左旋肉碱促进碱烧伤后角膜上皮修复的作用及机制  

Effect and mechanism of L-carnitine on corneal epithelial repair after alkali burn

作　　者：余冰洁 程钧 宋珊 杨玲玲 Bing-Jie Yu;Jun Cheng;Shan Song;Ling-Ling Yang(Qingdao University,Qingdao 266071,Shandong Province,China;State Key Laboratory Cultivation Base&Shandong Provincial Key Laboratory of Ophthalmology,Shandong Eye Institute,Shandong First Medical University,Shandong Academy of Medical Sciences,Qingdao 266071,Shandong Province,China;Eye Hospital of Shandong First Medical University,Qingdao 266071,Shandong Province,China;Department of Ophthalmology,Chengwu Hospital Affiliated to Shandong First Medical University,Heze 274200,Shandong Province,China)

机构地区：[1]青岛大学,中国山东省青岛市266071 [2]山东第一医科大学,山东省医学科学院,山东省眼科研究所,山东省眼科学重点实验室-省部共建国家重点实验室培育基地,中国山东省青岛市266071 [3]山东第一医科大学附属青岛眼科医院,中国山东省青岛市266071 [4]山东第一医科大学附属成武医院眼科,中国山东省菏泽市274200

出　　处：《国际眼科杂志》2021年第7期1150-1155,共6页International Eye Science

基　　金：山东省自然科学青年基金项目(No.ZR2017BH026)。

摘　　要：目的:探讨在角膜碱烧伤所致高渗和炎症环境中,左旋肉碱(LC)对角膜上皮修复的作用及其调控的分子机制。方法:将96只健康C57/6J小鼠随机分为空白对照组、磷酸缓冲盐溶液(PBS)组和LC组,空白对照组不做任何处理、LC组及PBS组制备急性碱烧伤模型。自模型制作前1d,LC组给予60mmol/L LC滴眼液,PBS组给予磷酸缓冲盐溶液连续每天点眼,每天6次。建模后0h,3、7d裂隙灯显微镜下荧光素钠染色观察角膜上皮修复情况。在第3d,取眼球进行冰冻切片免疫荧光检测Ki-67、IL-1β蛋白表达情况;提取角膜上皮全蛋白进行Western blot检测P63、NLRP3、Caspase-1的表达及STAT3的活化水平。结果:角膜荧光素钠染色结果显示在碱烧伤后3、7d时PBS组与LC组比,残留角膜上皮缺损面积占原始缺损面积的百分比分别为(29.38±6.83)%vs(17.78±4.11)%、(14.23±4.51)%vs(4.10±2.10)%(P<0.01)。LC组比PBS组角膜上皮修复快。造模后3d,与空白对照组比,碱烧伤处理的各组小鼠角膜上皮中焦亡相关蛋白NLRP3、Caspase-1表达上调,P63表达降低,p-STAT3/STAT3水平升高,除cleaved Caspase-1空白对照组vs LC组,差异均有意义。与PBS组比,LC组的NLRP3、pro Caspase-1、cleaved Caspase-1蛋白表达降低,P63表达量上调,p-STAT3/STAT3水平升高,差异均有意义。免疫荧光显示,碱烧伤后3d,PBS组和LC组与空白对照组比IL-1β、Ki-67表达上调。与PBS组比,LC组Ki-67蛋白表达上调,IL-1β表达降低。结论:LC可通过抑制细胞焦亡信号通路,促进STAT3信号通路的活化,促使小鼠角膜上皮干/祖细胞增生,进而促进碱烧伤后角膜上皮的修复。AIM:To investigate the effect of L-carnitine(LC)on corneal epithelial repair and its regulatory molecular mechanism in the hypertonic and inflammatory environment caused by alkali burn.METHODS:Ninety-six healthy C57/6J mice were randomly divided into blank control group,phosphate buffered solution(PBS)group and LC group.The blank control group did not receive any treatment,LC group and PBS group were prepared acute alkali burn models.LC group was given 60mmol/L LC eye drops,and PBS group was given PBS eye drops,6 times/d,for continuous days from one day before alkali burn.The repair of corneal epithelium was observed by fluorescein sodium staining under slit lamp microscope at 0h,3 and 7d.On the 3d,the expressions of Ki-67 and IL-1βproteins in cornea were detected by immunofluorescence,the total proteins of corneal epithelial were extracted for Western blot to detect the expression of P63,NLRP3,Caspase-1 and phosphorylation level of STAT3.RESULTS:The results of corneal fluorescein sodium staining showed that on the 3 and 7d after alkali burn,the percentage of residual corneal epithelial defect area in PBS group compared with LC group was(29.38±6.83)%vs(17.78±4.11)%and(14.23±4.51)%vs(4.10±2.10)%,respectively(P<0.01).The repair of corneal epithelium in LC group was faster than that in PBS group.On the 3d,compared with the blank control group,the expressions of pyroptosis related proteins NLRP3 and Caspase-1 in the corneal epithelium of the alkali burn treated mice were up-regulated,the expression of P63 was decreased,and the p-STAT3/STAT3 level was increased,all the differences were significant except cleaved Caspase-1 of blank control group vs LC group.Compared with PBS group,in LC group,the expression of NLRP3,pro Caspase-1 and cleaved Caspase-1 protein were decreased,P63 was up-regulated,and p-STAT3/STAT3 was increased,all the differences were significant.Immunofluorescence showed that compared with the blank control group,the expressions of IL-1βand Ki-67 were up-regulated in the alkali burned group.Compa

关 键 词：左旋肉碱 碱烧伤 焦亡 角膜缘干细胞 

分 类 号：R73[医药卫生—肿瘤]