Gorman Jennifer L, Liu Sammy T K, Slopack Dara, Shariati Khashayar, Hasanee Adam, Olenich Sara, Olfert I Mark, Haas Tara L
School of Kinesiology and Health Science, Angiogenesis Research Group and Muscle Health Research Centre, York University, Toronto, Ontario, Canada.
West Virginia University School of Medicine, Center for Cardiovascular and Respiratory Sciences, Division of Exercise Physiology, Morgantown, West Virginia, United States of America.
PLoS One. 2014 Jan 9;9(1):e85537. doi: 10.1371/journal.pone.0085537. eCollection 2014.
Skeletal muscle overload induces the expression of angiogenic factors such as vascular endothelial growth factor (VEGF) and matrix metalloproteinase (MMP)-2, leading to new capillary growth. We found that the overload-induced increase in angiogenesis, as well as increases in VEGF, MMP-2 and MT1-MMP transcripts were abrogated in muscle VEGF KO mice, highlighting the critical role of myocyte-derived VEGF in controlling this process. The upstream mediators that contribute to overload-induced expression of VEGF have yet to be ascertained. We found that muscle overload increased angiotensinogen expression, a precursor of angiotensin (Ang) II, and that Ang II signaling played an important role in basal VEGF production in C2C12 cells. Furthermore, matrix-bound VEGF released from myoblasts induced the activation of endothelial cells, as evidenced by elevated endothelial cell phospho-p38 levels. We also found that exogenous Ang II elevates VEGF expression, as well as MMP-2 transcript levels in C2C12 myotubes. Interestingly, these responses also were observed in skeletal muscle endothelial cells in response to Ang II treatment, indicating that these cells also can respond directly to the stimulus. The involvement of Ang II in muscle overload-induced angiogenesis was assessed. We found that blockade of AT1R-dependent Ang II signaling using losartan did not attenuate capillary growth. Surprisingly, increased levels of VEGF protein were detected in overloaded muscle from losartan-treated rats. Similarly, we observed elevated VEGF production in cultured endothelial cells treated with losartan alone or in combination with Ang II. These studies conclusively establish the requirement for muscle derived VEGF in overload-induced angiogenesis and highlight a role for Ang II in basal VEGF production in skeletal muscle. However, while Ang II signaling is activated following overload and plays a role in muscle VEGF production, inhibition of this pathway is not sufficient to halt overload-induced angiogenesis, indicating that AT1-independent signals maintain VEGF production in losartan-treated muscle.
骨骼肌超负荷会诱导血管生成因子如血管内皮生长因子(VEGF)和基质金属蛋白酶(MMP)-2的表达,从而导致新的毛细血管生长。我们发现,在肌肉VEGF基因敲除小鼠中,超负荷诱导的血管生成增加以及VEGF、MMP-2和MT1-MMP转录本的增加被消除,这突出了肌细胞源性VEGF在控制这一过程中的关键作用。导致超负荷诱导VEGF表达的上游介质尚未确定。我们发现肌肉超负荷会增加血管紧张素原的表达,血管紧张素原是血管紧张素(Ang)II的前体,并且Ang II信号在C2C12细胞的基础VEGF产生中起重要作用。此外,成肌细胞释放的与基质结合的VEGF诱导内皮细胞活化,内皮细胞磷酸化p38水平升高证明了这一点。我们还发现外源性Ang II会提高C2C12肌管中VEGF的表达以及MMP-2转录本水平。有趣的是,在骨骼肌内皮细胞对Ang II处理的反应中也观察到了这些反应,表明这些细胞也能直接对刺激做出反应。评估了Ang II在肌肉超负荷诱导的血管生成中的作用。我们发现使用氯沙坦阻断AT1R依赖性Ang II信号并不会减弱毛细血管生长。令人惊讶的是,在氯沙坦处理的大鼠的超负荷肌肉中检测到VEGF蛋白水平升高。同样,我们观察到单独用氯沙坦或与Ang II联合处理的培养内皮细胞中VEGF产生增加。这些研究最终确定了肌肉源性VEGF在超负荷诱导的血管生成中的必要性,并突出了Ang II在骨骼肌基础VEGF产生中的作用。然而,虽然Ang II信号在超负荷后被激活并在肌肉VEGF产生中起作用,但抑制该途径不足以阻止超负荷诱导的血管生成,这表明在氯沙坦处理的肌肉中,不依赖AT1的信号维持VEGF的产生。