中国循证儿科杂志
中國循證兒科雜誌
중국순증인과잡지
CHINESE JOURNAL OF EVIDENCE-BASED PEDIATRICS
2014年
5期
377-380
,共4页
腺病毒%IL-24%系膜细胞%脂多糖%凋亡%细胞周期调节蛋白
腺病毒%IL-24%繫膜細胞%脂多糖%凋亡%細胞週期調節蛋白
선병독%IL-24%계막세포%지다당%조망%세포주기조절단백
Adenovirus%Interleukin-24%Lipopolysaccharide%Glomerular mesangial cells%Apoptosis%Cell cycle regulatory protein
目的：探讨腺病毒介导的IL-24转移对脂多糖（ LpS）诱导的大鼠肾小球系膜细胞（ GMCs）和细胞周期调节蛋白p21、p27及CyclinE的影响。方法采用10%FBS/DMEM培养293细胞，扩增腺病毒载体，GMCs培养4~6代细胞后用于分析。①观察IL-24对LpS诱导的GMCs凋亡影响：分为对照组、Ad. IL-24组、LpS组和LpS+ Ad. IL-24组，其中对照组和LpS组GMCs不转染携带IL-24腺病毒（ Ad. IL-24），Ad. IL-24组和LpS+Ad. IL-24组以Ad. IL-24转染 GMCs，LpS组和LpS+Ad. IL-24组加LpS培养，以流式细胞术检测培养后24和48 h的GMCs凋亡率；②考察IL-24对GMCs周期蛋白影响：对照组为5%FBS/DMEM培养GMCs，Ad-GFp组采用携带绿色荧光蛋白的对照载体（ Ad. GFp）转染GMCs，Ad. IL-24+LpS组以Ad. IL-24转染GMCs，采用Western-blot检测p21、p27及CyclinE表达。结果①GMCs在培养后24和48 h细胞凋亡率：对照组分别为（0.86±0.15）%和（0.98±0.4）%；IL-24组分别为（1.02±0.22）%和（1.43±0.31）%；LpS组分别为（2.19±0.81）%和（2.49±0.12）%，LpS+Ad. IL-24组分别为（18.01±1.17）%、（26.82±5.01）%；2个观察时间点细胞凋亡率对照组与IL-24组差异无统计学意义，LpS组显著高于对照组（ P<0.05），LpS+ Ad. IL-24组细胞凋亡率最高（P<0.05）。②Ad-GFp组p21、p27表达显著低于对照组（P<0.05），CyclinE表达显著高于对照组（P<0.05）；Ad. IL-24+LpS组p21、p27表达较Ad-GFp组显著上调（P<0.05），CyclinE表达较Ad-GFp组下调。结论 IL-24腺病毒载体对正常GMCs无影响，但可使LpS诱导增生的 GMCs凋亡增加，其可能机制是上调关键细胞周期负调控蛋白 p21、p27及下调CyclinE。
目的：探討腺病毒介導的IL-24轉移對脂多糖（ LpS）誘導的大鼠腎小毬繫膜細胞（ GMCs）和細胞週期調節蛋白p21、p27及CyclinE的影響。方法採用10%FBS/DMEM培養293細胞，擴增腺病毒載體，GMCs培養4~6代細胞後用于分析。①觀察IL-24對LpS誘導的GMCs凋亡影響：分為對照組、Ad. IL-24組、LpS組和LpS+ Ad. IL-24組，其中對照組和LpS組GMCs不轉染攜帶IL-24腺病毒（ Ad. IL-24），Ad. IL-24組和LpS+Ad. IL-24組以Ad. IL-24轉染 GMCs，LpS組和LpS+Ad. IL-24組加LpS培養，以流式細胞術檢測培養後24和48 h的GMCs凋亡率；②攷察IL-24對GMCs週期蛋白影響：對照組為5%FBS/DMEM培養GMCs，Ad-GFp組採用攜帶綠色熒光蛋白的對照載體（ Ad. GFp）轉染GMCs，Ad. IL-24+LpS組以Ad. IL-24轉染GMCs，採用Western-blot檢測p21、p27及CyclinE錶達。結果①GMCs在培養後24和48 h細胞凋亡率：對照組分彆為（0.86±0.15）%和（0.98±0.4）%；IL-24組分彆為（1.02±0.22）%和（1.43±0.31）%；LpS組分彆為（2.19±0.81）%和（2.49±0.12）%，LpS+Ad. IL-24組分彆為（18.01±1.17）%、（26.82±5.01）%；2箇觀察時間點細胞凋亡率對照組與IL-24組差異無統計學意義，LpS組顯著高于對照組（ P<0.05），LpS+ Ad. IL-24組細胞凋亡率最高（P<0.05）。②Ad-GFp組p21、p27錶達顯著低于對照組（P<0.05），CyclinE錶達顯著高于對照組（P<0.05）；Ad. IL-24+LpS組p21、p27錶達較Ad-GFp組顯著上調（P<0.05），CyclinE錶達較Ad-GFp組下調。結論 IL-24腺病毒載體對正常GMCs無影響，但可使LpS誘導增生的 GMCs凋亡增加，其可能機製是上調關鍵細胞週期負調控蛋白 p21、p27及下調CyclinE。
목적：탐토선병독개도적IL-24전이대지다당（ LpS）유도적대서신소구계막세포（ GMCs）화세포주기조절단백p21、p27급CyclinE적영향。방법채용10%FBS/DMEM배양293세포，확증선병독재체，GMCs배양4~6대세포후용우분석。①관찰IL-24대LpS유도적GMCs조망영향：분위대조조、Ad. IL-24조、LpS조화LpS+ Ad. IL-24조，기중대조조화LpS조GMCs불전염휴대IL-24선병독（ Ad. IL-24），Ad. IL-24조화LpS+Ad. IL-24조이Ad. IL-24전염 GMCs，LpS조화LpS+Ad. IL-24조가LpS배양，이류식세포술검측배양후24화48 h적GMCs조망솔；②고찰IL-24대GMCs주기단백영향：대조조위5%FBS/DMEM배양GMCs，Ad-GFp조채용휴대록색형광단백적대조재체（ Ad. GFp）전염GMCs，Ad. IL-24+LpS조이Ad. IL-24전염GMCs，채용Western-blot검측p21、p27급CyclinE표체。결과①GMCs재배양후24화48 h세포조망솔：대조조분별위（0.86±0.15）%화（0.98±0.4）%；IL-24조분별위（1.02±0.22）%화（1.43±0.31）%；LpS조분별위（2.19±0.81）%화（2.49±0.12）%，LpS+Ad. IL-24조분별위（18.01±1.17）%、（26.82±5.01）%；2개관찰시간점세포조망솔대조조여IL-24조차이무통계학의의，LpS조현저고우대조조（ P<0.05），LpS+ Ad. IL-24조세포조망솔최고（P<0.05）。②Ad-GFp조p21、p27표체현저저우대조조（P<0.05），CyclinE표체현저고우대조조（P<0.05）；Ad. IL-24+LpS조p21、p27표체교Ad-GFp조현저상조（P<0.05），CyclinE표체교Ad-GFp조하조。결론 IL-24선병독재체대정상GMCs무영향，단가사LpS유도증생적 GMCs조망증가，기가능궤제시상조관건세포주기부조공단백 p21、p27급하조CyclinE。
