Pedrotty Dawn M, Klinger Rebecca Y, Badie Nima, Hinds Sara, Kardashian Ara, Bursac Nenad
Dept. of Biomedical Engineering, Duke Univ., Durham, NC 27708, USA.
Am J Physiol Heart Circ Physiol. 2008 Jul;295(1):H390-400. doi: 10.1152/ajpheart.91531.2007. Epub 2008 May 23.
Well-controlled studies of the structural and functional interactions between cardiomyocytes and other cells are essential for understanding heart pathophysiology and for the further development of safe and efficient cell therapies. We established a novel in vitro assay composed of a large number of individual micropatterned cell pairs with reproducible shape, size, and region of cell-cell contact. This assay was applied to quantify and compare the frequency of expression and distribution of electrical (connexin43) and mechanical (N-cadherin) coupling proteins in 5,000 cell pairs made of cardiomyocytes (CMs), cardiac fibroblasts (CFs), skeletal myoblasts (SKMs), and mesenchymal stem cells (MSCs). We found that for all cell pair types, side-side contacts between two cells formed 4.5-14.3 times more often than end-end contacts. Both connexin43 and N-cadherin were expressed in all homotypic CM pairs but in only 13.4-91.6% of pairs containing noncardiomyocytes, where expression was either junctional (at the site of cell-cell contact) or diffuse (inside the cytoplasm). CM expression was exclusively junctional in homotypic pairs but predominantly diffuse in heterotypic pairs. Noncardiomyocyte homotypic pairs exhibited diffuse expression 1.7-8.7 times more often than junctional expression, which was increased 2.6-4.4 times in heterotypic pairs. Junctional connexin43 and N-cadherin expression, respectively, were found in 38.6 +/- 7.3 and 39.6 +/- 6.2% of CM-MSC pairs, 21.9 +/- 5.0 and 13.6 +/- 1.9% of CM-SKM pairs, and in only 3.8-9.6% of CM-CF pairs. Measured frequencies of protein expression and distribution were stable for at least 4 days. Described studies in micropatterned cell pairs shed new light on cellular interactions relevant for cardiac function and cell therapies.
对心肌细胞与其他细胞之间结构和功能相互作用进行严格对照研究，对于理解心脏病理生理学以及安全有效的细胞治疗的进一步发展至关重要。我们建立了一种新型体外分析方法，该方法由大量具有可重复形状、大小和细胞间接触区域的单个微图案化细胞对组成。此分析方法用于量化和比较由心肌细胞（CMs）、心脏成纤维细胞（CFs）、骨骼肌成肌细胞（SKMs）和间充质干细胞（MSCs）组成的5000个细胞对中电耦合蛋白（连接蛋白43）和机械耦合蛋白（N-钙黏着蛋白）的表达频率和分布情况。我们发现，对于所有细胞对类型，两个细胞之间的侧面接触形成频率比端面接触高4.5至14.3倍。连接蛋白43和N-钙黏着蛋白在所有同型CM对中均有表达，但在仅含非心肌细胞的细胞对中，表达率仅为13.4%至91.6%，其表达要么是连接性的（在细胞间接触部位），要么是弥散性的（在细胞质内）。CM在同型对中的表达仅为连接性的，但在异型对中主要是弥散性的。非心肌细胞同型对中弥散性表达比连接性表达多1.7至8.7倍，在异型对中增加了2.6至4.4倍。在CM-MSC对中，连接性连接蛋白43和N-钙黏着蛋白的表达分别为38.6±7.3%和39.6±6.2%，在CM-SKM对中分别为21.9±5.0%和13.6±1.9%，而在CM-CF对中仅为3.8%至9.6%。所测量的蛋白表达频率和分布至少在4天内保持稳定。在微图案化细胞对中所描述的研究为与心脏功能和细胞治疗相关的细胞相互作用提供了新的见解。
