中华糖尿病杂志
中華糖尿病雜誌
중화당뇨병잡지
CHINES JOURNAL OF DLABETES MELLITUS
2011年
2期
138-141
,共4页
李树颖%于德民%小川涉%春日雅人
李樹穎%于德民%小川涉%春日雅人
리수영%우덕민%소천섭%춘일아인
核糖体蛋白质S6激酶类%胰岛素%糖原异生%基因沉默
覈糖體蛋白質S6激酶類%胰島素%糖原異生%基因沉默
핵당체단백질S6격매류%이도소%당원이생%기인침묵
Ribosomal protein S6 kinase%Insulin%Gluconeogenesis%Gene silencing
目的 探讨核糖体蛋白S6激酶1基因沉默对db/db小鼠肝脏胰岛素抵抗的影响及机制.方法 采用随机数字表法将24只9周龄健康雄性db/db小鼠(体质量44.5～48.2 g)随机分至对照组(n1=12)和实验组(n=12),实验组小鼠尾静脉注射核糖体蛋白S6激酶1短发夹RNA重组基因腺病毒,对照组尾静脉注射含U6启动子空载体腺病毒.注射病毒后第6天,对照组和实验组采用随机数字表进一步分为进食组(n=6)及空腹组(n=6).在进食和空腹状态处死小鼠,提取肝脏,应用Western blot法观测胰岛素受体底物-1和胰岛素受体底物-2蛋白表达水平.提取肝脏总RNA,应用实时荧光定量逆转录聚合酶链反应检测糖异生基因mRNA表达水平.经尾静脉取血,采用酶联免疫吸附法测定胰岛素水平,采用比色法检测游离脂肪酸、甘油三酯、总胆固醇水平.组间比较采用单因素方差分析.结果 与对照组相比,进食及空腹状态时,实验组胰岛素受体底物-1、胰岛素受体底物-2蛋白表达均增加.在空腹状态下,实验组与对照组相比,糖异生基因过氧化酶体增殖物受体共激活因子1α(分别为0.072±0.024和0.028±0.012)、磷酸烯醇式丙酮酸羧激酶(分别为23.8±4.0和12.3±2.4)、葡萄糖6磷酸酶(分别为3.0±0.8和1.5±0.4)mRNA表达均下降(F值分别为7.58、5.76、9.15,均P＜0.01).对照组和实验组胰岛素水平无显著差异(t=1.26,P＞0.05).空腹及进食状态下,实验组胆固醇水平低于对照组(F=15.48,P＜0.01).与空腹对照组相比,空腹实验组游离脂肪酸降低(t=2.56,P＜0.05).结论 db/db小鼠肝脏核糖体蛋白S6激酶1基因沉默后,空腹糖异生和血清游离脂肪酸水平下降,胰岛素受体底物-1和胰岛素受体底物-2蛋白表达增加,提示营养过剩条件下核糖体蛋白S6激酶1可能在肝脏胰岛素抵抗中发挥重要作用.
目的 探討覈糖體蛋白S6激酶1基因沉默對db/db小鼠肝髒胰島素牴抗的影響及機製.方法 採用隨機數字錶法將24隻9週齡健康雄性db/db小鼠(體質量44.5～48.2 g)隨機分至對照組(n1=12)和實驗組(n=12),實驗組小鼠尾靜脈註射覈糖體蛋白S6激酶1短髮夾RNA重組基因腺病毒,對照組尾靜脈註射含U6啟動子空載體腺病毒.註射病毒後第6天,對照組和實驗組採用隨機數字錶進一步分為進食組(n=6)及空腹組(n=6).在進食和空腹狀態處死小鼠,提取肝髒,應用Western blot法觀測胰島素受體底物-1和胰島素受體底物-2蛋白錶達水平.提取肝髒總RNA,應用實時熒光定量逆轉錄聚閤酶鏈反應檢測糖異生基因mRNA錶達水平.經尾靜脈取血,採用酶聯免疫吸附法測定胰島素水平,採用比色法檢測遊離脂肪痠、甘油三酯、總膽固醇水平.組間比較採用單因素方差分析.結果 與對照組相比,進食及空腹狀態時,實驗組胰島素受體底物-1、胰島素受體底物-2蛋白錶達均增加.在空腹狀態下,實驗組與對照組相比,糖異生基因過氧化酶體增殖物受體共激活因子1α(分彆為0.072±0.024和0.028±0.012)、燐痠烯醇式丙酮痠羧激酶(分彆為23.8±4.0和12.3±2.4)、葡萄糖6燐痠酶(分彆為3.0±0.8和1.5±0.4)mRNA錶達均下降(F值分彆為7.58、5.76、9.15,均P＜0.01).對照組和實驗組胰島素水平無顯著差異(t=1.26,P＞0.05).空腹及進食狀態下,實驗組膽固醇水平低于對照組(F=15.48,P＜0.01).與空腹對照組相比,空腹實驗組遊離脂肪痠降低(t=2.56,P＜0.05).結論 db/db小鼠肝髒覈糖體蛋白S6激酶1基因沉默後,空腹糖異生和血清遊離脂肪痠水平下降,胰島素受體底物-1和胰島素受體底物-2蛋白錶達增加,提示營養過剩條件下覈糖體蛋白S6激酶1可能在肝髒胰島素牴抗中髮揮重要作用.
