长链非编码LINC00312通过内质网IRE1-JNK途径促进病理性瘢痕成纤维细胞凋亡  

Long Non-coding RNA LINC00312 Promotes Apoptosis of Pathological Scar Fibroblasts Via the Endoplasmic Reticulum IRE1-JNK Pathway

作　　者：李涛 林简 王荣坤 李娜 LI Tao;LIN Jian;WANG Rongkun;LI Na(Department of Dermatology&STD,Sanya People′s Hospital,Sanya 572000,China)

机构地区：[1]三亚市人民医院皮肤性病科,海南三亚572000

出　　处：《中国皮肤性病学杂志》2020年第11期1221-1227,共7页The Chinese Journal of Dermatovenereology

摘　　要：目的探讨长链非编码RNA LINC00312(LncRNA LINC00312)对人病理性瘢痕成纤维细胞凋亡的影响及其可能机制。方法体外分离培养人病理性瘢痕成纤维细胞,并采用LINC00312 shRNA慢病毒和阴性对照慢病毒感染细胞,qRT-PCR法检测感染前后细胞中LINC00312的表达水平。根据实验需求,将细胞分为4组:空白对照组(Blank)、阴性对照慢病毒组(NC-shRNA)、LINC00312 shRNA慢病毒组(LINC00312-shRNA)及IRE1α特异性抑制剂组(LINC00312-shRNA+KIRA6)。采用CCK-8法检测各组细胞增殖情况;Hoechst染色法观察各组细胞形态变化;Annexin V-FITC/PI法检测各组细胞凋亡情况;Western blot检测各组细胞凋亡相关蛋白cleaved Caspase-3、Bax、Bcl-2以及内质网应激相关蛋白GRP78、IRE1、p-IRE1、JNK、p-JNK的表达水平。结果LINC00312 shRNA慢病毒感染后细胞中LINC00312的表达水平显著降低(P<0.05);与Blank组和NC-shRNA组比较,LINC00312-shRNA组细胞增殖活性明显降低,细胞核呈大量碎块致密浓染,凋亡水平明显升高,cleaved Caspase-3、Bax、GRP78、p-IRE1、p-JNK等蛋白表达水平明显升高,Bcl-2蛋白表达水平明显降低(P<0.05);与LINC00312-shRNA组比较,LINC00312-shRNA+KIRA6组细胞增殖活性明显升高,细胞核的碎块致密浓染数量减少,凋亡水平明显降低,cleaved Caspase-3、Bax、GRP78、p-IRE1、p-JNK蛋白表达水平明显降低,而Bcl-2蛋白表达水平明显升高(P<0.05)。结论沉默LINC00312可能通过激活内质网IRE1-JNK介导的凋亡途径,并诱导人病理性瘢痕成纤维细胞凋亡。Objective To investigate the effects of long non-coding RNA LINC00312(LncRNA LINC00312)on the apoptosis of human pathological scar fibroblast and its possible mechanism.Methods Human pathological scar fibroblasts were isolated and cultured in vitro,and the cells were infected with LINC00312 shRNA lentivirus and negative control lentivirus.Then detection of the expression of LINC00312 in lentivirus-infected cells by qRT-PCR assay.According to the experimental requirements,the cells were divided into four groups:blank control group(Blank),negative control lentivirus group(NC-shRNA),LINC00312 shRNA lentivirus group(LINC00312-shRNA),and IRE1αspecific inhibitor group(LINC00312-shRNA+KIRA6).CCK-8 method was used to detect the cell proliferation of each group.Hoechst staining was used to observe the cell morphological changes of each group.The apoptosis of cells was measured by Annexin V-FITC/PI method in each group.The expression of apoptosis related proteins cleaved Caspase-3,Bax and Bcl-2,and endoplasmic reticulum stress related proteins GRP78,IRE1,p-IRE1,JNK and p-JNK proteins were detected by Western blot assay.Results The expression of LINC00312 decreased significantly in the cells infected with LINC00312 shRNA lentivirus(P<0.05).Compared with the blank group and the NC-shRNA group,the proliferation activity of cells was significantly reduced,the nucleus was densely stained with a large number of fragments,and the apoptosis rate was markedly increased,the expression of cleaved Caspsae-3,Bax,GRP78,p-IRE1 and p-JNK proteins were significantly increased,and the expression of Bcl-2 protein was decreased(P<0.05)in the LINC00312-shRNA group.Compared with the LINC00312-shRNA group,the proliferation activity of cells was significantly increased,the number of fragments of the nucleus was reduced,and the apoptosis rate was significantly suppressed,the expression of cleaved Caspase-3,Bax,GRP78,p-IRE1 and p-JNK proteins were significantly down-regulated,but the expression of Bcl-2 protein was markedly up-regulated(P<0.05)in

关 键 词：长链非编码RNA LINC00312 人病理性瘢痕成纤维细胞 内质网应激 细胞凋亡 

分 类 号：R622[医药卫生—整形外科]