Fraccarollo Daniela, Galuppo Paolo, Bauersachs Johann
Department of Cardiology and Angiology, Medical School Hannover, Carl-Neuberg-Str. 1, Hannover, 30625, Germany.
Methods Mol Biol. 2017;1627:123-137. doi: 10.1007/978-1-4939-7113-8_9.
Cardiac (myo)fibroblasts play a key role in the regulation of wound healing and pathogenic remodeling after myocardial infarction. Impaired scar formation and alterations of the extracellular matrix network precipitate cardiac dysfunction leading to increased morbidity and mortality. Therapeutic approaches addressing (myo)fibroblast phenotype appear to be useful in preventing progressive structural, electrical, and functional impairment and heart failure.Permanent ligation of the left anterior descending coronary artery has proven to be a valuable experimental model to investigate the arrays of pathways/mechanisms involved in cardiac repair and extracellular matrix remodeling in ischemic heart failure. Here we describe the surgical procedure to occlude the left coronary artery in mice. Moreover, we present an accurate method to isolate (myo)fibroblasts from ischemic myocardium, with maintenance of the functional phenotype, using the specific marker for mouse cardiac fibroblasts mEF-SK4. The protocol can be completed within a few hours, and the isolated fibroblasts/myofibroblasts are suitable for downstream molecular biology applications, like gene expression profiling and cell culture.
心脏（肌）成纤维细胞在心肌梗死后伤口愈合和病理性重塑的调节中起关键作用。瘢痕形成受损和细胞外基质网络改变会促使心脏功能障碍，导致发病率和死亡率增加。针对（肌）成纤维细胞表型的治疗方法似乎有助于预防进行性结构、电和功能损害以及心力衰竭。左冠状动脉前降支的永久性结扎已被证明是一种有价值的实验模型，可用于研究缺血性心力衰竭中参与心脏修复和细胞外基质重塑的一系列途径/机制。在此，我们描述了在小鼠中闭塞左冠状动脉的手术过程。此外，我们提出了一种准确的方法，使用小鼠心脏成纤维细胞的特异性标志物mEF-SK4，从缺血心肌中分离（肌）成纤维细胞，并维持其功能表型。该方案可在数小时内完成，分离出的成纤维细胞/肌成纤维细胞适用于下游分子生物学应用，如基因表达谱分析和细胞培养。
