Characterization of novel resident heart cells identified as atypically-shaped cardiomyocytes(ACMs)

我们研究了成年小鼠体内存在有别于心肌细胞的常驻心肌细胞。心脏冠状灌注酶，切除两个心室并进一步消化。将心肌细胞旋转后，取上清液(心肌细胞耗竭部分，CMDF)在甲基纤维素半固体培养液中培养。培养三到五天后，一些附着在培养皿底部的小细胞逐渐形成形状并开始自发跳动。这些细胞大多是多核的，肌节组织良好，表达心脏特异蛋白，被定义为一种新型的心脏细胞，非典型形状的心肌细胞(ACM)。ACM细胞数量在新生期最高，5周内显著下降，成人期达到平台期。从新生和老年小鼠获得的ACM都表达胎儿心脏基因产物，如心钠素(ANP)和电压门控钙通道CaV3.2。我们研究了CMDF细胞，包括ACM，在经历模拟的致命性缺血后是否会发育成搏动的细胞。缺血预暴露的CMDF细胞培养6d后，其细胞数约为常氧条件下的50%。对ACM的电子显微镜分析表明，即使在常氧条件下，培养过程中也有结构性活跃的自噬。这一结果表明，即使存活的细胞数量很少，心脏损伤也可能发生跳动中的ACM。

We have investigated the presence of resident heart cells that are distinct from cardiomyocytes in adult mice. The heart was coronary perfused with enzymes and both ventricles were excised and further digested. After spinning the ventricular myocytes down, the supernatant fraction (cardiac myocyte-depleted fraction, CMDF) was cultured in methylcellulose-based semisolid culture medium. Three to five days after plating, some of the small cells adhered to the bottom of the dishes gradually developed their shapes and started beating spontaneously. These cells were mostly multinucleated, well sarcomeric-organized and expressed cardiac specific proteins, defined as a novel type of heart cells, atypically-shaped cardiomyocytes (ACMs). The cell population of ACMs was highest at neonatal period and significantly decreased within the first 5 weeks and reached a plateau in the adult stage. The ACMs obtained from both newborn and aged mice express the fetal cardiac gene products, such as atrial natriuretic peptide (ANP) and voltage-gated Ca2+channel CaV3.2. We examined whether CMDF cells, including ACMs, that underwent simulated lethal ischemia could develop into beating cells. When CMDF cells pre-exposed to ischemia were cultured for 6 days, the cell number of beating ACMs was ~50% of normoxic preparations. Electron microscopic analyses of ACMs displayed constitutively active autophagy during the culture even in the normoxic conditions. The results suggest the possibility that the development of beating ACMs could occur in injured heart, even if the surviving cell population is small.