Miller C E, Donlon K J, Toia L, Wong C L, Chess P R
Department of Pediatrics, University of Rochester School of Medicine and Dentistry, New York 14642, USA.
In Vitro Cell Dev Biol Anim. 2000 Nov-Dec;36(10):633-9. doi: 10.1290/1071-2690(2000)036<0633:CSIPOC>2.0.CO;2.
Embryonic heart cells undergo cyclic strain as the developing heart circulates blood to the embryo. Cyclic strain may have an important regulatory role in formation of the adult structure. This study examines the feasibility of a computerized cell-stretching device for applying strain to embryonic cardiocytes to allow measurement of the cellular response. A primary coculture of myocytes and a secondary culture of nonmyocytes from stage-31 (7 d) embryonic chick hearts were grown on collagen-coated membranes that were subsequently strained at 2 Hz to 20% maximal radial strain. After 24 h, total cell number increased by 37+/-6% in myocyte cocultures and by 26+/-6% in nonmyocyte cultures over unstrained controls. Lactate dehydrogenase and apoptosis assays showed no significant differences in cell viabilities between strained and unstrained cells. After 2 h strain, bromodeoxyuridine incorporation was 38+/-1.2% versus 19+/-0.2% (P < 0.01) in strained versus unstrained myocyte cocultures, and 35+/-2.1% versus 16+/-0.2% (P = 0.01) in nonmyocyte cultures. MF20 antibody labeling and periodic acid-Schiff (PAS) staining estimated the number of myocytes in strained wells as 50-67% larger than in control wells. Tyrosine phosphorylation may play a role in the cellular response to strain, as Western blot analysis showed an increase in tyrosine phosphorylation of two proteins with approximate molecular weights of 63 and 150 kDa within 2 min of strain. The results of this study indicate that embryonic chick cardiocytes can be cultured in an active mechanical environment without significant detachment and damage and that increased proliferation may be a primary response to strain.
随着发育中的心脏将血液输送到胚胎，胚胎心脏细胞会经历周期性应变。周期性应变可能在成体结构的形成中发挥重要的调节作用。本研究探讨了一种计算机化细胞拉伸装置对胚胎心肌细胞施加应变以测量细胞反应的可行性。从第31阶段（7日龄）胚胎鸡心脏分离的心肌细胞原代共培养物和非心肌细胞传代培养物，在涂有胶原蛋白的膜上生长，随后以2Hz的频率施加至最大径向应变20%。24小时后，与未受应变的对照相比，心肌细胞共培养物中的总细胞数增加了37±6%，非心肌细胞培养物中的总细胞数增加了26±6%。乳酸脱氢酶和凋亡检测显示，受应变和未受应变的细胞之间的细胞活力没有显著差异。应变2小时后，在受应变与未受应变的心肌细胞共培养物中，溴脱氧尿苷掺入率分别为38±1.2%和19±0.2%（P<0.01），在非心肌细胞培养物中分别为35±2.1%和16±0.2%（P = 0.01）。MF20抗体标记和过碘酸希夫（PAS）染色估计，受应变孔中的心肌细胞数量比对照孔中的大50 - 67%。酪氨酸磷酸化可能在细胞对应变的反应中起作用，因为蛋白质印迹分析显示，在应变后2分钟内，两种分子量约为63和150kDa的蛋白质的酪氨酸磷酸化增加。本研究结果表明，胚胎鸡心肌细胞可以在活跃的机械环境中培养，而不会发生明显的脱离和损伤，并且增殖增加可能是对应变的主要反应。
