GLP-1类似物利拉鲁肽对高糖诱导胰岛细胞凋亡及HSP72表达、ERK1/2通路活性的影响

目的 探讨胰高血糖素样多肽(GLP)-1类似物利拉鲁肽对高糖诱导胰岛细胞凋亡及热休克蛋白(HSP)72表达、细胞外信号调节激酶(ERK)1/2通路的影响。方法 体外培养大鼠胰岛INS-1细胞，分为对照组、高糖组、利拉鲁肽低剂量组(1 nmol/L)、利拉鲁肽中剂量组(3 nmol/L)和利拉鲁肽高剂量组(5 nmol/L)。检测各组INS-1细胞凋亡、HSP72、ERK1、ERK2、Bcl-2相关X蛋白(Bax)mRNA表达和HSP72、p-ERK1/2、ERK1/2、Bax蛋白表达情况。结果 与对照组相比，高糖组INS-1细胞凋亡率、HSP72、ERK1/2、Bax mRNA及HSP72、p-ERK1/2/ERK1/2、Bax蛋白表达水平显著升高(P<0.05);与高糖组相比，利拉鲁肽低、中、高剂量组INS-1细胞凋亡率、HSP72、ERK1/2、Bax mRNA及HSP72、p-ERK1/2/ERK1/2、Bax蛋白表达水平均显著降低(P<0.05);随着利拉鲁肽使用剂量升高，INS-1细胞凋亡率、HSP72、ERK1/2、Bax mRNA及HSP72、p-ERK1/2...     

利拉鲁肽降低心肌微循环内皮细胞缺血再灌注损伤的机制研究

目的探讨利拉鲁肽降低微循环缺血再灌注损伤的机制。方法体外培养人脐静脉血内皮细胞系,并建立细胞缺氧复氧(HR)模型。正常对照组(Con组,正常培养),HR组(等量PBS),低剂量利拉鲁肽组(低Lir组,加入利拉鲁肽终浓度1nmol/L),中剂量利拉鲁肽组(中Lir组,加入利拉鲁肽终浓度10nmol/L),高剂量利拉鲁肽组(高Lir组,加入利拉鲁肽终浓度100nmol/L)。小干扰RNA技术敲低三磷酸肌醇受体(IP3R)和VDAC蛋白表达,并作为干扰IP3R组和干扰VDAC组。MTT法检测细胞活性;Fura-2AM检测细胞内钙超载([Ca~(2+)]c)情况;Western blot检测钙泵蛋白IP3R;观察利拉鲁肽作用下线粒体钙([Ca~(2+)]m)的浓度变化。结果与Con组比较,HR组促进IP3R表达增加和[Ca~(2+)]c超载(P0.05),表现为Fura-2AM荧光增强(P0.05);同时[Ca~(2+)]c超载现象通过VDAC进入线粒体,诱发[Ca~(2+)]m超载、膜电位丢失、膜孔道开放增加、细胞色素C释放,最终TUNEL阳性凋亡细胞增加(P0.05)。与HR组比较,低、中和高Lir组减少IP3R表达(P0.05),抑制[Ca~(2+)]c/[Ca~(2+)]m超载,降低细胞凋亡(P0.05)。结论 HR通过激活IP3R-[Ca~(2+)]c/VDAC-[Ca~(2+)]m信号通路诱导内皮细胞凋亡,利拉鲁肽可以通过抑制上述信号保护微循环内皮,降低再灌注损伤。     

