扇贝多肽通过调节c-jun和COX-2抑制UVA诱导的HaCaT细胞凋亡

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

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。

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 polypeptidefrom Chlamys farreri （PCF） protecting HaGaT ceils from UVA-induced apoptosis. Methods Ceils were divided into five groups： control group, UVA model group, UVA ＋ 5. 6 9 mmol · L^-1 PCF group, UVA ＋2.84 mmol· L^-1 PCF group, PCF group. Expression level Real-Time PCR and Western ern blot analysis were used to UVA ＋ 1.42 mmol· L^-1 of c-jun was assayed by blot. RT-PCR and Westdetermine the mRNA and protein levels of COX-2. Using agarose gel electropho- resis, the effects of PCF and COX-2 inhibitor celecoxib on UVA-induced apoptosis were also in,instigated. Resuits PCF and celecoxib had inhibitory effect on 8 J · cm^-2 UVA-induced apoptosis of HaCaT ceils. 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^-1PCF 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 cjun and COX-2 expression which might attribute to its anti-apoptotic effect.