Mitochondria/Mass Spectroscopy Core

心肌线粒体存在于两个不同的群体中，即肌细胞内的肌膜下线粒体（SSM）， 位于肌膜下，和间纤维线粒体（IFM）之间的居住， 肌原纤维SSM通过组织均质化释放，而IFM的释放需要短暂的暴露 用胰蛋白酶处理。这两种线粒体是不同的 在生理和病理生理条件下都受到影响，因此需要研究每种 个别人群分开。线粒体/质谱核心（核心D）将执行以下操作 作为不同项目的服务。 1.从实验心脏分离肌膜下和原纤维间线粒体;评估质量控制。 2.获取心肌样品和分离的SSM和IFM群体，用于电子传递链复合物 分析. 3.获取心脏、SSM和IFM样本，用于脂质提取和磷脂分析。特别注意 心磷脂（CL）分析：CL含量，CL种类和CL氧化。 4.获取心脏组织用于酰基辅酶A和酰基肉毒碱的分析，特别强调丙二酰辅酶A是关键 代谢物在燃料利用中的转换。 5.进行质谱分析，以鉴定ETC上的翻译后和氧化修饰 亚基，尤其是复合体IV。 6.通过透射电镜对心脏组织和心脏SSM和IFM进行结构分析。 7.异前列烷和氧化低密度脂蛋白（oxLDL）定量作为体内氧化性脂蛋白的测量 应力 8.将对项目1和4的线粒体进行蓝色天然电泳。 相关性（参见说明）： 线粒体是发电站和细胞。在心力衰竭期间，心脏发生能量危机 与线粒体功能障碍有关。这个核心将提供线粒体分离的心脏， 狗和小鼠在实验条件下，并执行分析方法，以支持4个项目内 计划项目补助金。

Cardiac mitochondria exist in two distinct populations wathin the myocyte, subsarcolemmal mitochondria (SSM), located beneath the sarcolemmal membrane, and interfibrillar mitochondria (IFM) that reside between the myofibrils. SSM are released by tissue homogenization, whereas the liberation of IFM requires a brief exposure of the resulting skinned myocytes to trypsin treatment. The two populations of mitochondria are differently affected under both physiological and pathophysiological conditions and thus requires to study of each individual population separately. The Mitochondrial/Mass Spec Core (Core D) will perform the following experiments as a service to the different Projects. 1. Isolate subsarcolemmal and interfibrillar mitochondria from experimental hearts; assess quality control. 2. Obtain samples of myocardium and isolated populations of SSM and IFM for electron transport chain complex analysis. 3. Obtain samples of heart, SSM, and IFM for lipid extraction and phospholipid analysis. Special attention is paid to cardiolipin (CL) analysis: CL content, CL species and CL oxidation. 4. Aquire heart tissue for analysis of acyl-CoAs and acylcarnitines with special emphasis on malonyl-CoA a key metabolite in switch in fuel utilization. 5. Carry out mass spectrometric analysis to identify posttranslational and oxidative modifications on ETC subunits, especially complex IV. 6. Perform structural analysis on cardiac tissue and cardiac SSM and IFM by transmission electron microscopy. 7. Isoprostane and oxidized low density lipoprotein (oxLDL) quantitation as a measure of in vivo oxidative stress. 8. Blue Native Electrophoresis will be done on mitochondria from Project 1 and 4. RELEVANCE (See instructions): Mitochondrial are the powerhouses and the cell. During heart failure an energy crises occurs in the heart and is related to the dysfunction of mitochondria. This Core will provide mitochondrial isolation from heart of dogs and mice under experimental conditions and perform analytical methods to support the 4 project within the Program Project Grant.