利拉鲁肽通过调节SIRT1改善高糖诱导的大鼠H9c2细胞氧化应激损伤

目的探究利拉鲁肽(liraglutide, LRG)对高糖诱导的心肌细胞(H9c2)氧化应激损伤的影响及其潜在机制。方法采用高糖处理H9c2细胞24 h建立心肌细胞体外损伤模型, 给予不同浓度利拉鲁肽(10、100、1 000 nmol/L)干预, 用CCK-8检测细胞活力, 倒置显微镜观察细胞形态结构的改变。利拉鲁肽(100 nmol/L)干预高糖处理H9c2细胞24 h后检测细胞上清液中乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)和丙二醛含量;RT-PCR和Western印迹法检测沉默信息调节因子1(SIRT1)和叉头转录因子1(FOXO1)mRNA及蛋白水平;Western印迹法检测FOXO1蛋白乙酰化水平;应用小干扰RNA(siRNA)技术沉默SIRT1的H9c2细胞株验证SIRT1在其中的作用。结果与对照组相比, 高糖组细胞活力降低, 细胞结构受损, 细胞上清液中LDH、丙二醛含量显著升高, SOD含量降低, 氧化应激加重, SIRT1表达降低, FOXO1乙酰化水平升高(均P<0.05);与高糖组比较, 给予利拉鲁肽干预后细胞活力升高, 心肌细胞结构形态和氧化应激水...     

利拉鲁肽对2型糖尿病大鼠胰岛β细胞凋亡的调控机制

目的探讨利拉鲁肽对2型糖尿病大鼠胰岛β细胞凋亡的调控机制。方法将60只SPF级SD雄性大鼠随机分为正常对照组、模型组、阳性对照组(二甲双胍,140 mg·kg~(-1)·d~(-1))、利拉鲁肽低剂量组(0. 12 mg·kg~(-1)·d~(-1))和利拉鲁肽高剂量组(6. 30 mg·kg~(-1)·d~(-1)),每组12只。除正常对照组外,其余各组均采用高脂、高糖饲养,同时联合腹腔注射链脲佐菌素(STZ,30 mg/kg),建立大鼠2型糖尿病模型。成功建模7 d后,利拉鲁肽高、低剂量组皮下注射利拉鲁肽;阳性对照组灌胃给予二甲双胍(140 mg·kg~(-1)·d~(-1));正常对照组和模型组均灌胃给予等量的生理盐水(10 ml·kg~(-1)·d~(-1)),1次/d。治疗8 w后,检测各组空腹血糖浓度、胰岛素和抵抗素水平,免疫组化法检测各组胰岛β细胞Bcl-2和Bax蛋白表达水平,Western印迹测定各组胰岛β细胞Caspase-3蛋白的表达水平。结果与正常对照组比较,模型组空腹血糖浓度、胰岛素和抵抗素水平、Bax蛋白和Caspases-3蛋白的表达水平显著升高(P<0. 05),Bcl-2表达水平明显降低(P<0. 05);与模型组比较,利拉鲁肽组和阳性对照组空腹血糖浓度、胰岛素和抵抗素水平、Bax蛋白和Caspases-3蛋白表达水平显著降低(P<0. 05),Bcl-2蛋白表达水平显著升高(P<0. 05)。结论利拉鲁肽能显著改善2型糖尿病大鼠胰岛素抵抗、抑制胰岛β细胞凋亡,其机制可能与提高Bcl-2表达,下调Bax表达,进而降低Caspase-3的活性有关。     

激动β_3肾上腺素能受体上调载脂蛋白A-Ⅰ促进泡沫细胞胆固醇流出的研究

目的 通过激动或抑制HepG2细胞的β3肾上腺素能受体(β3-AR),探讨β3-AR调节胆固醇逆转运过程的可能机制。方法 将培养的HepG2细胞随机分为对照组、β3-AR激动剂组(激动剂组)和β3-AR拮抗剂组(拮抗剂组),ELISA法检测上清液载脂蛋白(apo)A-Ⅰ、apoA-Ⅱ及β3-AR水平;测定细胞内胆固醇、游离胆固醇和胆固醇酯水平,3 H标记的胆固醇测定胆固醇流出率,实时定量PCR和蛋白印迹法分别检测细胞中三磷酸腺苷结合盒转运蛋白A1(ABCA1)和肝X受体α亚型(LXRα)的表达。结果 与对照组比较,激动剂组apoA-Ⅰ、游离胆固醇、胆固醇流出率显著增加,胆固醇、胆固醇酯显著降低,ABCA1和LXRαmRNA及ABCA1和LXRα蛋白显著增加;拮抗剂组胆固醇、胆固醇酯显著升高,apoA-Ⅰ、胆固醇流出率显著减少,ABCA1和LXRαmRNA及ABCA1和LXRα蛋白显著降低。与激动剂组比较,拮抗剂组ABCA1和LXRαmRNA及ABCA1和LXRα蛋白显著降低(0.49±0.10 vs1.24±0.02,0.85±0.05 vs 1.32±0.05,0.38±0.01 vs 1.45±0.20,0.08±0.01 vs 0.76±0.02,P0.01)。结论激动HepG2细胞的β3-AR,可上调apoA-Ⅰ表达,促进巨噬细胞源性泡沫细胞胆固醇逆转运相关蛋白的表达。     

