扇贝多肽通过调节c-jun和COX-2抑制UVA诱导的HaCaT细胞凋亡

目的复制UVA诱导的HaCaT细胞凋亡模型,研究UVA对细胞内c-jun和环氧合酶-2(cyclooxygenase-2,COX-2)的影响,从而探究扇贝多肽(Polypeptide from Chlamys far-reri,PCF)抑制UVA引起的HaCaT细胞凋亡的分子机制。方法实验分为5组:正常对照组、UVA模型组、UVA+5.69mmol.L-1PCF组、UVA+2.84mmol.L-1PCF组、UVA+1.42mmol.L-1PCF组。应用实时荧光定量PCR和蛋白印迹法检测细胞内c-jun的表达;RT-PCR结合蛋白质印迹法检测细胞内COX-2的表达;琼脂糖凝胶电泳分析PCF和COX-2特异性抑制剂celecoxib对UVA诱导HaCaT细胞凋亡的影响。结果预先加入PCF和celecoxib均可明显抑制8J.cm-2UVA诱导的HaCaT细胞凋亡;UVA照射HaCaT细胞后COX-2mRNA及蛋白表达水平增加,与对照组相比差异有显著性(P<0.01);1.42~5.69mmol.L-1剂量范围内的PCF可剂量依赖性抑制UVA引起的细胞内COX-2mR-NA及蛋白表达(P<0.05,P<0.01);PCF也抑制了UVA引起的HaCaT细胞内c-jun表达的增加,且呈量效关系(P<0.05,P<0.01)。结论UVA诱导HaCaT细胞发生凋亡时,细胞内COX-2和c-jun的表达明显增加,PCF通过抑制细胞内COX-2和c-jun的表达而发挥其抗凋亡作用。

Inhibition of UVA-induced apoptosis by the polypeptide from Chlamys farreri via regulation of c-jun and COX-2 expression in HaCaT cells

Aim A UVA-induced apoptotic model of HaCaT cells was established to investigate the impact of UVA on c-jun/cyclooxygenase-2(COX-2)and explore related molecular mechanism of the polypeptide from Chlamys farreri(PCF)protecting HaCaT cells from UVA-induced apoptosis.Methods Cells were divided into five groups:control group,UVA model group,UVA+5.69 mmol·L-1 PCF group,UVA+2.84 mmol·L-1 PCF group,UVA+1.42 mmol·L-1 PCF group.Expression level of c-jun was assayed by Real-Time PCR and Western blot.RT-PCR and Western blot analysis were used to determine the mRNA and protein levels of COX-2.Using agarose gel electrophoresis,the effects of PCF and COX-2 inhibitor celecoxib on UVA-induced apoptosis were also investigated.Results PCF and celecoxib had inhibitory effect on 8 J·cm-2 UVA-induced apoptosis of HaCaT cells.COX-2 mRNA and protein levels increased after UVA radiation and the discrepancy was significant compared with control group(P<0.01).1.42~5.69 mmol·L-1 PCF dose-dependently inhibited UVA-induced expression of COX-2 in HaCaT cells(P<0.05,P<0.01).PCF also attenuated UVA-induced expression of c-jun in a dose-dependent manner(P<0.05,P<0.01).Conclusion The expression levels of c-jun and COX-2 in HaCaT cells significantly increased in UVA-induced apoptosis;PCF could attenuate UVA-induced c-jun and COX-2 expression which might attribute to its anti-apoptotic effect.