中华传染病杂志
中華傳染病雜誌
중화전염병잡지
CHINESE JOURNAL OF INFECTIOUS DISEASES
2011年
7期
385-389
,共5页
余东山%安方梅%龚邦东%赵钢德%项晓刚%林兰意%俞红%王晖%谢青
餘東山%安方梅%龔邦東%趙鋼德%項曉剛%林蘭意%俞紅%王暉%謝青
여동산%안방매%공방동%조강덕%항효강%림란의%유홍%왕휘%사청
肝功能衰竭,急性%细胞凋亡%半胱天冬氨酸蛋白酶-8%微小RNA-1187
肝功能衰竭,急性%細胞凋亡%半胱天鼕氨痠蛋白酶-8%微小RNA-1187
간공능쇠갈,급성%세포조망%반광천동안산단백매-8%미소RNA-1187
Liver failure,acute%Apoptosis%Caspase-8%miR-1187
目的 通过微小RNA-1187(miR-1187)靶向调节半胱天冬氨酸蛋白酶-8(caspase-8)mRNA的表达,观察miR-1187对肝细胞凋亡的调控作用.方法 应用D-氨基半乳糖联合脂多糖构建BALB/c小鼠急性肝功能衰竭模型,取肝组织进行锁核酸(LNA)-miRNA微阵列分析和miRNA靶基因功能分析.体外培养胚胎小鼠肝细胞株2(BNL-CL2细胞),予以TNFα联合D-氨基半乳糖分别诱导转染miR-1187模拟物组及未转染组,以正常生长细胞为对照,应用实时(RT)-PCR、Western印迹等方法检测各组细胞miR-1187、caspase-8 mRNA及caspase-8蛋白的表达,并采用流式细胞技术测定细胞凋亡率.组间均数比较采用t检验.结果 miR-1187的信号值随急性肝功能衰竭进展明显下降.miR-1187与caspase-8 mRNA的3'非编码端(3'UTR)有直接的结合位点关系;miR-1187在未转染组表现为显著下调,在转染组下调减缓(t=6.371,P<0.01);caspase-8 mRNA 在未转染组表达显著增高.在转染组上升明显减少(t=4.539,P<0.01);转染组细胞凋亡率明显低于未转染组(t=3.365,P<0.05).结论 miR-1187为肝细胞凋亡的抑制因子之一;高表达miR-1187通过靶向调节caspase-8 mRNA的表达,阻抑肝细胞凋亡的发生.
目的 通過微小RNA-1187(miR-1187)靶嚮調節半胱天鼕氨痠蛋白酶-8(caspase-8)mRNA的錶達,觀察miR-1187對肝細胞凋亡的調控作用.方法 應用D-氨基半乳糖聯閤脂多糖構建BALB/c小鼠急性肝功能衰竭模型,取肝組織進行鎖覈痠(LNA)-miRNA微陣列分析和miRNA靶基因功能分析.體外培養胚胎小鼠肝細胞株2(BNL-CL2細胞),予以TNFα聯閤D-氨基半乳糖分彆誘導轉染miR-1187模擬物組及未轉染組,以正常生長細胞為對照,應用實時(RT)-PCR、Western印跡等方法檢測各組細胞miR-1187、caspase-8 mRNA及caspase-8蛋白的錶達,併採用流式細胞技術測定細胞凋亡率.組間均數比較採用t檢驗.結果 miR-1187的信號值隨急性肝功能衰竭進展明顯下降.miR-1187與caspase-8 mRNA的3'非編碼耑(3'UTR)有直接的結閤位點關繫;miR-1187在未轉染組錶現為顯著下調,在轉染組下調減緩(t=6.371,P<0.01);caspase-8 mRNA 在未轉染組錶達顯著增高.在轉染組上升明顯減少(t=4.539,P<0.01);轉染組細胞凋亡率明顯低于未轉染組(t=3.365,P<0.05).結論 miR-1187為肝細胞凋亡的抑製因子之一;高錶達miR-1187通過靶嚮調節caspase-8 mRNA的錶達,阻抑肝細胞凋亡的髮生.
목적 통과미소RNA-1187(miR-1187)파향조절반광천동안산단백매-8(caspase-8)mRNA적표체,관찰miR-1187대간세포조망적조공작용.방법 응용D-안기반유당연합지다당구건BALB/c소서급성간공능쇠갈모형,취간조직진행쇄핵산(LNA)-miRNA미진렬분석화miRNA파기인공능분석.체외배양배태소서간세포주2(BNL-CL2세포),여이TNFα연합D-안기반유당분별유도전염miR-1187모의물조급미전염조,이정상생장세포위대조,응용실시(RT)-PCR、Western인적등방법검측각조세포miR-1187、caspase-8 mRNA급caspase-8단백적표체,병채용류식세포기술측정세포조망솔.조간균수비교채용t검험.결과 miR-1187적신호치수급성간공능쇠갈진전명현하강.miR-1187여caspase-8 mRNA적3'비편마단(3'UTR)유직접적결합위점관계;miR-1187재미전염조표현위현저하조,재전염조하조감완(t=6.371,P<0.01);caspase-8 mRNA 재미전염조표체현저증고.재전염조상승명현감소(t=4.539,P<0.01);전염조세포조망솔명현저우미전염조(t=3.365,P<0.05).결론 miR-1187위간세포조망적억제인자지일;고표체miR-1187통과파향조절caspase-8 mRNA적표체,조억간세포조망적발생.
Objective To observe the regulatory role of microRNA-1187(miR-1187)in hepatocyte apoptosis through miR-1187 targeting regulation of caspase-8 mRNA expression.Methods The acute liver failure model was established by injection of D-galactosamine plus lipopolysaccharides(LPS)in BALB/c mice.The liver tissues were collected for LNA-miRNA array analysis and functional analysis of genes targeted by miRNA.Embryonic murine hepatocyte cell line 2(BNL-CL2)was cultivated in vitro and treated with tumor necrosis factor(TNF)-α and D-galactosamine to induce the transfection of miR-1187 in transfected group or untransfected group.The expressions of miR-1187 and caspase-8 mRNA were detected by real-time polymeramse chain reaction(PCR)and caspase-8 protein was determined by Western blot.The apoptosis rate was detected by flow cytometry.The comparison of means between groups was done by t test.Results The miR-1187 signal was deceased with the development of acute liver failure.The 3'UTR of caspase-8 mRNA had direct binding sites with miR1187.In BNL-CL2 cell experiments,miR-1187 was down-regulated in untransfected group and decreased more slowly in transfected group(t=6.371,P<0.01).The expression of caspase-8 mRNA was up-regulated in untransfected group and increased less in transfected group(t=4.539,P<0.01).The apoptosis rate in transfected group was significantly lower than untransfected group(t=3.365,P<0.05).Conclusios miR-1187 is one of inhibitors of hepatocyte apoptosis.High expression of miR-1187 could regulate the expression of caspase-8 mRNA,thus inhibit the apoptosis of hepatocytes.