利拉鲁肽经AMPK途径对平滑肌细胞KCa3.1蛋白表达的影响

目的探讨利拉鲁肽经激活腺苷酸活化的蛋白激酶(AMPK)途径对平滑肌细胞KCa3.1蛋白表达的影响。方法制备SD大鼠血管平滑肌细胞模型,大鼠主动脉平滑肌细胞采用利拉鲁肽进行干预,测定未加入利拉鲁肽及加入利拉鲁肽15 min后磷酸化AMPK(PAMPK)/AMPK比值。将平滑肌细胞分为4组培养:A组为正常组,B组采用血管紧张素Ⅱ100 nmol/L,C组采用利拉鲁肽1μmol/L+血管紧张素Ⅱ100 nmol/L,D组采用利拉鲁肽1μmol/L+AMPK阻滞剂4μmol/L+血管紧张素Ⅱ100 nmol/L。培养72 h后使用Western-Blotting检测KCa3.1蛋白表达。结果显微镜下可见细胞呈长梭状、三角状,α-actin细胞免疫荧光染色后,胞浆着色为红色,确认为血管平滑肌细胞。利拉鲁肽与大鼠血管平滑肌作用15 min后,与基线比较PAMPK水平明显上升,PAMPK与AMPK比值明显上调,差异有统计学意义(P0.05)。C组平滑肌细胞KCa3.1蛋白表达较B组下降(P0.05)。结论利拉鲁肽可激活AMPK细胞通路,抑制平滑肌细胞上KCa3.1蛋白表达。     

利拉鲁肽通过抑制核因子κB p65磷酸化调控人脐静脉内皮细胞一氧化氮合酶的表达

目的 探讨利拉鲁肽在高糖环境下对人脐静脉内皮细胞（HUVECs）功能的影响及其可能机制.方法 高糖环境下（25 mmol/L葡萄糖）培养人脐静脉内皮细胞,分别给予10、100、1 000 μg/L利拉鲁肽进行干预,采用实时定量聚合酶链反应（RT-PCR）和Western blotting法分别检测内皮型一氧化氮合酶（eNOS）、诱导型一氧化氮合酶（iNOS）以及核因子κB p65 （NF-κB p65）的mRNA、蛋白表达水平;应用肿瘤坏死因子-α（TNF-α）干预利拉鲁肽处理后的HUVECs,观察eNOS、iNOS、NF-κB p65的mRNA、蛋白表达水平的变化.研究结果多组间采用单因素方差分析,两组间比较采用SLD分析.结果 与普通培养基（7 mmol/L葡萄糖）组相比,高糖环境下HUVECs的eNOS mRNA和蛋白表达均降低,iNOS mRNA和蛋白表达及磷酸化NF-κB p65蛋白表达均增高（t=2.79、5.75、4.32、4.85、7.12,均P＜0.05）.与0μg/L组相比,1 000μg/L利拉鲁肽干预组HUVECs的eNOS mRNA、eNOS蛋白表达均上调,iNOS mRNA、iNOS蛋白表达及NF-κB p65磷酸化蛋白表达均下调（t=5.12、9.34、6.70、5.50、8.94,均P＜0.05）.与单独使用利拉鲁肽组相比,TNF-α联合利拉鲁肽组HUVECs的eNOSmRNA和蛋白表达均下调,iNOS mRNA和蛋白表达及磷酸化NF-κB p65蛋白表达均上调（t=3.33～7.87,均P＜0.05）.结论 利拉鲁肽可通过抑制NF-κB p65磷酸化在转录和翻译水平上增加eNOS表达、降低iNOS的表达,从而改善内皮细胞功能,预防糖尿病动脉粥样硬化.     

