Hypocrellin A通过ROS/JNK通路促进瘢痕疙瘩成纤维细胞凋亡的作用及机制

The effect and mechanism of Hypocrellin A promotes apoptosis of Keloid fibroblasts through ROS/JNK pathway

目的:探索Hypocrellin A(HA)通过活性氧簇(ROS)/c-Jun氨基末端激酶(JNK)通路促进瘢痕疙瘩成纤维细胞凋亡的作用及机制。方法:原代培养KFs并进行传代,第3代KFs分为对照组、不同剂量HA组(0.125、0.25、0.5、1.0μmol/L HA处理)、HA+乙酰半胱氨酸(NAC)组(1.0μmol/L HA+1.0 mmol/L NAC处理)。检测细胞增殖的光密度OD450,凋亡率,胶原代谢指标I型胶原(Col-I)、α-平滑肌肌动蛋白(α-SMA)、纤维连接蛋白(FN)、氧化应激指标丙二醛(MDA)、超氧化物歧化酶(SOD)、总抗氧化力(T-AOC)及ROS的含量,内质网应激标志物GRP78、p-IRE-1及ROS/JNK通路中p-JNK、cleaved caspase-3的表达水平。结果:不同剂量HA组的OD450、Col-I、α-SMA、FN、SOD、T-AOC的含量均低于对照组,凋亡率、MDA、ROS含量及GRP78、p-IRE-1、p-JNK、cleaved caspase-3的表达水平均高于对照组(P<0.05)且HA剂量越高,上述指标的变化越显著;HA+NAC组的OD450、Col-I、α-SMA、FN、SOD、T-AOC的含量均高于1.0μmol/L HA组,凋亡率、MDA、ROS含量及GRP78、p-IRE-1、p-JNK、cleaved caspase-3的表达水平均低于1.0μmol/L HA(P<0.05)。结论:HA通过促进KFs凋亡,这一作用与激活ROS/JNK通路介导的氧化应激和内质网应激有关。

Objective:To explore the role and mechanism of Hypocrellin A(HA)in promoting apoptosis of Keloid fibroblasts(KFs)through reactive oxygen species(ROS)/c-Jun N-terminal kinase(JNK)pathway.Methods:KFs were primarily cultured and passaged.The 3rd generation KFs were divided into control group and different dose HA groups(0.125,0.25,0.5,1.0μmol/L HA treatment),HA+N-acetylcysteine(NAC)group(1.0μmol/L HA+1.0 mmol/L NAC treatment).The optical density OD450 of cell proliferation,apoptosis rate,the contents of collagen metabolism indicators type I collagen(Col-I),α-smooth muscle actin(α-SMA),fibronectin(FN),oxidative stress indicators malondialdehyde(MDA),superoxide dismutase(SOD),total antioxidant capacity(T-AOC)and ROS,and the expression levels of endoplasmic reticulum stress markers GRP78,p-IRE-1 and p-JNK,cleaved caspase-3 in the ROS/JNK pathway were detected.Results:The OD450 level,the contents of Col-Iα-SMA,FN,SOD,T-AOC in the control group were lower than those in the control group,while the apoptosis rate,the contents of MDA,ROS,and the expression levels of GRP78,p-IRE-1,p-JNK and cleared caspase-3 were higher than those in the control group(P<0.05).The higher the HA dose,the more significant the changes in the above indicators.The OD450 level,the contents of Col-Iα-SMA,FN,SOD,T-AOC in the control group were lower than those in the HA+NAC group were higher than those in the 1.0μmol/L HA group,while the apoptosis rate,the contents of MDA,ROS,and the expression levels of GRP78,p-IRE-1,p-JNK and cleared caspase-3 were lower than those in the 1.0μmol/L HA group(P<0.05).Conclusion:HA promotes the apoptosis of KFs,which is related to the activation of ROS/JNK pathway mediated oxidative stress and endoplasmic reticulum stress.