【摘要】： 初上高原的人常常不能很快的適應高原低氧環(huán)境,從而產(chǎn)生一系列的病理生理過程和臨床綜合征,出現(xiàn)高原反應,嚴重的發(fā)展為高原腦水腫和高原肺水腫,危及生命。其根本原因是由于外界氧分壓降低,導致低張性缺氧。線粒體是細胞的“能量工廠”,其通過消耗氧的氧化磷酸化過程為機體提供95%的能量。解偶聯(lián)蛋白家族(Uncouplingprotein,UCPs)是線粒體內(nèi)膜質(zhì)子轉(zhuǎn)運蛋白,作為質(zhì)子漏通道,使氧化磷酸化形成的質(zhì)子梯度降低,但不伴有ATP的生成,產(chǎn)生“無效氧耗”。近年來對UCPs的研究多涉及營養(yǎng)、代謝等領域,但在高原缺氧時UCPs在機體對于高原習服和適應中的作用未見報道。UCP4和BMCP-1是近年來才發(fā)現(xiàn)的特異在腦組織中豐富表達的UCPs家族成員,而缺氧時UCP4和BMCP-1在腦組織中的表達、異構(gòu)體分布、功能特異性以及線粒體氧耗和ATP生成的關系尚無人問津。因此研究缺氧時UCPs活性和含量的變化及其對線粒體能量代謝和呼吸氧耗的影響,對提高氧的利用和能量的生成有重要的意義。 方法 健康雄性SD大鼠暴露于模擬海拔5000米高原低壓艙內(nèi),23小時/天,分別連續(xù)缺氧3天(急性缺氧組)和30天(慢性缺氧組),同時設立平原對照組;各缺氧組與對照組分別在模擬高原低壓艙內(nèi)和平原斷頭處死,分離大鼠腦線粒體,Clark氧電極法測定線粒體氧化呼吸活性,寡霉素抑制法測定F_0-F_1ATP酶活性,羅丹明123法測定線粒體膜電位,HPLC分析腦組織線粒體內(nèi)腺苷酸含量,[~3H]-GTP結(jié)合法同時測定腦組織UCPs的活性與含量,RT-PCR檢測UCP4、BMCP-1mRNA表達量,Western blot測定UCP4、BMCP-1蛋白表達。 結(jié)果 1.急性缺氧暴露可顯著降低大鼠腦線粒體ST3、RCR、OPR,而ST4則顯著增高;慢性缺氧腦線粒體ST3、OPR較對照顯著下降,而與急性組比較,ST4顯著降低,RCR則顯著升高。 2.急、慢性缺氧組大鼠腦線粒體ATP含量均顯著下降,分別為對照的58.4%和71.4%。而慢性組則有所回升,顯著高于急性組;急、慢性組AMP含量均較對照組顯著增高;急性組ADP含量顯著增高;急、慢性組ATP/ADP均顯著降低,其中慢性組顯著高于急性組。急、慢性缺氧組能荷均顯著下降,相較于對照組分別降低20.1%、22.3%。 3.急、慢性組腦線粒體UCPs與[~3H]-GTP結(jié)合的解離常數(shù)Kd均顯著下降,各為對照的58.1%、75.8%;而慢性組則高于急性組。急性組線粒體UCPs結(jié)合[~3H]-GTP的最大量Bmax顯著增高,為對照組的2.9倍;慢性組則顯著低于急性組,而與對照組無統(tǒng)計學意義差別。 急性組UCP4和BMCP-1 mRNA和蛋白的表達均顯著高于對照組及慢性組,其中UCP4、BMCP-1 mRNA表達分別為對照組的2.34倍和4.08倍。慢性組BMCP-1 mRNA和蛋白表達較對照組顯著增高,而UCP4 mRNA和蛋白表達則恢復至對照水平。 4.急、慢性缺氧組腦組織線粒體膜電位(MMP)和F_0-F_1ATP酶活性顯著下降,其中急性組分別降為對照組的80.5%和55.4%,而慢性組則分別降為對照組的87.4%和77.5%,比急性組均有回升。 結(jié)論 1、缺氧可顯著增加大鼠腦線粒體UCPs活性。急性缺氧暴露可誘導大鼠腦線粒體UCP4、BMCP-1 mRNA和蛋白的表達,增加腦組織中UCPs的含量,慢性缺氧時則有所回降,而急、慢性缺氧均可增加腦線粒體UCPs活性,表明UCPs(UCP4、BMCP-1)表達的增加是活性增高的主要因素。 2、缺氧時UCPs活性增強,驅(qū)散線粒體內(nèi)膜質(zhì)子梯度,同時增強線粒體ST4呼吸,使無效氧耗增強,從而使ATP合成減少,能量生成效率降低。 3、慢性缺氧暴露可在一定程度上使大鼠腦組織線粒體UCPs活性回降,增強氧化磷酸化功能,部分恢復ATP合成酶活性和線粒體內(nèi)ATP含量。提示經(jīng)長期缺氧習服-適應后腦組織UCPs活性的下降,使無效氧耗減少,能量生成效率提高,從而保持能量平衡,這無疑是慢性高原缺氧習服-適應的重要機制之一。
[Abstract]:The people of the first upper plateau often do not adapt to the high altitude hypoxia environment very quickly, so as to produce a series of pathophysiological processes and clinical syndromes, and the high altitude reaction occurs, and the serious development is high altitude cerebral edema and high altitude pulmonary edema, which is life-threatening. The root cause is due to the lower external oxygen partial pressure, resulting in low-tension oxygen deficiency. The mitochondria are the "energy plant" of the cells that provide 95% energy to the body by the oxidative phosphorylation of oxygen. Uncodingprotein (UCPs) is the mitochondrial membrane proton transport protein, which is used as the proton leakage channel, so that the proton gradient formed by the oxidative phosphorylation is reduced, but without the generation of ATP, the "dead oxygen consumption" is generated. In recent years, the research of UCPs in the field of nutrition, metabolism and so on, but the role of UCPs in the acclimatization and adaptation of the high altitude is not reported. UCP4 and BMCP-1 are members of the UCPs which are found only in the brain tissue in recent years, and the expression of UCP4 and BMCP-1 in the brain tissue, the distribution of the isomers, the function specificity and the relationship between the mitochondrial oxygen consumption and the ATP production are not yet known. Therefore, it is of great significance to study the changes of the activity and content of UCPs and its effect on the energy metabolism and the oxygen consumption of the mitochondria in the absence of hypoxia, and it is of great significance to improve the utilization of oxygen and the generation of energy. Methods Healthy male SD rats were exposed to the low-pressure cabin of the simulated altitude of 5000 m,23 hours/ day, respectively for 3 days (acute hypoxia group) and 30 days (chronic hypoxia group). in that control group of the plain, the hypoxia group and the control group were sacrificed at the low-pressure