瓣膜病慢性心房颤动心房肌缝隙连接CX43和CX40的重构

Remodeling of gap-junctional proteins, connexin40 and connexin43, in human atrial myocardium of valvular permanent atrial fibrillation

目的 探讨人类瓣膜病慢性心房颤动 (房颤 )心房肌中缝隙连接通道蛋白Connexin 4 3(CX4 3)和Connexin 4 0 (CX4 0 )的重构及其与房颤的关系。方法 (1)取 11例风湿性心脏瓣膜病 (风心病 )患者的右心耳心肌 (房颤 8例 ,窦性心律 3例 ) ,以Northernblot印迹法比较窦性心律和房颤者CX4 3和CX4 0mRNA表达量的变化 ;(2 )另取 2 0例风心病患者右心耳心肌 (窦性心律 8例 ,房颤 12例 ,房颤持续时间从 3个月至 15年 )制成冰冻切片 ,在激光共聚焦显微镜 (confocallaserscanningmi croscopy)下分别观察窦性心律和房颤心房肌CX4 3和CX4 0蛋白的荧光斑面积、空间分布及形态变化。结果 (1)窦性心律和房颤CX4 3和CX4 0mRNA表达量差异无显著性 ;(2 )CX4 3和CX4 0荧光斑面积在窦性心律和房颤时差异也无显著性 ,但CX4 0在心肌纵切面上的数目明显多于横切面 (P <0 0 5 ) ,而CX4 3数目无变化。提示房颤时CX4 0通道在心肌空间上的分布发生了显著改变 ,即端 端连接大于侧 侧连接。结论 人类风心病慢性房颤时缝隙连接蛋白CX4 3和CX4 0的mRNA表达量和蛋白量均未发生显著改变 ,但CX4 0蛋白的空间分布发生了显著变化 ,端 端连接明显增多 ,侧 侧连接显著减少 ,提示房颤时CX4 0通道发生了空间排列上的重构 。

Objective To reveal the changes of connexin40 (CX40) and connexin43 (CX43) mRNA level, spatial distribution pattern, and their morphology in human atrial myocardium of permanent atrial fibrillation and sinus rhythm in valvular heart disease. Methods Eleven cases of human right atrial appendage(RAA) samples were collected from rheumatic heart disease during valve replacement, eight of which were atrial fibrillation(AF) samples that average sustained AF period were (5.6±6.5) years, three of which remained sinus rhythm(SR). Total RNA was purified, CX40 and CX43 mRNA level were analysised through Northern blot technique. Twenty cases of human RAA samples from patients with rheumatic heart disease were collected(eight SR and twelve AF). Frozen sections (8 μm) were cut from longitudinal direction and transverse direction of myocardium respectively. All sections were incubated with anti-CX43 and CX40 antibody, and with appropriate secondary antibody (IgG), and then immunolabeled sections were examined using a confocal microscopy. The fluorescent plaque area and spatial distribution pattern and morphology of CX40 and CX43 proteins were evaluated between AF and SR groups. Results (1)Northern blot analysis show that the levels of CX43 and CX40 mRNA were no statistically significant differences between AF and SR groups; (2)Immunoconfocal laser scanning microscopy examination shows that CX40 gap junction in transverse orientation was reduced in large number (P<0.05), while argument markedly in longitudinal orientation in atrial fibrillation group. It is indicated that the side-to-side of CX40 gap junction was lost at large extends during atrial fibrillation. Comprising to CX43 gap junction, the spatial distribution of CX40 gap junction are more heterogeneous in atrial fibrillation group. Results also show that neither CX40 nor CX43 proteins, the average area of single gap junction fluorescent plaque were no statistical significantly difference between AF groups and SR groups. Conclusion These results suggest that the remodeling of CX40 gap junction was involved in initiation and maintenance of human valvular chronic atrial fibrillation.