA and protein expressions of RhoA,Rock1,MRP1 and Bcl-2,and IC50(P<0.05).There were no significant differences in the above indicators between the blank group versus L-OHP group,and between naringin group versus naringin+L-OHP group(P>0.05).Conclusion Naringin can significantly reduce the proliferation of L-OHP-resistant human colorectal cancer resistant cell line HT-29/L-OHP and improve its chemosensitivity to L-OHP by downregulating RhoA,Rock1,MRP1 and Bcl-2.

关 键 词：结直肠肿瘤 柚皮苷 实验 

分 类 号：R574.62[医药卫生—消化系统] R574.63[医药卫生—内科学] R-33[医药卫生—临床医学]