González-Muniesa P, Milagro F I, Campión J, Martínez J A
Department of Physiology and Nutrition, University of Navarra, 31008 Pamplona, Spain.
J Physiol Biochem. 2005 Jun;61(2):389-93. doi: 10.1007/BF03167056.
The UCP1 is an uncoupling protein located in the inner mitochondrial membrane of brown adipocytes, which has a well-documented role in diet-induced thermogenesis. The current study assessed whether UCP1 transfected liver cells demand more fuel substrates in the oxidative phosphorylation processes. Therefore, the purpose of this experiment was to achieve an ectopic expression of UCP1 in HepG2 cells to significantly decrease the production of ATP. The UCP1 gene was transferred into the hepatic cells by using a calcium phosphate precipitation protocol. The efficiency of the transfection was tested, 48 hours later, by bioluminescence of luciferase previously transfected, while the expression of mRNA of UCP1 was demonstrated by RT-PCR. In addition, measuring the production of ATP by using a bioluminescence procedure assessed the functionality of this protein. Transfected liver cells with UCP1 showed a decrease of 23% in ATP production in comparison with control cells without expression of UCP1 (2.23 vs. 2.90 RLU/pg protein, p=0.015). In conclusion, the ectopic expression of UCP1 decreased the production of ATP, possibly uncoupling the oxidative phosphorylation, which could be a novel approach for understanding thermogenic processes and eventually for energy metabolism and body weight management.
解偶联蛋白1（UCP1）是一种位于棕色脂肪细胞线粒体内膜的解偶联蛋白，在饮食诱导的产热过程中发挥着有充分记录的作用。本研究评估了转染UCP1的肝细胞在氧化磷酸化过程中是否需要更多的燃料底物。因此，本实验的目的是在HepG2细胞中实现UCP1的异位表达，以显著降低ATP的产生。通过磷酸钙沉淀法将UCP1基因导入肝细胞。48小时后，通过先前转染的荧光素酶的生物发光检测转染效率，同时通过RT-PCR证明UCP1的mRNA表达。此外，使用生物发光程序测量ATP的产生评估了该蛋白的功能。与未表达UCP1的对照细胞相比，转染UCP1的肝细胞的ATP产生减少了23%（2.23对2.90相对光单位/微克蛋白，p = 0.015）。总之，UCP1的异位表达降低了ATP的产生，可能使氧化磷酸化解偶联，这可能是理解产热过程以及最终能量代谢和体重管理的一种新方法。
