论文部分内容阅读
目的探讨RNA激活p21对肝癌HepG2、Hep3b和SMMC-7721细胞生长和侵袭力的影响。方法化学合成靶向p21的saRNA、阴性对照dsRNA,将其转染肝癌细胞系HepG2、Hep3b和SMMC-7721。每个细胞系分为3组,分别为p21-322组、阴性对照组和空白对照组,每组复3孔;p21-322组和阴性对照组分别采用p21-322saRNA和阴性对照dsRNA进行转染,空白对照组不干预。转染后72h,采用RT-PCR和Western blot检测各组细胞中的p21 mRNA和P21蛋白表达水平,采用MTT法检测细胞生长情况及划痕实验观察细胞侵袭能力的变化。结果HepG2、Hep3b和SMMC-7721细胞p21-322组p21mRNA相对表达水平分别为23.43±2.29、16.87±1.61、31.77±5.06,P21蛋白相对表达水平分别为55.93±12.66、32.91±5.17、24.96±6.81;空白对照组分别为3.53±0.07、2.39±0.02、5.70±0.89,3.21±0.03、2.91±0.14、4.15±0.12;阴性对照组分别为3.87±0.97、2.57±0.71、5.87±1.73,3.11±0.70、3.01±0.97、5.13±2.14;p21-322组p21mRNA和蛋白相对表达水平明显高于空白对照组和阴性对照组(P<0.01),空白对照组与阴性对照组比较差异无统计学意义(P>0.05);细胞转染后第6天时HepG2、Hep3b、SMMC-7721细胞p21-322转染组细胞平均生长抑制率分别为41%、48%和52%;HepG2细胞p21-322组24h后空白区域占原划痕区域面积的百分比((76±11)%)明显高于空白对照组((13±6)%)和阴性对照组((17±8)%)(P<0.01),阴性对照组与空白对照组比较差异无统计学意义(P>0.05)。结论靶向p21的RNAa能抑制肝癌细胞的生长和侵袭力,p21可作为一个具有肝癌治疗应用价值的靶基因。
Objective To investigate the effects of p21 activated by RNA on the growth and invasiveness of HepG2, Hep3b and SMMC-7721 cells. Methods The saRNA targeting p21 and the negative control dsRNA were chemically synthesized and transfected into HepG2, Hep3b and SMMC-7721 cells. Each cell line was divided into three groups, p21-322 group, negative control group and blank control group, each group complex 3 wells; p21-322 group and negative control group were transfected with p21-322saRNA and negative control dsRNA , The blank control group did not interfere. The expression of p21 mRNA and p21 protein in each group was detected by RT-PCR and Western blot 72 h after transfection. The cell growth was detected by MTT assay and the invasion ability of cells was observed by scratch test. Results The relative expression levels of p21 mRNA in p21-322 group were 23.43 ± 2.29, 16.87 ± 1.61 and 31.77 ± 5.06 respectively, and the relative expression levels of P21 protein in HepG2, Hep3b and SMMC-7721 cells were 55.93 ± 12.66, 32.91 ± 5.17 and 24.96 ± 6.81, respectively. The blank control group was 3.53 ± 0.07,2.39 ± 0.02,5.70 ± 0.89,3.21 ± 0.03,2.91 ± 0.14,4.15 ± 0.12 respectively; the negative control group was 3.87 ± 0.97,2.57 ± 0.71,5.87 ± 1.73,3.11 ± 0.70 respectively, 3.01 ± 0.97 and 5.13 ± 2.14 respectively. The relative expression level of p21 mRNA and protein in p21-322 group was significantly higher than that in blank control group and negative control group (P <0.01), but there was no significant difference between blank control group and negative control group (P> 0.05). The average cell growth inhibition rates of HepG2, Hep3b and SMMC-7721 cells transfected with p21-322 were 41%, 48% and 52% respectively on the 6th day after transfection; in p21-322 HepG2 cells, the blank area (76 ± 11)% of the original scratched area was significantly higher than that of the blank control group (13 ± 6%) and the negative control group (17 ± 8%) (P <0.01). The negative control There was no significant difference between the group and the blank control group (P> 0.05). Conclusion RNAi targeting p21 can inhibit the growth and invasiveness of hepatocellular carcinoma cells. P21 can be used as a target gene with therapeutic value for hepatocellular carcinoma.