Influence of cyanide p-trifluoromethoxyphenyl-hydrazone on insulin sensitivity and mitochondrial function in 3T3-L1 adipocytes
10.3969/cma.j.issn.2095-428X.2013.19.006
- VernacularTitle:羰基-氰-对-三氟甲氧基本腙对成熟脂肪细胞胰岛素敏感性和线粒体功能的影响
- Author:
Chun-Mei SHI
1
;
Chen-Bo JI
;
Yu-Mei WANG
;
Ling CHEN
;
Lei YANG
;
Guang-Feng XU
;
Xi-Rong GUO
Author Information
1. 南京医科大学儿科研究所
- Keywords:
Obesity;
Cyanide p-trifluoromethoxyphenyl-hydrazone;
Adipocytes;
Mitochondria function;
Insulin sensitivity
- From:
Chinese Journal of Applied Clinical Pediatrics
2013;28(19):1455-1458
- CountryChina
- Language:Chinese
-
Abstract:
Objective To observe the effect of uncoupler of mitochondrial oxidative phosphorylation——Cyanide p-trifluoromethoxyphenyl-hydrazone (FCCP) on mitochondrial function and insulin sensitivity in mature adipocytes.Me-thods 3T3-L1 pre-adipocytes were differentiated into mature adipocytes and then induced and maintained in medium that contained the chemical uncoupler 7.5 μmol/L FCCP.Glucose uptake was determined in the adipocytes by measu-ring 2-deoxy-D-[3H] glucose uptake.Western blot was used to detect the translocation of insulin-sensitive glucose transporter 4 (GLUT4) and measure the phosphorylation and total protein contents of insulin signaling proteins such as insulin receptor substrate(IRS)-1,Akt.The mitochondrial morphology was performed by transmission electron microscope.The mitochondrial DNA (mtDNA) copy number was evaluated by real time PCR.Luciferase-based luminescence assay was used to determine cellular ATP production.The mitochondrial membrane potential(△Ψm) and reactive oxygen species(ROS) were detected by flow cytometry.Results (1) Exposure of mature adipocytes to FCCP basal glucose uptake was similar to mature adipocytes without FCCP(t =-0.07,P ＞ 0.05) ; however,the insulin-stimulated glucose uptake was significantly decreased in FCCP group (t =5.87,P ＜ 0.01).(2)FCCP decreased insulin-stimulated GLUT4 translocation to the plasmalemma and inhibition of insulin-induced phosphorylation of IRS-1 and Akt in 3T3-L1 adipocytes.(3)The size of mitochondria in FCCP-treated adipocytes was smaller than that in 3T3-L1 adipocytes without FCCP,and the morphology was condensed and abnormal.(4) mtDNA copy number in FCCP-treated adipocytes was significantly lower than that in adipocytes without FCCP(t =-1.73,P ＜ 0.001).(5) Exposure of 3T3-L1 adipocytes to FCCP significantly decreased △Ψm (t =4.83,P ＜ 0.01) and total cellular ATP production compared with cells without FCCP (t =6.08,P ＜ 0.0001),as well as the increased intracellular ROS levels (t =-6.82,P ＜ 0.01).Conclusions FCCP may impair mitochondrial morphology and mitochondrion dysfunction,and inhibited the activation of insulin-induced phosphorylation of IRS-1 and Akt in 3T3-L1 adipocytes,which suggests that FCCP-induced insulin resistance may be correlated with the FCCP induced generation of ROS in 3T3-L1 adipocytes.
