Tan Kezhe, Ding Zhaoping, Steckel Bodo, Hartwig Sonja, Lehr Stefan, Deng Xiaoming, Schrader Jürgen
Department of Anesthesiology, Changhai Hospital, Second Military Medical University; Department of Molecular Cardiology, Heinrich Heine University.
Department of Molecular Cardiology, Heinrich Heine University.
J Vis Exp. 2017 Jun 20(124):55849. doi: 10.3791/55849.
The present protocol describes a unique approach that enables the collection of cardiac transudate (CT) from the isolated, saline-perfused rat heart. After isolation and retrograde perfusion of the heart according to the Langendorff technique, the heart is inverted into an upside-down position and is mechanically stabilized by a balloon catheter inserted into the left ventricle. Then, a thin latex cap - previously cast to match the average size of the rat heart - is placed over the epicardial surface. The outlet of the latex cap is connected to silicon tubing, with the distal opening 10 cm below the base level of the heart, creating slight suction. CT continuously produced on the epicardial surface is collected in ice-cooled vials for further analysis. The rate of CT formation ranged from 17 to 147 µL/min (n = 14) in control and infarcted hearts, which represents 0.1-1% of the coronary venous effluent perfusate. Proteomic analysis and high performance liquid chromatography (HPLC) revealed that the collected CT contains a wide spectrum of proteins and purinergic metabolites.
本方案描述了一种独特的方法，可从离体的、用生理盐水灌注的大鼠心脏收集心脏渗出液（CT）。根据Langendorff技术对心脏进行分离和逆行灌注后，将心脏倒置，通过插入左心室的球囊导管进行机械固定。然后，将一个预先铸造以匹配大鼠心脏平均大小的薄乳胶帽放置在心外膜表面。乳胶帽的出口连接到硅胶管，远端开口位于心脏基线以下10 cm处，产生轻微吸力。在心外膜表面持续产生的CT收集在冰冷却的小瓶中用于进一步分析。在对照心脏和梗死心脏中，CT形成速率为17至147 μL/分钟（n = 14），占冠状静脉流出灌注液的0.1-1%。蛋白质组学分析和高效液相色谱（HPLC）显示，收集到的CT含有多种蛋白质和嘌呤能代谢产物。
