中华眼底病杂志
中華眼底病雜誌
중화안저병잡지
CHINESE JOURNAL OF OCULAR FUNDUS DISEASES
2014年
2期
161-165
,共5页
视网膜色素上皮/损伤%瘦素/中毒%活性氧/中毒
視網膜色素上皮/損傷%瘦素/中毒%活性氧/中毒
시망막색소상피/손상%수소/중독%활성양/중독
Retinal pigment epithelium/injuries%Leptin/poisoning%Reactive oxygen species/poisoning
目的 观察瘦素对胰岛素抵抗状态下人视网膜色素上皮(RPE)细胞氧化损伤的影响.方法 体外培养的人RPE细胞随机分为正常对照组和胰岛素抵抗组.分别加入0、10、100 ng/ml瘦素干预24、48、72 h.2 ',7'-二氯荧光素二乙酸酯(DCFH-DA)法检测细胞内活性氧(ROS)的产生情况;免疫细胞化学法检测细胞内8-羟基-2'-脱氧鸟嘌呤(8-OHdG)的表达量;蛋白质免疫印迹法检测细胞浆中8-羟基鸟嘌呤DNA糖苷酶1(hOGGl)的表达量.结果 干预后24、48、72 h,正常对照组、胰岛素抵抗组RPE细胞ROS(正常对照组:F=37.136、37.178、49.634;胰岛素抵抗组:F=9.822、28.881、71.150)、8-OHdG表达量(正常对照组:F=88.643、390.920、1039.276;胰岛素抵抗组:F=273.311、299.155、82.237)均随瘦素浓度增加而增加,hOGG1的表达量(正常对照组:F=470.062、1073.113、295.456;胰岛素抵抗组:F=240.032、592.389、527.760)随瘦素浓度的增加而递增,差异均有统计学意义(P＜0.05).正常对照组、胰岛素抵抗组在相同浓度瘦素干预下,随干预时间延长,胰岛素抵抗组RPE细胞8-OHdG表达量呈现高于正常对照组的趋势.干预后24 h,胰岛素抵抗组RPE细胞hOGG1表达量与正常对照组比较,差异无统计学意义(F=23.392,P＞0.05);干预后72 h,胰岛素抵抗组PRE细胞hOGG1表达量低于正常对照组,差异有统计学意义(F=129.394,P＜0.05).在相同浓度瘦素干预下,随干预时间延长,RPE细胞hOGG1表达量呈现先升高后降低的趋势.干预后48 h,正常对照组(F=83.673、268.000、373.492)、胰岛素抵抗组(F=49.021、304.293、294.293)RPE细胞hOGG1表达量均高于干预后24、72 h RPE细胞hOGG1表达量,差异有统计学意义(P＜0.05).结论 正常状态及胰岛素抵抗状态下瘦素能引起人RPE细胞氧化损伤,随着瘦素浓度的增加、作用时间的延长氧化损伤的程度呈加重趋势;氧化损伤的修复随着时间的延长呈先强后弱的趋势,随着瘦素浓度的增加而呈增强的趋势.
目的 觀察瘦素對胰島素牴抗狀態下人視網膜色素上皮(RPE)細胞氧化損傷的影響.方法 體外培養的人RPE細胞隨機分為正常對照組和胰島素牴抗組.分彆加入0、10、100 ng/ml瘦素榦預24、48、72 h.2 ',7'-二氯熒光素二乙痠酯(DCFH-DA)法檢測細胞內活性氧(ROS)的產生情況;免疫細胞化學法檢測細胞內8-羥基-2'-脫氧鳥嘌呤(8-OHdG)的錶達量;蛋白質免疫印跡法檢測細胞漿中8-羥基鳥嘌呤DNA糖苷酶1(hOGGl)的錶達量.結果 榦預後24、48、72 h,正常對照組、胰島素牴抗組RPE細胞ROS(正常對照組:F=37.136、37.178、49.634;胰島素牴抗組:F=9.822、28.881、71.150)、8-OHdG錶達量(正常對照組:F=88.643、390.920、1039.276;胰島素牴抗組:F=273.311、299.155、82.237)均隨瘦素濃度增加而增加,hOGG1的錶達量(正常對照組:F=470.062、1073.113、295.456;胰島素牴抗組:F=240.032、592.389、527.760)隨瘦素濃度的增加而遞增,差異均有統計學意義(P＜0.05).正常對照組、胰島素牴抗組在相同濃度瘦素榦預下,隨榦預時間延長,胰島素牴抗組RPE細胞8-OHdG錶達量呈現高于正常對照組的趨勢.榦預後24 h,胰島素牴抗組RPE細胞hOGG1錶達量與正常對照組比較,差異無統計學意義(F=23.392,P＞0.05);榦預後72 h,胰島素牴抗組PRE細胞hOGG1錶達量低于正常對照組,差異有統計學意義(F=129.394,P＜0.05).在相同濃度瘦素榦預下,隨榦預時間延長,RPE細胞hOGG1錶達量呈現先升高後降低的趨勢.榦預後48 h,正常對照組(F=83.673、268.000、373.492)、胰島素牴抗組(F=49.021、304.293、294.293)RPE細胞hOGG1錶達量均高于榦預後24、72 h RPE細胞hOGG1錶達量,差異有統計學意義(P＜0.05).結論 正常狀態及胰島素牴抗狀態下瘦素能引起人RPE細胞氧化損傷,隨著瘦素濃度的增加、作用時間的延長氧化損傷的程度呈加重趨勢;氧化損傷的脩複隨著時間的延長呈先彊後弱的趨勢,隨著瘦素濃度的增加而呈增彊的趨勢.