胆固醇逆向转运关键蛋白介导药物和小分子调控脂代谢的机制研究

胆固醇逆向转运(RCT)异常导致脂代谢紊乱是动脉粥样硬化发病重要环节,探讨介导RCT的关键蛋白在脂代谢中的作用及调控机制对阐明动脉粥样硬化发病的分子机制具有重要意义。本期专题收集的论文探讨了利拉鲁肽、苦瓜素以及生长分化因子11对细胞胆固醇流出及介导RCT中关键蛋白三磷酸腺苷结合盒转运体A1(ABCA1)、ABCG1和B类Ⅰ型清道夫受体表达的影响,分别从不同角度阐释RCT关键蛋白参与药物或小分子物质调控胆固醇流出的分子机制。     

利拉鲁肽对非酒精性脂肪肝大鼠肝脏组织中胰岛素JNK1信号通路的影响

目的探究利拉鲁肽对非酒精性脂肪肝(NAFLD)大鼠肝脏组织中胰岛素JNK1信号通路的影响。方法选取40只6周龄SPF级大鼠,采用高脂饮食喂养12周建立NAFLD大鼠模型。将NAFLD大鼠随机分为空白对照组(control)、模型组(model)、低剂量利拉鲁肽组(low lira)和高剂量利拉鲁肽组(high lira)。利拉鲁肽干预18 d后,测量大鼠体质量及肝指数变化;全自动生化分析仪测定大鼠血清丙氨酸氨基转移酶(ALT)、空腹血糖(FBG)、甘油三酯(TG)、总胆固醇(TC)和血清胰岛素(FINS)变化;酶联免疫吸附法测定大鼠肝组织中肿瘤坏死因子(TNF-α)、超氧化物歧化酶(SOD)、丙二醛(MAD)及游离脂肪酸(FFAs)含量; Western blot检测大鼠肝脏组织中胰岛素受体(IR)、磷酸化胰岛素受体底物1(p-IRS1)、C-Jun氨基端激酶1(JNK1)及磷酸化C-Jun氨基端激酶1(p-JNK1)表达情况; HE染色观察大鼠肝脏组织病理学变化。结果与对照组相比,模型组大鼠体质量,肝指数,血清ALT、TG、TC、FINS、TNF-α、MAD、FFAs含量,p-IRS1、JNK1、p-JNK1蛋白表达量显著升高,SOD含量显著降低,差异有统计学意义(P 0. 01);血清FBG含量和IR蛋白表达量无显著变化(P 0. 05);相比模型组,利拉鲁肽组大鼠体质量,肝指数,血清ALT、TG、TC、FINS、TNF-α、MAD、FFAs含量,p-IRS1、JNK1、p-JNK1蛋白表达量显著降低,且高剂量组显著低于低剂量组(P 0. 01); SOD含量显著升高,且高剂量组显著高于低剂量组(P 0. 01);血清FBG含量和IR蛋白表达量无显著变化(P 0. 05)。结论利拉鲁肽可以缓解大鼠脂肪肝的发展,其作用机制可能与抑制JNK1信号通路及JNK1磷酸化相关,且在一定范围内呈浓度依赖性。     

