长链非编码RNA-TUG1调控小梁网细胞内质网应激和炎症的小鼠实验研究  

Regulation of endoplasmic reticulum stress and inflammation in trabecular reticulum cells by long non-coding RNA-TUG1

作　　者：陈雪红 吴子东 庄海容 陈圣文[1] CHEN Xue-hong;WU Zi-dong;ZHUANG Hai-rong(Department of Ophthalmology,Second Affiliated Hospital of Hainan Medical College,Haikou Hainan 570100,China)

机构地区：[1]海南医学院第二附属医院眼科,海南海口570100

出　　处：《临床和实验医学杂志》2023年第9期897-902,共6页Journal of Clinical and Experimental Medicine

基　　金：海南省卫生健康行业科研项目(编号:18A200169)。

摘　　要：目的观察长链非编码RNA牛磺酸上调基因1(Lnc-TUG1)对小梁网细胞内质网应激及其主要标志物的影响,并探讨Lnc-TUG1在小梁网功能障碍、细胞凋亡中所起的作用。方法选取C57BL/6J小鼠48只,随机数字表法分为两组,AAV2-Lnc-TUG1组(24只,24眼)单眼前房注射Lnc-TUG1过表达腺病毒,AAV2-GFP组(24只,24眼)单眼前房注射GFP腺病毒。在注射后的不同时间点,分别取AAV2-GFP组和AAV2-Lnc-TUG1组小鼠的前房角组织,用冰冻切片和免疫荧光染色法观察病毒荧光及内质网应激相关蛋白C/EBP同源蛋白(CHOP)的表达;荧光原位杂交法观察Lnc-TUG1的表达和定位。用实时荧光定量PCR法检测炎症因子白细胞介素(IL)-1α、IL-1β、IL-6以及内皮细胞白细胞黏附分子1(ELAM-1)在mRNA水平的表达;用TUNEL染色法观察小梁网细胞的凋亡情况;用超薄切片和透射电镜观察小梁网细胞的超微结构。在注射前及注射后3、7、10、13 d分别用回弹式眼压计测量小鼠的日间及夜间眼压。结果Lnc-TUG1过表达腺病毒在注射后24 h即可观察到注射眼小梁网内的病毒荧光,且AAV2-Lnc-TUG1组的小梁网Lnc-TUG1的表达明显高于AAV2-GFP组,差异有统计学意义(P<0.05)。在注射后3 d,AAV2-Lnc-TUG1组小梁网组织的IL-1α、IL-1β、IL-6、ELAM-1在mRNA水平的表达明显高于AAV2-GFP组,差异有统计学意义(P<0.05)。注射后7 d,与AAV2-GFP组比,AAV2-GFP组和AAV2-Lnc-TUG1组小梁网组织CHOP的表达上调,差异有统计学意义(P<0.05),且小梁网TUNEL阳性细胞数显著增加,小梁网细胞内可见明显扩张的粗面内质网。在注射后7、10、13 d,AAV2-Lnc-TUG1组日间眼压及夜间眼压均高于AAV2-GFP组,但仅术后7 d的差异有统计学意义(P<0.05)。结论单眼前房注射Lnc-TUG1过表达腺病毒可通过上调Lnc-TUG1的表达增加小鼠眼压,并促进小梁网细胞的内质网应激、炎症和细胞凋亡。Objective To investigate the effect of long non-coding RNA taurine up-regulated gene 1(Lnc-TUG1)on endoplasmic reticulum stress and its main markers in trabecular meshwork cells,and to explore the role of Lnc-TUG1 in trabecular meshwork dysfunction and apoptosis.Methods A total of 48 C57BL/6 mice were selected.The experimental group(24,24 eyes)was injected with Lnc-TUG1 overexpressed adenovirus in single eye and the control group(24,24 eyes)was injected with GFP adenovirus in single eye.The expression of viral fluorescence and er stress-related protein C/EBP homologous protein(CHOP)was observed by freezing section and immunofluorescence staining at different time points after injection.The expression and localization of Lnc-TUG1 were observed by fluorescence in situ hybridization.The mRNA levels of inflammatory cytokines interleukin(IL)-1α,IL-1β,IL-6 and endothelial leukocyte adhesion molecule-1(ELAM-1)were detected by real-time fluorescence quantitative PCR.The apoptosis of trabecular meshwork cells was observed by TUNEL staining.Ultrastructure of trabecular meshwork cells was observed by ultrathin section and transmission electron microscope.The intraocular and nocturnal intraocular pressure of mice were measured by rebound tonometer before and at 3,7,10 and 13 days after injection.Results The fluorescence of Lnc-TUG1 overexpressed adenovirus in trabecular meshwork could be observed 24 h after injection,and the expression of Lnc-TUG1 in trabecular meshwork in the experimental group was significantly up-regulated compared with that in the control group,the difference was statistically significant(P<0.05).On day 3 after injection,the mRNA levels of IL-1α,IL-1β,IL-6 and ELAM-1 in trabecular meshwork in the experimental group were significantly higher than those in the control group,the differences were statistically significant(P<0.05).On day 7 after injection,compared with the control group,the expression of CHOP in trabecular meshwork was up-regulated in the experimental group,the difference was statisticall

关 键 词：小鼠 长链非编码RNA-TUG1 小梁网细胞 内质网应激 炎症 细胞凋亡 

分 类 号：R775[医药卫生—眼科]