목적 관찰수소대이도소저항상태하인시망막색소상피(RPE)세포양화손상적영향.방법 체외배양적인RPE세포수궤분위정상대조조화이도소저항조.분별가입0、10、100 ng/ml수소간예24、48、72 h.2 ',7'-이록형광소이을산지(DCFH-DA)법검측세포내활성양(ROS)적산생정황;면역세포화학법검측세포내8-간기-2'-탈양조표령(8-OHdG)적표체량;단백질면역인적법검측세포장중8-간기조표령DNA당감매1(hOGGl)적표체량.결과 간예후24、48、72 h,정상대조조、이도소저항조RPE세포ROS(정상대조조:F=37.136、37.178、49.634;이도소저항조:F=9.822、28.881、71.150)、8-OHdG표체량(정상대조조:F=88.643、390.920、1039.276;이도소저항조:F=273.311、299.155、82.237)균수수소농도증가이증가,hOGG1적표체량(정상대조조:F=470.062、1073.113、295.456;이도소저항조:F=240.032、592.389、527.760)수수소농도적증가이체증,차이균유통계학의의(P＜0.05).정상대조조、이도소저항조재상동농도수소간예하,수간예시간연장,이도소저항조RPE세포8-OHdG표체량정현고우정상대조조적추세.간예후24 h,이도소저항조RPE세포hOGG1표체량여정상대조조비교,차이무통계학의의(F=23.392,P＞0.05);간예후72 h,이도소저항조PRE세포hOGG1표체량저우정상대조조,차이유통계학의의(F=129.394,P＜0.05).재상동농도수소간예하,수간예시간연장,RPE세포hOGG1표체량정현선승고후강저적추세.간예후48 h,정상대조조(F=83.673、268.000、373.492)、이도소저항조(F=49.021、304.293、294.293)RPE세포hOGG1표체량균고우간예후24、72 h RPE세포hOGG1표체량,차이유통계학의의(P＜0.05).결론 정상상태급이도소저항상태하수소능인기인RPE세포양화손상,수착수소농도적증가、작용시간적연장양화손상적정도정가중추세;양화손상적수복수착시간적연장정선강후약적추세,수착수소농도적증가이정증강적추세.
Objective To investigate the effects of leptin on the oxidative damage in human retinal pigment epithelial (RPE) cells.Methods Human RPE cells (ARPE-19) were cultured in vitro,and randomly divided into control group and insulin resistance group.RPE cells were treated with 0,10,100 ng/mL leptin for 24,48,72 hours respectively.Then the levels of reactive oxygen species (ROS) expression in RPE cells were detected by 2',7'-dichlorofluorescin-diacetate (DCFH-DA),and the levels of 8-hydroxy-2′-deoxyguanosine (8-OHdG) expression in RPE cells were observed by immunocytochemistry (ICC),and the levels of human 8-oxoguanine DNA glycosylase 1 (hOGG1) expression in lysate were measured by Western blot.Results After 24,48,72 hours,the level of ROS (Control group:F=37.136,37.178,49.634; P＜0.05.Insulin resistance group:F=9.822,28.881,71.150;P＜0.05),8-OHdG (Control group:F =88.643,390.920,1039.276; P ＜ 0.05.Insulin resistance group:F =273.311,299.155,82.237;P＜0.05) and hOGGl (Control group:F=470.062,1073.113,295.456;P＜0.05.Insulin resistance group:F =240.032,592.389,527.760 ; P＜0.05) expression increased significantly with the increase of leptin concentration in control group and insulin resistance group.Under the same leptin concentration,the level of 8-OHdG has a trend that it was higher in the insulin resistance group than the control group.After 24 hours,the difference of hOGGl expression between control group and insulin resistance group was not significant (F=23.392,P＞0.05).After 72 hours,the level of hOGGl expression was significantly higher in the insulin resistance group than the control group (F=129.394,P＜0.05).The level of hOGGl expression was significantly higher at 48 hours than that at 24 hours and 72 hours (P＜ 0.05).Conclusion Leptin could induce the oxidative damage of RPE cells in normal and insulin resistance status.With the increase of leptin concentration and time extended,the degree of oxidative damage and its repair were both increased.The degree of oxidative repair increased with the increase of leptin concentration,but decreased with time extended.
