线粒体氧化应激与心肌重构研究进展

贾春澍,范志民,陈晓亮,任立群*

(1.吉林大学药学院实验药理与毒理学教研室,吉林长春130021;2.吉林大学白求恩第一医院乳腺外科;3.吉林大学中日联谊医院泌尿外科)

心力衰竭是工业化国家致死的首要原因[1]。这也是一个日益严重的公共卫生问题,主要是由于人口老龄化和老人心力衰竭患病率升高。大量的基础、临床和人口科学研究促进了心力衰竭的现代治疗,但左心室(left ventricular,LV)衰竭发生和发展最根本的机制仍未完全阐明。活性氧簇(Reactive oxygen species,R OS)如超氧负离子(·O2-)和羟自由基(-OH)可导致膜磷脂、蛋白和DNA氧化[2],并与一系列病理状态有关,如缺血再灌注损伤[3],神经变形性疾病[4]及衰老[5]。在生理状态下,活性氧簇的毒性效应可以被超氧化物歧化酶(superoxide dismutase,SOD)、谷胱甘肽过氧化物酶(glutathione peroxidase,GSHPx)、过氧化氢酶及其他非酶抗氧化剂清除。但是,当ROS的产生超过抗氧化剂的防御能力时,氧化应激即对生物组织产生功能和结构上的破坏效应。ROS随即导致心肌收缩力下降及结构损伤。氧化应激在心肌衰竭发展过程中的LV重构的病理生理机制中所起的作用日益受到重视。现有的证据表明,ROS的突出作用是作为一个信号分子对激素、生长和凝血因子、细胞因子和氧分压改变的作用做出反应[6]。在低氧状态下,高水平的线粒体ROS能够诱导低氧诱导因子(hypoxiainducible factor,HIF-1α)的激活。HIFs是细胞对低氧适应性反应的主要因子。线粒体ROS与低氧状态下HIF-1α的表达密切相关,多种蛋白激酶参与ROS与HIF-1α表达之间的信号转导[7],其具体机制仍待阐明。另外,也有研究认为抗氧化剂加重心力衰竭[8]。

1 心力衰竭过程中的线粒体氧化应激

目前的实验及临床证明ROS的产生促进心力衰竭[9-12]。脂质过氧化物水平和 8-iso-前列腺素 F2α(8-isoprostaglandin F2a,8-iso-PGF2a)是ROS产生的主要生物学标志物。心力衰竭患者的血清和心包液中这两项指标明显升高并与其严重程度相关[9,12]。Belch等[9]报道丙二醛水平与LV射血分数呈现明显负相关(r=-0.35)。Mallat等[12]应用NYHA分类和超声心动图评价心肌衰竭的严重度,证明其与心包液中8-iso-PGF2a含量相关:有症状的心力衰竭患者(NYHA II和III)心包液中8-iso-PGF2a含量明显高于无症状(NYHA I)心力衰竭患者,并与病情严重程度相关(P=0.000 3)。另外,心包液中8-iso-PGF2a与LV舒张末期及收缩末期直径呈正相关(P=0.008,P=0.026,respectively)[12]。心脏中R OS的细胞来源包括心肌细胞,上皮细胞和中性粒细胞。心肌细胞ROS产生的亚细胞定位包括线粒体电子传递,NAD(P)H氧化酶,和黄嘌呤脱氢酶/黄嘌呤氧化酶。心脏是摄氧率最高的器官,基础代谢状态下每分钟每克体重消耗约0.1 ml O2[13]。为满足氧化代谢ATP合成的需要,整个机体中心肌细胞具最高的线粒体体积密度。线粒体通过单电子经呼吸链传递至氧分子产生ROS。在生理状态下,线粒体呼吸链传递过程产生微量ROS,肌细胞的内源性清除机制随即发挥清除作用。

应用电子自旋共振(electron spin resonance,ESR)光谱学,5,5’二甲基-1-吡咯啉-N-氧化物(DMPO)作为捕捉物,在正常的亚线粒体结构抑制电子传递链的复合体I和复合体 Ш导致大量·O2-的产生[14]。在NADH作用下,衰竭心脏的线粒体较正常心脏产生更多的·O2-,说明线粒体电子传递链是·O2-的主要来源。进一步来说,线粒体功能衰竭伴随着复合酶活力的降低。因此,线粒体是衰竭心脏ROS的主要来源,同时也证明线粒体衰竭和氧化应激[15]的病理生理关联。

2 线粒体DNA损伤,功能衰竭和氧化应激

线粒体具其单独的基因组,即mtDNA,一个闭合环状双链DNA分子,约16.5 KB。mtDNA有两个启动子,轻链(LSP)和重链启动子(HSP)。线粒体功能受mtDNA的调控,同时mtDNA转录和/或复制因子也对其产生影响[16]。这就提出了mtDNA损伤和线粒体基因转录或复制的异常与心力衰竭相关的可能。实际上,很多证据表明心力衰竭与mtDNA在质和量上的缺陷相关[17-20]。线粒体功能和mtDNA拷贝数的下降在心肌缺血后心力衰竭的发展中发挥着重要作用[17,21]。