cabin and the plain end of the simulated plateau, respectively, the mitochondria of the brain of the rat were isolated, the activity of the mitochondrial oxidative respiration was determined by the Clark oxygen electrode method, the activity of the F _ 0-F _ 1 ATPase was determined by the oligomycin inhibition method, and the determination of the line by the rhodamine 123 method The activity and contents of UCP4 and BMCP-1 were determined by RT-PCR, and the expression of UCP4 and BMCP-1 was determined by Western blot. 1 egg Results:1. Acute hypoxia exposure could significantly lower the mitochondrial ST3, RCR and OPR of the rat brain, while the ST4 increased significantly; the chronic hypoxic-brain mitochondria ST3 and OPR decreased significantly compared with the acute group, and ST4 was significantly higher than that in the acute group. 2. The content of ATP in the brain of the rats in the acute and chronic hypoxia group decreased significantly, respectively. The control group was 58.4% and 71.4%, while the chronic group recovered, significantly higher than that of the acute group, and the content of AMP in the acute and chronic group was significantly higher than that in the control group; the content of ADP in the acute group was significantly increased; and the ATP/ ADP in the acute and chronic groups decreased significantly. The chronic group was significantly higher in the acute and chronic hypoxia group than in the control group. The dissociation constant (Kd) of the mitochondrial UCPs and[~ 3H]-GTP in the acute and chronic group decreased significantly, each of which was 58.1% of the control group. The most significant Bmax of the acute group of UCPs binding to[~ 3H]-GTP was significantly higher in the acute group than in the control group, and the chronic group was significantly lower than that in the control group. The expression of UCP4 and BMCP-1 mRNA and protein in the acute group was significantly higher than that in the control group and the chronic group, of which the UCP4 and BMCP-1 mRNA were significantly higher in the acute group than in the control group and the chronic group. The expression of BMCP-1 mRNA and protein in the chronic group was significantly higher than that of the control group, while the expression of BMCP-1 mRNA and protein in the chronic group was significantly higher than that of the control group. In the acute and chronic hypoxia group, the mitochondrial membrane potential (MMP) and F _ 0-F _ 1 ATPase activity of the brain tissue of the acute and chronic hypoxia group decreased significantly, and the acute group decreased to 80.5% and 55.4% in the control group, respectively, while the chronic group decreased to the control group, respectively. irradiation group 87.4% and 77.5% in the acute group. Conclusion 1. Hypoxia can significantly increase the activity of UCPs in the brain of rats. Acute hypoxia exposure can induce the expression of UCP4, BMCP-1 mRNA and protein in the brain of rats, increase the content of UCPs in the brain, and decrease the activity of UCPs in chronic hypoxia. The increase of the expression of s (UCP4, BMCP-1) is the main factor of the increase of activity. T4 respiration, which increases the effective oxygen consumption, decreases the synthesis of ATP and decreases the energy generation efficiency.3. Chronic hypoxia exposure can increase the activity of the mitochondrial UCPs in the brain of rats to a certain extent. In order to decrease the activity of UCPs in the brain tissue after long-term hypoxic acclimatization, the activity of UCPs in brain tissue can be reduced, and the effective oxygen consumption is reduced and the energy is effective.
【學位授予單位】：第三軍醫(yī)大學
【學位級別】：碩士
【學位授予年份】：2006
【分類號】：R363

【引證文獻】

相關博士學位論文 前1條

1 徐丁潔;婦科實寒癥與虛寒癥代謝組學及證候形成過程中相關網(wǎng)絡調(diào)控的比較研究[D];河北醫(yī)科大學;2012年

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