Isolation and Characterization of Intact Cardiomyocytes from Frozen and Fresh Human Myocardium and Mouse Hearts.
Isolation and Characterization of Intact Cardiomyocytes from Frozen and Fresh Human Myocardium and Mouse Hearts.
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DOI：
10.1007/978-1-0716-0668-1_15
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Kühn B
中科院分区：
文献类型：
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作者：
Liu H;Bersell K;Kühn B
完整人体细胞的获取和表征对于再生医学和转化医学研究至关重要。目前分离完整细胞的方法的选择取决于心脏的年龄。为了从胎儿或新生儿心肌中分离心肌细胞，可以将心肌切碎成小组织块，然后进行酶孵育。而胎儿和新生儿心肌细胞非常柔软，分离后形态由长杆状或梭形变为球形。由于成年心肌细胞的致密性和细胞间的紧密连接，仅用酶孵育法很难从成年心肌中分离心肌细胞。灌注方法是将酶溶液输送到心肌深层所必需的。然而，灌注方法需要非常罕见的完整心脏。因此，缺乏从不同年龄的心肌中有效分离心肌细胞的方法，在基础研究和临床研究之间建立了障碍。在这里，我们描述了一种从新鲜或冷冻的人心肌或新鲜小鼠心脏中分离完整心肌细胞的方法，并对多核化、心肌细胞大小、细胞周期活性和每颗心脏的心肌细胞总数进行定量。我们通过从各种小鼠器官（包括肝脏、肺和胸腺）中分离细胞来推广这种固定消化方法。
Procurement and characterization of intact human cells is essential for studies in regenerative medicine and translational medical research. The selection of the currently available approaches to isolate intact cells depends on the age of the hearts. To isolate cardiomyocytes from the fetal or neonatal myocardium, the myocardium can be minced into small tissue blocks followed by enzyme incubation. However, the fetal and neonatal cardiomyocytes are very soft and the morphology changes from long-rod or spindle shape to spheres after isolation. Because the dense packing of the cardiomyocytes and the strong cell-cell connection in adult myocardium, it is difficult to isolate the cardiomyocytes from adult myocardium only by enzyme incubation. Perfusion method is necessary to deliver the enzyme solution to the deep layers of the myocardium. However, intact hearts, which is very rare, are required for perfusion method. Therefore, lacking methods to efficiently isolate cardiomyocytes from myocardium of various ages builds a barrier between basic research and clinical studies. Here, we describe a method for the isolation of intact cardiomyocytes from fresh or frozen human myocardium or fresh mouse hearts and the quantification of multinucleation, cardiomyocyte size, cell cycle activity, and total cardiomyocyte count per heart. We generalize this fixation-digestion method by isolating cells from a variety of mouse organs, including the liver, lung, and thymus.