CAJ | 학술논문

目的：探讨胰岛素及格列齐特治疗2型糖尿病对大鼠肝脏脂质沉积的影响及机制。方法：制备高脂及链脲佐菌素诱导的2型糖尿病大鼠模型，随机分为糖尿病组、胰岛素组和格列齐特组，并设正常对照组。通过肝脏油红O染色观察其肝细胞脂质沉积情况；ELISA检测血清脂联素水平；实时荧光定量PCR检测肝脏脂联素受体1（AdipoR1）mRNA表达；Western blotting检测肝脏腺苷酸活化蛋白激酶（AMPK）、磷酸化的腺苷酸活化蛋白激酶（Thr172p-AMPK）、固醇调节因子结合蛋白1c（SREBP-1c）、磷酸化的固醇调节因子结合蛋白1c（Ser372p-SREBP1-1c）、乙酰辅酶A羧化酶（ACC）、磷酸化的乙酰辅酶A羧化酶（Ser79p-ACC）和免疫球蛋白结合蛋白（BiP）的表达。结果：糖尿病组肝细胞脂质沉积较正常对照组明显增多，胰岛素和格列齐特治疗后肝细胞脂质沉积明显改善。胰岛素治疗后，血清脂联素的水平及肝脏AdipoR1 mRNA水平较糖尿病组和正常对照组显著升高（P＜0．01），而格列齐特治疗后两者水平恢复至正常对照组水平。 Western blotting 结果显示，糖尿病大鼠与正常对照组比较，肝脏Thr172p-AMPK／AMPK和Ser372p-SREBP-1c／SREBP-1c和Ser79p-ACC／ACC表达明显降低（P＜0．01），BiP表达明显升高（P＜0．01）。胰岛素治疗后，Thr172p-AMPK／AMPK和Ser372p-SREBP-1c／SREBP-1c显著升高（P＜0．01）， Ser79p-ACC／ACC和BiP蛋白表达恢复至正常对照组水平。而格列齐特治疗后Thr172p-AMPK／AMPK和Ser372p-SREBP-1c／SREBP-1c恢复至正常对照组水平，BiP蛋白表达显著下降（P＜0．01），Ser79p-ACC／ACC与糖尿病组比较无明显改善。结论：胰岛素和格列齐特治疗均能通过激活脂联素-AMPK减轻2型糖尿病大鼠肝脏的脂质沉积。但两者作用的分子机制有所不同。胰岛素激活 AMPK，通过抑制SREBP-1c表达、直接磷酸化 SREBP-1c 抑制SREBP-1c入核等短期和长期的作用以及抑制内质网应激影响SREBP-1c，减少脂质合成；而格列齐特仅通过磷酸化的短期作用和抑制内质网应激对SREBP-1c产生影响，且对脂肪酸氧化无作用。目的：探討胰島素及格列齊特治療2型糖尿病對大鼠肝髒脂質沉積的影響及機製。方法：製備高脂及鏈脲佐菌素誘導的2型糖尿病大鼠模型，隨機分為糖尿病組、胰島素組和格列齊特組，併設正常對照組。通過肝髒油紅O染色觀察其肝細胞脂質沉積情況；ELISA檢測血清脂聯素水平；實時熒光定量PCR檢測肝髒脂聯素受體1（AdipoR1）mRNA錶達；Western blotting檢測肝髒腺苷痠活化蛋白激酶（AMPK）、燐痠化的腺苷痠活化蛋白激酶（Thr172p-AMPK）、固醇調節因子結閤蛋白1c（SREBP-1c）、燐痠化的固醇調節因子結閤蛋白1c（Ser372p-SREBP1-1c）、乙酰輔酶A羧化酶（ACC）、燐痠化的乙酰輔酶A羧化酶（Ser79p-ACC）和免疫毬蛋白結閤蛋白（BiP）的錶達。結果：糖尿病組肝細胞脂質沉積較正常對照組明顯增多，胰島素和格列齊特治療後肝細胞脂質沉積明顯改善。胰島素治療後，血清脂聯素的水平及肝髒AdipoR1 mRNA水平較糖尿病組和正常對照組顯著升高（P＜0．01），而格列齊特治療後兩者水平恢複至正常對照組水平。 Western blotting 結果顯示，糖尿病大鼠與正常對照組比較，肝髒Thr172p-AMPK／AMPK和Ser372p-SREBP-1c／SREBP-1c和Ser79p-ACC／ACC錶達明顯降低（P＜0．01），BiP錶達明顯升高（P＜0．01）。胰島素治療後，Thr172p-AMPK／AMPK和Ser372p-SREBP-1c／SREBP-1c顯著升高（P＜0．01）， Ser79p-ACC／ACC和BiP蛋白錶達恢複至正常對照組水平。而格列齊特治療後Thr172p-AMPK／AMPK和Ser372p-SREBP-1c／SREBP-1c恢複至正常對照組水平，BiP蛋白錶達顯著下降（P＜0．01），Ser79p-ACC／ACC與糖尿病組比較無明顯改善。結論：胰島素和格列齊特治療均能通過激活脂聯素-AMPK減輕2型糖尿病大鼠肝髒的脂質沉積。但兩者作用的分子機製有所不同。胰島素激活 AMPK，通過抑製SREBP-1c錶達、直接燐痠化 SREBP-1c 抑製SREBP-1c入覈等短期和長期的作用以及抑製內質網應激影響SREBP-1c，減少脂質閤成；而格列齊特僅通過燐痠化的短期作用和抑製內質網應激對SREBP-1c產生影響，且對脂肪痠氧化無作用。
목적：탐토이도소급격렬제특치료2형당뇨병대대서간장지질침적적영향급궤제。방법：제비고지급련뇨좌균소유도적2형당뇨병대서모형，수궤분위당뇨병조、이도소조화격렬제특조，병설정상대조조。통과간장유홍O염색관찰기간세포지질침적정황；ELISA검측혈청지련소수평；실시형광정량PCR검측간장지련소수체1（AdipoR1）mRNA표체；Western blotting검측간장선감산활화단백격매（AMPK）、린산화적선감산활화단백격매（Thr172p-AMPK）、고순조절인자결합단백1c（SREBP-1c）、린산화적고순조절인자결합단백1c（Ser372p-SREBP1-1c）、을선보매A최화매（ACC）、린산화적을선보매A최화매（Ser79p-ACC）화면역구단백결합단백（BiP）적표체。결과：당뇨병조간세포지질침적교정상대조조명현증다，이도소화격렬제특치료후간세포지질침적명현개선。이도소치료후，혈청지련소적수평급간장AdipoR1 mRNA수평교당뇨병조화정상대조조현저승고（P＜0．01），이격렬제특치료후량자수평회복지정상대조조수평。 Western blotting 결과현시，당뇨병대서여정상대조조비교，간장Thr172p-AMPK／AMPK화Ser372p-SREBP-1c／SREBP-1c화Ser79p-ACC／ACC표체명현강저（P＜0．01），BiP표체명현승고（P＜0．01）。이도소치료후，Thr172p-AMPK／AMPK화Ser372p-SREBP-1c／SREBP-1c현저승고（P＜0．01）， Ser79p-ACC／ACC화BiP단백표체회복지정상대조조수평。이격렬제특치료후Thr172p-AMPK／AMPK화Ser372p-SREBP-1c／SREBP-1c회복지정상대조조수평，BiP단백표체현저하강（P＜0．01），Ser79p-ACC／ACC여당뇨병조비교무명현개선。결론：이도소화격렬제특치료균능통과격활지련소-AMPK감경2형당뇨병대서간장적지질침적。단량자작용적분자궤제유소불동。이도소격활 AMPK，통과억제SREBP-1c표체、직접린산화 SREBP-1c 억제SREBP-1c입핵등단기화장기적작용이급억제내질망응격영향SREBP-1c，감소지질합성；이격렬제특부통과린산화적단기작용화억제내질망응격대SREBP-1c산생영향，차대지방산양화무작용。
