龙胆苦苷对肿瘤坏死因子-α诱导的HaCaT细胞增殖的抑制作用

Inhibition of Tumor Necrosis Factor-α-induced HaCaT cell Proliferation by Gentiopicroside

目的通过观察龙胆苦苷对肿瘤坏死因子(TNF)-α诱导的人永生化角质形成细胞(HaCaT细胞)增殖、凋亡及血管内皮生长因子(VEGF)A、角蛋白(K)17表达的影响,探讨龙胆苦苷治疗角质形成细胞异常增生性疾病的可能机制。方法通过噻唑兰(MTT)实验和流式细胞术检测对照组、模型组、氨甲蝶呤(MTX)组及龙胆苦苷治疗组对TNF-α诱导的HaCaT细胞增殖和细胞凋亡的影响;此外通过荧光定量聚合酶链反应(RT-PCR)法检测VEGF A、K17的表达。结果与对照组比较,模型组HaCaT细胞存活率、VEGF A、K17 mRNA显著升高(P<0.01),细胞凋亡率明显下降(P<0.01);与模型组相比,MTX组、龙胆苦苷(100、200、400 ng/mL)治疗组细胞存活率、K17 mRNA表达量明显下降(P<0.01),细胞凋亡率明显上升(P<0.05);MTX组、龙胆苦苷(200、400 ng/mL)治疗组VEGF A mRNA表达量下降(P<0.01);与MTX组相比,龙胆苦苷(100、200 ng/mL)治疗组抑制TNF-α诱导的HaCaT细胞增殖作用比较差异无统计学意义(P>0.05),400 ng/mL治疗组抑制作用优于MTX组(P<0.05);龙胆苦苷治疗组促进细胞凋亡、抑制HaCaT细胞VEGF A、K17 mRNA的表达差异无统计学意义(P>0.05)。结论龙胆苦苷可以抑制TNF-α诱导的HaCaT细胞的增殖,促进HaCaT细胞凋亡,降低VEGF A、K17 mRNA的表达,可以作为治疗角质形成细胞异常增生性疾病的潜在药物。

Objective In order to investigate the possible mechanism of gentiopicroside in the treatment of keratinocyte dysproliferative diseases by observing the effects of gentiopicroside on tumor necrosis factor(TNF)-αinduced HaCaT cell proliferation,apoptosis,and expression of vascular endothelial growth factor(VEGF)A and keratin(K)17.Methods 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenytetrazoliumromide(MTT)experiment and flow cytometry were used to detect the effects of control group,model group,methotrexate group and gentiopicroside treatment group on TNF-α-induced HaCaT cell proliferation and apoptosis.In addition,the expression of VEGF A and K17 was detected by reverse transcription-polymerase chain reaction(RT-PCR)method.Results Compared with the control group,the HaCaT cell survival rate,VEGF A and K17 mRNA in the model group were significantly increased(P<0.01),and the apoptosis rate was significantly decreased(P<0.01).Compared with the model group,theMTX group and the gentiopicroside treatment group(100,200,400 ng/mL)cell survival rate,K17 mRNA expression decreased significantly(P<0.01),cell apoptosis rate increased significantly(P<0.05).The expression of VEGF A mRNA in the MTX group and the gentiopicroside treatment group(200,400 ng/mL)decreased(P<0.01).Compared with the MTX group,the gentiopicroside treatment group(100,200 ng/mL)had no statistical significance in inhibition of the proliferation of TNF-α-induced HaCaT cells(P>0.05),the inhibitory effect of 400 ng/mL treatment group was better than MTX group(P<0.05).Gentiopicroside treatment group promoted apoptosis and inhibited expression of VEGF A and K17 mRNA in HaCaT cells without statistical significance(P>0.05)Conclusion Gentiopicroside can inhibit the proliferation of HaCaT cells induced by TNF-α,promote the apoptosis of HaCaT cells,and reduce the expression of VEGF A and K17 mRNA.It can be used as a potential drug for the treatment of keratinocyte dysproliferative diseases.