Objective To explore the effect of interleukin-24(IL-24)gene transfer on glomerular mesangial cells(GMCs) apoptosis and to find out the effect of IL-24 on cell cycle regulatory protein p21,p27 and CyclinE of GMCs induced by LpS. Methods 293 cells were cultured in 10%FBS/DMEM and Ad. IL-24 and Ad. GFp were amplifycated in 293 cells. GMCs were analysed after 4 to 6 generations. ①They were divided into four groups:control group,Ad. IL-24 group,LpS group and LpS+Ad. IL-24 group. And control group and LpS group werenˊt infected with Ad. IL-24,Ad. IL-24 group and LpS+Ad. IL-24 group GMCs were infected with Ad. IL-24,then LpS+Ad. IL-24 group GMCs were cultured in 5%FBS/DMEM with LpS(10 mg·L-1 ). The apoptosis of the GMCs was examined by AnnexinV/FITC flow cytometry;②The effect of IL-24 on cell cycle regulatory protein p21, p27 and CyclinE of GMCs induced by LpS were determined. They were divided into three groups:control group,Ad-GFp group and IL-24 group. Control group GMCs were cultured in 5%FBS/DMEM. Ad-GFp group GMCs were infected with Ad. GFp and then cultured in 5%FBS/DMEM with LpS(10 mg·L-1 ). GMCs were infected with Ad. IL-24. The expressions of cell cycle regulatory protein p21,p27 and cyclinE were examined by Western-blotting. Results The GMCs were cultured for 24 hours and 48 hours. The apoptosis rate was(0. 86 ± 0. 15)% and(0. 98 ± 0. 4)% in the control group,(1. 02 ± 0. 22)% and(1. 43 ± 0. 31)% in the Ad. IL-24 group,(2. 19 ± 0. 81)% and(2. 49 ± 0. 12)% in the LpS group,(18. 01 ± 1. 17)% and(26. 82 ± 5. 01)% in LpS + Ad. IL-24 group. There was no difference between control group and Ad. IL-24 group,and the apoptosis rate of LpS group was higher than control group(P<0. 05). The apoptosis rate of LpS+Ad. IL-24 group was the highest while there was no change in Ad. IL-24 group(P<0. 05). ②The expressions of p21 and p27 were down-regulated while CyclinE expression was up-regulated in GMC by LpS(P<0. 05). Adenovirus mediated IL-24 gene transfer could abolish the role of LpS on regulation of p21, p27 and CyclinE expression(P<0. 05). Conclusion Adenovirus mediated exogenous IL-24 gene transfer could effectively inhibit LpS-induced proliferation of glomerular mesangial cells but had no effect on normal GMCs,and it could abolish the role of LpS on regulation of p21,p27 and CyclinE expression then increased the apoptosis of LpS-induced proliferation of GMCs.