慢性间歇性低氧对大鼠胰岛β细胞功能的影响及利拉鲁肽对胰岛β细胞的保护作用

目的观察慢性间歇性低氧对大鼠胰岛β细胞功能的影响,探讨胰高糖素样肽-1(GLP-1)类似物利拉鲁肽对胰岛β细胞的保护作用。方法 36只SD雄性大鼠随机分为常氧对照组12只,间歇性低氧组24只。造模第8周结束后,两组大鼠均进行胰岛β细胞原代提取,常氧对照组随机分为常氧组和高糖刺激组;间歇性低氧组随机分为模型组、利拉鲁肽(10、50、100 nmol/L)干预组。ELISA法检测各组大鼠胰岛β细胞上清中8-异前列腺素(8-ISO)及谷胱甘肽过氧化物酶(GSH-Px)表达水平;流式细胞仪检测各组胰岛β细胞凋亡率; Western-blot法检测胰岛β细胞凋亡相关蛋白Caspase-3表达。结果与正常对照组比较,模型组和高糖刺激组大鼠胰岛β细胞上清液中8-ISO水平升高,而GSH-Px在模型组及高糖刺激组低于正常对照组,差异有统计学意义(P 0. 05);同时,模型组及高糖刺激组胰岛β细胞凋亡率升高,Caspase-3在胰岛β细胞表达上调,差异有统计学意义(P 0. 05),但高糖刺激组较模型组差异更明显。与模型组比较,利拉鲁肽干预后胰岛β细胞上清液中8-ISO水平降低,而GSH-Px升高,差异有统计学意义(P 0. 05);药物干预后Caspase-3蛋白表达低于模型组,且高浓度组较中低浓度组变化更明显。结论间歇性低氧导致的氧化应激可能是睡眠呼吸暂停综合征胰岛功能损伤的重要机制,GLP-1类似物利拉鲁肽通过对氧化物质的调节,从而减轻胰岛β细胞凋亡,保护胰岛β细胞。     

Inflammatory stress exacerbates hepatic cholesterol accumulation via disrupting cellular cholesterol export

Background and Aim: Both inflammation and cholesterol accumulation play important roles in the development of non‐alcoholic fatty liver disease. This study was undertaken to investigate whether inflammation aggravated cholesterol accumulation via disrupting hepatic cholesterol export and we explored the underlying mechanisms. Methods: We used casein injection in C57BL/6J mice, and tumor necrosis factor alpha (TNF‐α) stimulation in human hepatoblastoma cell line (HepG2) cells to induce inflammation. Intracellular cholesterol level was examined by Oil Red O staining and quantitative analysis. Bile acid level was quantified by colorimetric analysis. ³[H] cholesterol assay by scintillation counting was performed to evaluate the cholesterol efflux. The mRNA and protein expression was examined by real‐time polymerase chain reaction and Western blot. Results: Inflammation increased cholesterol accumulation in livers of C57BL/6J mice and in HepG2 cells. High‐fat diet in mice and low‐density lipoprotein (LDL) loading in HepG2 cells increased bile acid synthesis and cholesterol efflux, enhanced the mRNA and protein expression of liver X receptor α (LXRα), peroxisome proliferator‐activated receptors (PPARα, γ), cholesterol 7α‐hydroxylase (CYP7A1) and ATP‐binding cassette transporter A1 (ABCA1). However, inflammation reduced bile acid synthesis and cholesterol efflux even in high‐fat‐diet‐fed mice and HepG2 cells in the presence of LDL loading. The enhanced effects of these genes and proteins expression due to high‐fat diet and LDL loading were inhibited by inflammation both in vivo and in vitro. Conclusions: Inflammation disrupted PPAR‐LXR‐CYP7A1/ABCA1‐mediated bile acid synthesis and cholesterol efflux resulting in exacerbated cholesterol accumulation in livers of C57BL/6J mice and HepG2 cells.     

