穿心莲内酯对口腔鳞状细胞癌体内外抑制作用的实验研究

Experimental study of andrographolide on oral squamous cell carcinoma in vivo and in vitro

目的:研究穿心莲内酯在体内外对口腔鳞状细胞癌(OSCC)生长的抑制作用。方法:穿心莲内酯浓度梯度处理CAL-27细胞,CCK8检测OSCC细胞活力,流式细胞术检测细胞周期,克隆形成实验检测克隆形成率,RT-PCR检测PCNA和P21表达水平,Hoechst观察细胞形态,Western blot检测caspase-9、caspase-3、cleaved PARP、E-cadherin、N-cadherin、Vimentin和TGF-β1蛋白表达水平。OSCC细胞系CAL-27皮下注射裸鼠建立移植瘤裸鼠模型,检测肿瘤重量,免疫组化检测Ki67、caspase-3和Vimentin表达水平。结果:与0μmol/L穿心莲内酯组比较,15μmol/L、30μmol/L穿心莲内酯处理组细胞活力显著降低(P<0.05),克隆形成率显著降低(P<0.05),PCNA mRNA表达水平下调,P21 mRNA表达水平显著上调(P<0.05),caspase-9、caspase-3蛋白表达水平升高(P<0.05),N-cadherin和Vimentin蛋白表达水平显著降低(P<0.05),E-cadherin蛋白表达水平显著升高(P<0.05)。与对照组比较,50 mg/kg和100 mg/kg穿心莲内酯组肿瘤重量明显下降(P<0.05),caspase-3细胞阳性率显著增高,Ki67和Vimentin细胞阳性率显著减少(P<0.05)。结论:穿心莲内酯在体内外均能抑制OSCC细胞生长。

Objective:To study the inhibitory effect of andrographolide on the growth of oral squamous cell carcinoma(OSCC in vivo and in vitro.Methods:The concentration gradient of andrographolide was used to treat CAL-27 cells,CCK8 was used to detect the viability of OSCC,cell cycle was detected by flow cytometry,the clone formation test was used to detect the clone formation rate,the expression levels of PCNA and P21 mRNA were detected by RT-PCR,the cell morphology was observed by Hoechst,and the Western blot was performed to test the expression levels of caspase-9,caspase-3,cleaved PARP,E-cadherin,N-cadherin,Vimentin and TGF-β1 protein.OSCC cell line CAL-27 was subcutaneously injected into nude mice to establish a nude mouse model of transplanted tumor,the tumor weight was measured,and the expression levels of Ki67,caspase-3 and Vimentin were detected by immunohistochemistry.Results:Compared with the 0μmol/L andrographolide group,the cell viability of the 15μmol/L and 30μmol/L andrographolide treatment groups was significantly reduced(P<0.05),the clone formation rate was significantly reduced(P<0.05),the PCNA mRNA expression level was down-regulated,and the P21 mRNA expression level was significantly up-regulation(P<0.05),caspase-9 and caspase-3 protein expression levels was increased(P<0.05),N-cadherin and Vimentin protein expression levels were significantly reduced(P<0.05),E-cadherin protein expression level was significantly increased(P<0.05).Compared with the control group,the tumor weight in the 50 mg/kg and 100 mg/kg andrographolide groups were significantly reduced(P<0.05),the positive rate of caspase-3 cells was significantly increased,and the positive rate of Ki67 and Vimentin cells were significantly reduced(P<0.05).Conclusion:Andrographolide can inhibit the growth of OSCC cells in vivo and in vitro.