Elliott Winston H, Tan Yan, Li Min, Tan Wei
Department of Mechanical Engineering, University of Colorado at Boulder, 1111 Engineering Dr, ECME 114, Boulder, CO 80309-0427 USA.
Department of Pediatrics, University of Colorado at Denver, Aurora, CO 80045 USA.
Cell Mol Bioeng. 2015 Mar 19;8(2):285-295. doi: 10.1007/s12195-015-0386-7. eCollection 2015 Jun.
Vascular fibrosis, the formation of excess fibrous tissue on the blood vessel wall, is characterized by unmitigated proliferation of fibroblasts or myofibroblast-like cells exhibiting α-smooth-muscle-actin in vessel lumen and other vascular layers. It likely contributes to vascular unresponsiveness to conventional therapies. This paper demonstrates a new flow-induced vascular fibrosis mechanism. Using our developed flow system which simulates the effect of vessel stiffening and generates unidirectional high pulsatility flow (HPF) with the mean shear flow at a physiological level, we have shown that HPF caused vascular endothelial dysfunction. Herein, we further explored the role of HPF in vascular fibrosis through endothelial-to-mesenchymal transdifferentiation (EndMT). Pulmonary arterial endothelial cells (ECs) were exposed to steady flow and HPF, which have the same physiological mean fluid shear but different in flow pulsatility. Cells were analyzed after being conditioned with flows for 24 or 48 h. HPF was found to induce EndMT of cells after 48 h stimulation; cells demonstrated drastically decreased expression in EC marker CD31, as well as increased transforming growth factor β, α-SMA, and collagen type-I, in both gene and protein expression profiles. Using the flow media from HPF-conditioned endothelial culture to cultivate arterial adventitial fibroblasts (AdvFBs) and ECs respectively, we found that the conditioned media respectively enhanced migration, proliferation and α-SMA expression of AdvFBs, and induced EndMT of ECs. It was further revealed that cells exposed to HPF exhibited much higher percentage of caspase-positive cells compared to those exposed to steady flow. Apoptotic cells together with remaining, caspase-negative cells suggested the presence of apoptosis-resistant dysfunctional ECs which likely underwent EndMT process and perpetuated fibrosis throughout vascular tissues. Therefore, our results indicate that prolonged HPF stimuli induce vascular fibrosis through triggering EndMT and EC-mediated AdvFB activation and migration, which follows initial endothelial inflammation, dysfunction and apoptosis.
血管纤维化是指在血管壁上形成过多的纤维组织，其特征是成纤维细胞或肌成纤维细胞样细胞在血管腔和其他血管层中不受控制地增殖，并表现出α-平滑肌肌动蛋白。它可能导致血管对传统疗法无反应。本文展示了一种新的血流诱导的血管纤维化机制。利用我们开发的流动系统，该系统模拟血管硬化的效果并产生生理水平平均剪切流的单向高搏动性血流（HPF），我们已经表明HPF会导致血管内皮功能障碍。在此，我们通过内皮-间充质转化（EndMT）进一步探讨了HPF在血管纤维化中的作用。将肺动脉内皮细胞（ECs）暴露于稳定流和HPF中，它们具有相同的生理平均流体剪切力，但血流搏动性不同。在用这些流处理细胞24或48小时后对细胞进行分析。发现HPF在刺激48小时后诱导细胞发生EndMT；在基因和蛋白质表达谱中，细胞的内皮细胞标志物CD31表达急剧下降，同时转化生长因子β、α-SMA和I型胶原蛋白表达增加。分别用来自HPF处理的内皮细胞培养物的流动培养基培养动脉外膜成纤维细胞（AdvFBs）和ECs，我们发现条件培养基分别增强了AdvFBs的迁移、增殖和α-SMA表达，并诱导了ECs的EndMT。进一步发现，与暴露于稳定流的细胞相比，暴露于HPF的细胞中半胱天冬酶阳性细胞的百分比要高得多。凋亡细胞与剩余的半胱天冬酶阴性细胞表明存在抗凋亡的功能失调的ECs，这些细胞可能经历了EndMT过程并使整个血管组织中的纤维化持续存在。因此，我们的结果表明，长时间的HPF刺激通过触发EndMT以及EC介导的AdvFB激活和迁移来诱导血管纤维化，这继发于最初的内皮炎症、功能障碍和凋亡。