高糖对THP-1巨噬细胞ABC转运体的表达和功能的影响

目的:探讨高糖对体外培养的THP-1巨噬细胞中三磷酸腺苷结合盒（ABC）转运体的表达及功能的影响。方法: 以不同浓度的D-葡萄糖干预培养的THP-1单核巨噬细胞5 d,用实时定量PCR和Western blot检测巨噬细胞中ABCG1、ABCA1 mRNA和其蛋白的表达。用酶荧光化学法检测培养基中及细胞内胆固醇的含量。结果: 高糖可抑制巨噬细胞中ABCG1的表达,但是对ABCA1的表达影响不明显。随着D-葡萄糖浓度的增加,从巨噬细胞中流出的胆固醇量减少,同时细胞内胆固醇的含量增加。结论: 高糖可抑制巨噬细胞中ABCG1的表达及功能,有助于促进巨噬细胞内脂质堆积。     

Molecular interaction between caveolin-1 and ABCA1 on high-density lipoprotein-mediated cholesterol efflux in aortic endothelial cells

OBJECTIVE: Caveolin-1 and ATP-binding cassette transporter A1 (ABCA1) are proteins that are involved in cellular cholesterol efflux. In this study, we analyzed the relationships between caveolin-1 and ABCA1 on high-density lipoprotein (HDL)-mediated cholesterol efflux in rat aortic endothelial cells. METHODS AND RESULTS: Overexpression of caveolin-1 by transfection with caveolin-1 cDNA in aortic endothelial cells up-regulated ABCA1 expression and enhanced cholesterol efflux. Suppression of caveolin-1 by siRNA decreased ABCA1 expression and reduced cholesterol efflux. The number of caveolae increased after transfection with caveolin-1 into cells. Immunoprecipitation assays revealed a molecular interaction between caveolin-1 and ABCA1 in the plasma membrane and in the cytoplasm after HDL incubation. Immunoelectron microscopy demonstrated that caveolin-1 colocalized with ABCA1 in the caveolae and in the cytoplasmic vesicles; it was also found that caveolin-1 and ABCA1 colocalized with cellular cholesterol by immunofluorescence microscopy. Blocking of intracellular lipid transport by inhibitors disrupted the interaction between caveolin-1 and ABCA1 and reduced cholesterol to methyl-beta-cyclodextrin and HDL. CONCLUSIONS: The molecular interaction between caveolin-1 and ABCA1 is associated with the HDL-mediated cholesterol efflux pathway in aortic endothelial cells.     

脂联素对泡沫细胞三磷酸腺苷结合盒转运体A1表达及细胞内胆固醇含量的影响

目的通过研究脂联素对细胞内胆固醇外流的影响以及三磷酸腺苷结合盒转运子A1的功能变化,探讨其抗动脉粥样硬化作用的可能机制。方法以THP-1来源的泡沫细胞模型为研究对象,用脂联素进行体外干预;采用逆转录聚合酶链反应、酶联免疫吸附试验和高效液相色谱等方法测定干预前后三磷酸腺苷结合盒转运子A1表达和细胞内胆固醇含量的变化。结果脂联素在体外上调泡沫细胞三磷酸腺苷结合盒转运子A1mRNA和蛋白的表达,与载脂蛋白A1联合作用还能进一步增加细胞内胆固醇的流出。脂联素单独作用促进细胞内胆固醇酯转化为游离胆固醇。结论脂联素上调泡沫细胞三磷酸腺苷结合盒转运子A1表达,并促进其介导的细胞内游离胆固醇的外流;脂联素还能促进细胞内胆固醇酯的水解。     

TNFalpha induces ABCA1 through NF-kappaB in macrophages and in phagocytes ingesting apoptotic cells

Recent evidence suggests that tumor necrosis factor alpha (TNFalpha) signaling in vascular cells can have antiatherogenic consequences, but the mechanisms are poorly understood. TNFalpha is released by free cholesterol-loaded apoptotic macrophages, and the clearance of these cells by phagocytic macrophages may help to limit plaque development. Macrophage cholesterol uptake induces ATP-binding cassette (ABC) transporter ABCA1 promoting cholesterol efflux to apolipoprotein A-I and reducing atherosclerosis. We show that TNFalpha induces ABCA1 mRNA and protein in control and cholesterol-loaded macrophages and enhances cholesterol efflux to apolipoprotein A-I. The induction of ABCA1 by TNFalpha is reduced by 65% in IkappaB kinase beta-deficient macrophages and by 30% in p38alpha-deficient macrophages, but not in jun kinase 1 (JNK1)- or JNK2-deficient macrophages. To evaluate the potential pathophysiological significance of these observations, we fed TNFalpha-secreting free cholesterol-loaded apoptotic macrophages to a healthy macrophage monolayer (phagocytes). ABCA1 mRNA and protein were markedly induced in the phagocytes, a response that was mediated both by TNFalpha signaling and by liver X receptor activation. Thus, TNFalpha signals primarily through NF-kappaB to induce ABCA1 expression in macrophages. In atherosclerotic plaques, this process may help phagocytic macrophages to efflux excess lipids derived from the ingestion of cholesterol-rich apoptotic corpses.     