목적 탐토핵당체단백S6격매1기인침묵대db/db소서간장이도소저항적영향급궤제.방법 채용수궤수자표법장24지9주령건강웅성db/db소서(체질량44.5～48.2 g)수궤분지대조조(n1=12)화실험조(n=12),실험조소서미정맥주사핵당체단백S6격매1단발협RNA중조기인선병독,대조조미정맥주사함U6계동자공재체선병독.주사병독후제6천,대조조화실험조채용수궤수자표진일보분위진식조(n=6)급공복조(n=6).재진식화공복상태처사소서,제취간장,응용Western blot법관측이도소수체저물-1화이도소수체저물-2단백표체수평.제취간장총RNA,응용실시형광정량역전록취합매련반응검측당이생기인mRNA표체수평.경미정맥취혈,채용매련면역흡부법측정이도소수평,채용비색법검측유리지방산、감유삼지、총담고순수평.조간비교채용단인소방차분석.결과 여대조조상비,진식급공복상태시,실험조이도소수체저물-1、이도소수체저물-2단백표체균증가.재공복상태하,실험조여대조조상비,당이생기인과양화매체증식물수체공격활인자1α(분별위0.072±0.024화0.028±0.012)、린산희순식병동산최격매(분별위23.8±4.0화12.3±2.4)、포도당6린산매(분별위3.0±0.8화1.5±0.4)mRNA표체균하강(F치분별위7.58、5.76、9.15,균P＜0.01).대조조화실험조이도소수평무현저차이(t=1.26,P＞0.05).공복급진식상태하,실험조담고순수평저우대조조(F=15.48,P＜0.01).여공복대조조상비,공복실험조유리지방산강저(t=2.56,P＜0.05).결론 db/db소서간장핵당체단백S6격매1기인침묵후,공복당이생화혈청유리지방산수평하강,이도소수체저물-1화이도소수체저물-2단백표체증가,제시영양과잉조건하핵당체단백S6격매1가능재간장이도소저항중발휘중요작용.
Objective To study the effects of ribosomal protein S6 kinase 1 ( S6K1 ) gene silencing in pathogenesis of hepatic insulin resistance. Methods Twenty-four healthy male db/db mice aged 9 weeks (body weight 44. 5 to 48. 2 g) were randomly assigned to the control group (n = 12) and the study group (n = 12). S6K1 short hairpin RNA recombinant adenovirus (S6K1Ax) was injected into the tail vein of db/db mice of the study group and U6 promoter recombinant adenovirus (pU6Ax) was injected into the control mice. At 6 days,those mice were randomly assigned to the fasting group ( n = 6) and the food-intake group ( n = 6 ). Insulin receptor substrate-1 ( IRS-1 ) and insulin receptor substrate-2 ( IRS-2 ) were determined by Western blot. Total RNAs were extracted to analyze gluconeogenic genes by using real-time quantitative revere transcription polymerase chain reaction. Insulin was determined by enzyme-linked immunosorbent assay. Free fatty acids (FFA),triglyceride (TG),cholesterol and alanine aminotransferase were checked by colorimetry. All data were analyzed by One-way analysis of variance. Results Compared with the control group,the protein expression of IRS-1 and IRS-2 of the S6K1Ax group was increased.Peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α),phosphoenolpyruvate carboxykinase (PEPCK),and glucose-6-phosphatase (G6Pase) mRNA expression was descended (PGC1-α: 0. 072 ±0. 024 vs 0. 028 ±0. 012,F =7. 58; PEPCK: 23.8 ±4. 0 vs 12. 3 ±2.4,F =5. 76; G6Pase: 3.0 ±0. 8 vs 1.5 ±0. 4,F = 9. 15; all P ＜ 0. 01 ). There was no difference in serum insulin between the control and the study group ( t = 1.26,P ＞ 0. 05 ). Serum cholesterol of the study group was reduced in both fasting and food-intake state ( F = 15.48,P ＜0. 01 ). Compared with the fasting control group,fasting blood FFA was decreased in fasting state of the study group ( t = 2. 56,P ＜ 0. 05 ). Conclusions Hepatic gluconeogenesis genes and serum FFA may reduce while IRS-1 and IRS-2 protein may increase in hepatic S6K1 gene silencing mice. These findings suggest that S6K1 may play an important role in hepatic insulin resistance.