ROS能够在其形成或靠近形成部位损伤线粒体大分子。因此,除了产生 R OS,线粒体本身也能被ROS损伤,mtDNA是主要的靶点。原因如下,首先,线粒体基因组无组蛋白参与组装,即缺少了对抗ROS损伤的一道屏障。第二,mtDNA的DNA修复能力有限。第三,线粒体内形成大量的·O2-,并且不能通过线粒体膜,因此,ROS损伤大部分局限在线粒体内。实际上,mtDNA聚集了大量DNA氧化产物,8-羟基,明显高于核DNA[22]。与核编码基因不同,线粒体编码基因表达调控主要依赖于mtDNA的拷贝数[23]。因此,线粒体损伤主要表现在mtDNA的损伤,即线粒体RNA(mtRNA)转录子,蛋白合成和线粒体功能的下降[24,25]。衰竭心脏中线粒体结构损伤和功能衰竭与ROS升高水平有关,主要表现为线粒体脂质过氧化物水平升高,mtDNA拷贝数下降,mtRNA转录子数目下降,及低复合酶活力导致的氧化能力下降[21]。更重要的是,衰竭心脏的Ⅰ,Ⅲ,和Ⅳ复合酶活力下降,而单纯由核DNA编码的复合酶Ⅱ和柠檬酸合成酶活力无下降表现。慢性的ROS产生增高与线粒体损伤和功能衰竭有关,即形成线粒体功能下降的恶性循环,大量的ROS产生引起细胞损伤。mtDNA损伤通过上述机制参与心肌重构和心力衰竭的发生和发展。

3 线粒体氧化应激在心肌重构中的作用

氧化应激对心肌细胞结构和功能有直接的作用,能够直接激活心肌重构和心力衰竭的信号分子。ROS导致心肌细胞的表型变化,即离体的心肌细胞肥大和凋亡[26]。

ROS另一个作用靶点是金属基质蛋白酶(matrix metalloproteinases,MMPs)。MMPs在正常组织重构中发挥重要的作用,如细胞迁移,侵袭,增殖和凋亡。并参与多种发育过程,如血管分枝形态建成,血管发生,创伤愈合及细胞外基质降解。MMPs表达于大量细胞和组织中并广泛的降解细胞外基质蛋白[27]。ROS有激活心肌成纤维细胞MMP的作用[28]。衰竭心肌细胞MMP活力增高[26,29]。MMP抑制剂具有限制心肌缺血大鼠模型早期LV扩张的作用[30]。心肌缺血模型中,MMP-2基因敲除明显抑制了早期心脏破裂和LV衰竭的发展,生存率明显提高[31]。由于 ROS能激活MMP[32],随即提出了ROS的过多产生过度激活MMPs导致LV重构的学说。持续的MMP激活可能通过提供一个异常的细胞外环境影响心肌的结构特点。·OH清除剂二甲叉三脲能够抑制心肌重构和心力衰竭相关的MMP-2激活[33]。这些证据均表明心肌缺血后的过度氧化应激是心肌MMP激活的刺激物,并在心力衰竭发展过程中发挥重要作用。

4 氧化应激与骨骼肌功能失调的作用

运动能力受限是心衰患者的主要症状[34],而不依赖于心力衰竭程度[35]。心衰患者运动能力受限与过度的氧化应激有关[36]。心肌缺血导致的心肌衰竭大鼠骨骼肌ROS含量明显升高,主要是线粒体产生的·O2-[37]。目前,Kinugawa等[38]阐述了·O2-与运动能力受限的关系,·O2-含量升高运动能力下降,同时与全身氧耗升高有关。解偶联蛋白(Uncoupling proteins,UCPs)是线粒体内膜质子运载体,能够降低质子线粒体内膜电化学梯度。电化学梯度的降低导致ATP生物合成的降低。衰竭心肌线粒体表达UCPs明显上调[39]。另外,Echtay等[40]的研究表明线粒体内ROS激活UCPs。这些结果表明ROS能通过上调UCPs表达引起能效改变,这可能在心力衰竭发展过程中骨骼肌的功能失调发挥作用。

5 结语

为改善心力衰竭患者的预后,我们需要基于心肌重构和心力衰竭病理生理过程的深入探寻发展一种新型的治疗方法。线粒体氧化应激和mtDNA损伤的调节方法的研究可能有助于建立有效治疗心力衰竭策略。氧化应激不仅涉及心力衰竭,而且与各种心血管疾病包括动脉粥样硬化,高血压及衰老过程相关。因此,以调节这种不当的适应性的反应作为治疗策略可能得到广泛的应用。

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