可溶性环氧化物水解酶抑制剂对脂肪细胞胆固醇流出的影响

目的 观察可溶性环氧化合物水解酶抑制剂(sEHi)tAUCB对脂肪细胞胆固醇流出的影响,并探讨其机制.方法 测定脂肪细胞过氧化物酶体增殖物激活受体γ(PPARγ)及三磷酸腺苷结合盒转运体A1(ABCA1)mRNA及蛋白的表达,检测细胞内胆固醇流出.结果 tAUCB呈剂量依赖性促进载脂蛋白(Apo)A1介导的胆固醇流出,1、10、50、100 μmol/L浓度的tAUCB干预后,胆固醇流出率分别为(5.93±0.66)%,(7.40±0.43)%,(8.30±0.34)%和(9.77±0.42)%.加入10～100μmol/L tAUCB干预组与空白组(5.67±0.17)%比较,胆固醇流出差异有统计学意义(P＜0.05).同时发现随着tAUCB干预浓度的增加,脂肪细胞ABCA1、PPARy mRNA及蛋白的表达也呈剂量依赖性地升高,而GW9662明显抑制tAUCB对脂肪细胞胆固醇流出及ABCA1和PPARγ表达的促进作用.结论 tAUCB可通过上调PPARy的表达,促进脂肪细胞Apo A1-ABCA1通路加速细胞内胆固醇流出,抑制脂肪细胞内胆固醇过度蓄积.

Abstract：

Objective To observe the effects of soluble epoxide hydrolase inhibitors tAUCB on cholesterol efflux in adipocytes. Methods 3T3-L1 preadipocytes were induced to differentiation and maturation. Cells were stimilated with 100μg/L LPS after starved for 24 hours, then tAUCB in various concentrations(1 ,10,50,100 μmol/L)were added for 24 h, or incubated with the peroxisome proliferator activated receptor gamma (PPARy) antagonist GW9662 (5 μmol/L).0μmol/L tAUCB treated group was taken as empty control. After then, the mRNA expression of PPARγ and adenosine triphosphate binding cassette transporter Al (ABCA1) in cells were determined via realtime-PCR, the amounts of protein expression of PPARγand ABCA1 in cells were detected by Western blot, the efflux rates of 3H-cholesterol in cells were detected by means of liquid scintillation counter. Results tAUCB could dose-dependently increase the apolipoprotein A1 (apoA1)-mediated cholesterol efflux in adipocytes. After stimulated by 1, 10,50,100 μmol/L tAUCB, cholesterol efflux rates were (5.93±0.66) %, (7.40 ± 0. 43) %, (8. 30 ±0. 34)% ,(9.77±0.42)% respectively, there were significant difference after treated by 10-100 μmol/L tAUCB compared with control(5.67±0.17)%(P＜0.05). With the concentration of tAUCB increased,ABCA1, PPAR mRNA and protein expression were also dose-dependently up-regulated. GW9662 could significantly inhibit the effects of tAUCB, and then reduce the cholesterol efflux and the expression of PPARγ and ABCA1 in adipocytes. Conclusions tAUCB could up-regulate PPARγ expression in adipocytes, and promote the cholesterol efflux of adipocytes via apoA1-ABCA1 pathway, which might decrease the cellular cholesterol accumulation in adipocytes.