Haddad J, Decker M L, Hsieh L C, Lesch M, Samarel A M, Decker R S
Department of Medicine, Northwestern University Medical School, Chicago, Illinois 60611.
Am J Physiol. 1988 Jul;255(1 Pt 1):C19-27. doi: 10.1152/ajpcell.1988.255.1.C19.
The present observations demonstrate that quiescent calcium-tolerant adult rabbit cardiac myocytes can be isolated by collagenase-hyaluronidase perfusion and maintained in primary culture for at least 2 wk. Culturing large numbers of myocytes requires that the freshly isolated cells be attached to a suitable substratum such as laminin, type IV collagen, or fetal bovine serum. The cultured myocytes retain their rod-like morphology for approximately 7 days before gradually spreading into a flattened conformation by 14 days. During the 1st wk of culture, contaminating interstitial cells rapidly proliferate, making cultures unsuitable for long-term study. Pure myocyte populations can be established and maintained if freshly isolated cells are cultured in the presence of cytosine arabinoside (Ara-C, 10 microM). This antimetabolite does not appear to adversely affect high-energy phosphates, since ATP and creatine phosphate (CrP) content of the myocytes is maintained at levels normally found in biopsy samples of rabbit myocardium. These results illustrate that an energetically stable population of adult cardiac myocytes can be maintained in primary culture in sufficient numbers to make them useful for future investigations of myocyte function.
目前的观察结果表明，通过胶原酶-透明质酸酶灌注可以分离出静止的耐钙成年兔心肌细胞，并在原代培养中维持至少2周。培养大量心肌细胞需要将新鲜分离的细胞附着在合适的基质上，如层粘连蛋白、IV型胶原或胎牛血清。培养的心肌细胞在大约7天内保持其杆状形态，到14天时逐渐伸展成扁平形态。在培养的第1周，污染的间质细胞迅速增殖，使培养物不适用于长期研究。如果在胞嘧啶阿拉伯糖苷（Ara-C，10 microM）存在的情况下培养新鲜分离的细胞，可以建立并维持纯心肌细胞群体。这种抗代谢物似乎不会对高能磷酸盐产生不利影响，因为心肌细胞中的ATP和磷酸肌酸（CrP）含量维持在兔心肌活检样本中通常发现的水平。这些结果表明，在原代培养中可以维持足够数量的能量稳定的成年心肌细胞群体，使其可用于未来心肌细胞功能的研究。
