Pimentel Rhea C, Yamada Kathryn A, Kléber André G, Saffitz Jeffrey E
Department of Medicine, Washington University School of Medicine, St Louis, MO, USA.
Circ Res. 2002 Apr 5;90(6):671-7. doi: 10.1161/01.res.0000014823.75393.4d.
Cardiac myocytes can rapidly adjust their expression of gap junction channel proteins in response to changes in load. Previously, we showed that after only 1 hour of linear pulsatile stretch (110% of resting cell length; 3 Hz), expression of connexin43 (Cx43) by cultured neonatal rat ventricular myocytes is increased by approximately 2-fold and impulse propagation is significantly more rapid. In the present study, we tested the hypothesis that vascular endothelial growth factor (VEGF), acting downstream of transforming growth factor-beta (TGF-beta), mediates stretch-induced upregulation of Cx43 expression by cardiac myocytes. Incubation of nonstretched cells with exogenous VEGF (100 ng/mL) or TGF-beta (10 ng/mL) for 1 hour increased Cx43 expression by approximately 1.8-fold, comparable to that observed in cells subjected to pulsatile stretch for 1 hour. Stretch-induced upregulation of Cx43 expression was blocked by either anti-VEGF antibody or anti-TGF-beta antibody. Stretch-induced enhancement of conduction was also blocked by anti-VEGF antibody. ELISA assay showed that VEGF was secreted into the culture medium during stretch. Furthermore, stretch-conditioned medium stimulated Cx43 expression in nonstretched cells. This effect was also blocked by anti-VEGF antibody. Upregulation of Cx43 expression stimulated by exogenous TGF-beta was blocked by anti-VEGF antibody, but VEGF-stimulation of Cx43 expression was not blocked by anti-TGF-beta antibody. Thus, stretch-induced upregulation of Cx43 expression is mediated, at least in part, by VEGF, which acts downstream of TGF-beta. Because the cultures contained only approximately 5% nonmyocytic cells, these results indicate that myocyte-derived VEGF, secreted in response to stretch, acts in an autocrine fashion to enhance intercellular coupling.
心肌细胞能够根据负荷变化迅速调节其缝隙连接通道蛋白的表达。此前，我们发现，培养的新生大鼠心室肌细胞在仅接受1小时的线性搏动拉伸（静息细胞长度的110%；3Hz）后，连接蛋白43（Cx43）的表达增加了约2倍，冲动传播明显加快。在本研究中，我们检验了这样一个假说：血管内皮生长因子（VEGF）在转化生长因子-β（TGF-β）的下游起作用，介导拉伸诱导的心肌细胞Cx43表达上调。将未拉伸的细胞与外源性VEGF（100 ng/mL）或TGF-β（10 ng/mL）孵育1小时，Cx43表达增加了约1.8倍，与接受1小时搏动拉伸的细胞中观察到的情况相当。拉伸诱导的Cx43表达上调被抗VEGF抗体或抗TGF-β抗体阻断。抗VEGF抗体也阻断了拉伸诱导的传导增强。ELISA分析表明，拉伸过程中VEGF分泌到培养基中。此外，拉伸条件培养基刺激了未拉伸细胞中Cx43的表达。这种作用也被抗VEGF抗体阻断。外源性TGF-β刺激引起的Cx43表达上调被抗VEGF抗体阻断，但VEGF刺激的Cx43表达未被抗TGF-β抗体阻断。因此，拉伸诱导的Cx43表达上调至少部分是由VEGF介导的，VEGF在TGF-β的下游起作用。由于培养物中仅含有约5%的非心肌细胞，这些结果表明，响应拉伸而分泌的心肌细胞源性VEGF以自分泌方式发挥作用，增强细胞间偶联。
