二脒那秦内源性激活血管紧张素转化酶2对细胞脂质沉积的抑制效应及其机制

旨在通过诱导3T3-L1前脂肪细胞转分化为3T3-L1脂肪细胞，探讨二脒那秦（DIZE）内源性激活血管紧张素转化酶2（ACE 2）对细胞脂质沉积的抑制效应及机制。对3T3-L1前脂肪细胞转分化，将分化成功的3T3-L1脂肪细胞分为对照组、DIZE组（12.5、25 μmol·L^-1处理），siACE2组和siACE2+DIZE组（siACE2干扰后+25 μmol·L^-1 DIZE），作用48 h，取上清和细胞进行试验：1）测定细胞上清中三酰甘油和葡萄糖含量；2）油红O染色细胞，并进行定量分析；3） Western blot检测细胞内ACE2和脂肪酸合成关键酶或因子FAS、ACC和SREBP-1c及葡萄糖转运蛋白GLUT4与氧化分解关键酶CS的蛋白表达水平。结果显示：1）成功诱导得到了3T3-L1脂肪细胞，前脂肪细胞诱导至第14天，有95%以上细胞内出现脂滴；2）确定了DIZE作用浓度为12.5和25 μmol·L^-1，处理时间为48 h，并用该药物浓度及时间处理3T3-L1脂肪细胞，细胞上清中三酰甘油含量显著降低（P<0.05），葡萄糖含量显著升高（P<0.05）；25 μmol·L^-1 DIZE处理组细胞脂滴减少，siACE2组无显著变化，siACE2+DIZE组脂滴蓄积介于对照组和DIZE组之间； 25 μmol·L^-1 DIZE组ACE2蛋白水平显著高于对照组（P<0.05）；siACE2组显著下调（P<0.05），siACE2+DIZE组较siACE2组无明显变化；3）与对照组相比25 μmol·L^-1 DIZE组细胞中FAS、ACC和SREBP-1c蛋白表达下调，siACE2组和siACE2+DIZE组则表达均上调；4）与对照组相比，25 μmol·L^-1 DIZE组GLUT4和CS蛋白表达水平显著上调（P<0.05），siACE2组和siACE2+DIZE组则均显著下调（P<0.05）。综上所述，DIZE处理通过介导脂肪细胞ACE2的内源性激活改善了脂肪细胞的脂肪沉积。其机理：一方面抑制脂质合成；另一方面促进葡萄糖摄取和氧化代谢，两方面协同减少了脂肪沉积。结果提示，ACE2或DIZE可作为防控脂肪沉积发生和发展的潜在靶点或药物。

Inhibitory Effect and Mechanism of Endogenous Activation of Angiotensin Converting Enzyme 2 by Diminazene Aceturate on Cell Lipid Deposition

In this study, 3T3-L1 preadipocytes were induced to transdifferentiate into 3T3-L1 adipocytes to explore the inhibitory effect and mechanism of diminazene aceturate (DIZE) endogenous activation of angiotensin converting enzyme 2 (ACE2) on cell lipid deposition. 3T3-L1 preadipocytes were transdifferentiated. The successfully differentiated 3T3-L1 adipocytes were divided into control group and DIZE group (12.5, 25 μmol·L^-1 treatment), siACE2 group and siACE2 + DIZE group (after siACE2 interference + 25 μmol·L^-1 (dice) for 48 h, supernatant and cells were taken for the following experiments. Results:1) 3T3-L1 adipocytes were successfully induced. Fat droplets appeared in more than 95% of preadipocytes on the 14th day; 2) The action concentrations of DIZE were determined to be 12.5 and 25 μmol·L^-1 When 3T3-L1 adipocytes were treated with the drug concentration and time for 48 hours, the content of triglyceride in cell supernatant decreased significantly (P<0.05) and the content of glucose increased significantly (P<0.05); The lipid droplets of cells in the 25 μmol·L^-1 DIZE treatment group decreased, but the siACE2 group had no significant change. The accumulation of lipid droplets in the siACE2+DIZE group was between the control group and the DIZE group; the ACE2 protein level was significantly higher in the 25 μmol·L^-1 DIZE group. Compared with the control group (P<0.05); the siACE2 group was significantly down-regulated (P<0.05), and the siACE2+DIZE group had no significant change compared with the siACE2 group; 3) Compared with the control group, the expressions of FAS, ACC and SREBP-1c proteins in the 25 μmol·L^-1 DIZE group were down-regulated, while those in the siACE2 group and siACE2+DIZE group were all up-regulated.; 4) Compared with the control group, the expression levels of GLUT4 and CS proteins in the 25 μmol·L^-1 DIZE group were significantly up-regulated (P<0.05), while those in the siACE2 group and siACE2+DIZE group were significantly down-regulated (P<0.05). DIZE treatment improved adipocyte fat deposition by mediating endogenous activation of adipocyte ACE2. Its mechanism:on the one hand, it inhibits lipid synthesis; on the other hand, it promotes glucose uptake and oxidative metabolism, which synergistically reduces fat deposition. The results suggest that ACE2 or DIZE can be used as potential targets or drugs to prevent and control the occurrence and development of fat deposition.