【作者】 刘畅；

【导师】 李冬海；

【作者基本信息】 南京大学 ， 生物化学和分子生物学， 2017， 硕士

【摘要】 解偶联是线粒体在氧化磷酸化过程中发生的一个反应。它主要能激活基底氧化反应并消耗氧化能量来产生热量。呼吸作用解偶联及消耗能量产热对于机体的能量平衡和体重控制有着重要作用,对肥胖等新陈代谢相关疾病具有积极意义,并且能够阻止线粒体产生的氧自由基(ROS)的积累。解偶联蛋白是位于线粒体内膜上的一类负责氧化磷酸化解偶联的载体蛋白。解偶联蛋白最主要的作用是参与体温调控,通过消耗能量转化为热量来维持体内热平衡。解偶联蛋白还参与抑制体内不良的活性氧的生成。非震颤性生热是哺乳动物在进化过程中获得的应对寒冷环境的生热方式,其与解偶联蛋白1(UCP1)高度相关,能被寒冷刺激和去甲肾上腺素注射诱导产生,引起UCP1表达升高。UCP1过去被认为是在褐色脂肪组织中特异性表达,直到近年来有研究发现其在人体皮肤组织中也有表达。UCP1在人和小鼠中具有高度同源性,但小鼠皮肤组织中UCP1的存在及功能尚不明确。在本研究中,我们通过对野生型和UCP1基因敲除小鼠的研究,将小鼠褐色脂肪组织作为阳性对照,测量了 UCP1在皮肤组织中mRNA及蛋白质的表达水平,发现小鼠皮肤组织中存在UCP1mRNA及蛋白质;通过分离表皮组织和真皮组织,我们进一步确定UCP1存在于皮肤组织中的表皮组织而非真皮组织中。为了进一步确定表皮组织中UCP1是否行使功能,我们试图通过寒冷刺激诱导表皮中UCP1表达的升高。结果发现寒冷刺激在表皮组织中能引起UCP1 mRNA表达水平升高,但UCP1在表皮组织中与其在褐色脂肪组织中对于短期寒冷刺激的反应存在差异。随后我们比较了寒冷刺激与去甲肾上腺素注射两种刺激方式在表皮组织与褐色脂肪组织中对UCP1表达及功能影响的异同。综上所述,线粒体解偶联蛋白UCP1存在于小鼠皮肤组织的表皮组织中,同时在寒冷刺激条件下表达水平升高,发挥解偶联功能,与其在褐色脂肪组织中的功能存在异同。本论文扩大了我们对于线粒体解偶联蛋白分布和功能的了解,或对能量代谢相关疾病的治疗及癌症病人活性氧的抑制具有潜在价值。但是后期动物水平和临床应用仍需要进一步的深入探索。更多还原

【Abstract】 Uncoupling is a reaction which occurs in the process of oxidative phosphorylation in mitochondria.It mainly can activate the substrate oxidation and consume the energy of oxidation to produce heat.Uncoupling in respiration,along with heat production during energy consumption,plays an important role in energy balance and body weight control.It also has positive significance for metabolic diseases such as obesity and can prevent the accumulation of oxygen free radicals(ROS)produced by mitochondria.Uncoupling proteins are a kind of carrier proteins located in mitochondrial inner membrane,which is responsible for uncoupling during oxidative phosphorylation.The most important role of uncoupling proteins is participating in the regulation of body temperature,maintaining the body’s thermal balance by converting energy into heat.Uncoupling proteins are also involved in the Inhibition of the formation of undesirable ROS in vivo.Non-shivering thermogenesis is a heat-producing way obtained in the process of the mammalian evolution in response to cold environment.It is highly correlated with the uncoupling protein 1(UCPI)and can be induced by cold exposure and norepinephrine injection,resulting in increased expression of UCP1.UCP1 was previously thought to be specifically expressed in brown adipose tissue.Until recent years,it has been found that UCP1 is also expressed in human skin tissue.UCP1 is highly homologous to human and mice,but the existence and function of UCP1 in mice skin tissue is unclear.In this study,based on the study on wild-type and UCP1 knock-out mice,we used the mice brown adipose tissue as a positive control,and measured the expression level of UCP1 mRNA and protein.We found that UCP1 mRNA and protein exist in mice skin tissue.By separating the epidermis and the dermis,we further confirmed that the existence of UCP1 in the mice skin tissue is in the epidermal tissue,not in the dermal tissue.In order to further determine whether UCP1 is functional in epidermal tissue,we tried to induce the increase of UCP1 expression in the epidermal tissue by cold exposure.The result suggested that cold exposure can increase the expression level of UCP1 mRNA in the epidermal tissue,but there are differences in the response of UCP1 to the short-term cold exposure between epidermal tissue and brown adipose tissue.Then,we compared the effects of cold exposure and norepinephrine injection on the expression and function of UCP1 in the epidermal tissue and brown adipose tissue.In summary,the mitochondrial uncoupling protein UCP1 exists in the epidermis of mice skin tissue.While under cold exposure conditions,UCP1 expression level elevates and plays the uncoupling function.There are similarities and differences in the function of UCP1 between epidermal tissue and brown adipose tissue.This paper extends our understanding of the distribution and functions of mitochondrial uncoupling proteins.And it may have potential value in the treatment of energy metabolism-related diseases and the inhibition of ROS in cancer patients.However,the research on animal level and clinical application still need further exploration.更多还原