AIM:To investigate the effect of insulin and gliclazide therapies on the liver fat accumulation in type 2 diabetic rats .METHODS:A high-fat diet plus low-dose streptozotocin was implemented to establish a type 2 dia-betic rat model, and the rats were randomly divided into diabetes mellitus (DM) group, diabetic rats treated with insulin ( INS) group, diabetic rats treated with gliclazide per os ( PO) group, and normal control ( NC) group.The diabetic rats in INS group and PO group were given insulin and gliclazide for 3 weeks, respectively.The changes of the liver fatty were evaluated with oil red O staining .Fasting plasma adiponectin concentration was measured by ELISA .The expression of adi-ponectin receptor 1 ( AdipoR1 ) was detected by real-time PCR.The protein levels of AMP-activated protein kinase (AMPK), phosphorylated AMPK on threonine 172 ( Thr172p-AMPK), sterol regulatory element-binding protein 1c (SREBP-1c), phosphorylated SREBP-1c on serine 372 (Ser372p-SREBP-1c), acetyl-CoA carboxylase (ACC), phospho-rylated ACC on serine79 (Ser79p-ACC) and immunoglobulin-binding protein (BiP) in the liver homogenate were deter-mined by Western blotting .RESULTS:Compared with the normal rats , in DM group, the presence of cytoplasmic lipid deposits was confirmed by oil red O staining .In INS group, these changes were significantly lower than those in DM group . Similar results were obtained in PO group .Insulin therapy significantly increased the plasma concentration of diponectin and liver tissue levels of AdipoR1 compared with DM group.At the same time, these 2 indicators returned to normal levels after gliclazide therapy .Thr172p-AMPK/AMPK, Ser372p-SREBP-1c/SREBP-1c and Ser79p-ACC/ACC expression ratios were significantly reduced in DM group compared with control values .The expression of BiP was increased on the contrary . After insulin therapy, Thr172p-AMPK/AMPK and Ser372p-SREBP-1c/SREBP-1c were significantly increased, and Ser79p-ACC/ACC and BiP returned to the normal levels .After gliclazide treatment, Thr172p-AMPK/AMPK and Ser372p-SREBP-1c/SREBP-1c returned to the normal levels , the expression ratio of Ser79p-ACC/ACC had no significant improve-ment compared with DM group , and the expression of BiP significantly declined .CONCLUSION: Both the insulin and gliclazide therapies reduce the lipid deposition in the liver of rats with type 2 diabetes by activating AMPK , but the extent and mechanism are not the same.In insulin therapy, AMPK restrains the expression of SREBP-1c directly, increases the phosphorylation of SREBP-1c, and affects SREBP-1c by inhibiting the endoplasmic reticulum stress .Gliclazide treatment, which has no effect on the lipid oxidation , reduces lipid deposition in the liver only through the phosphorylation of SREBP-1c and the suppression of the endoplasmic reticulum stress .