In vitro elastogenesis: instructing human vascular smooth muscle cells to generate an elastic fiber-containing extracellular matrix scaffold
In vitro elastogenesis: instructing human vascular smooth muscle cells to generate an elastic fiber-containing extracellular matrix scaffold
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DOI：
10.1088/1748-6041/10/3/034102
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发表时间：
2015-06-01
影响因子：
4
通讯作者：
Schenke-Layland, Katja
中科院分区：
文献类型：
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作者：
Hinderer, Svenja;Shen, Nian;Schenke-Layland, Katja
弹性纤维对器官的正常功能是必不可少的，包括心血管组织，如心脏瓣膜和血管。尽管之前已经描述了在组织工程构建中生产(Tropo)弹性蛋白，但使用人类细胞在体外组装成功能性弹性纤维一直是非常具有挑战性的。在本研究中，我们将原代分离的人血管平滑肌细胞(VSMCs)种植到3D电纺支架上，并使用定制的生物反应器系统将它们暴露在定义的层流剪应力下。6天后，弹性蛋白表达增加，随后弹性蛋白沉积到电纺支架和新形成的纤维上。最有趣的是，我们鉴定了弹性发生相关蛋白的成功沉积，包括纤维蛋白-1和-2，纤连蛋白-4和-5，纤维连接蛋白，弹性蛋白微纤维界面定位蛋白1(Emily in-1)和赖氨酰氧化酶(LOX)。超微结构分析显示，发育中的细胞外基质(ECM)类似于天然的人类胎儿组织，由胶原蛋白、微纤维和弹性蛋白组成。综上所述，一种新型的动态流动生物反应器和电纺杂化聚合物支架的结合使得含有弹性纤维的ECM的生产和组装成为可能。
Elastic fibers are essential for the proper function of organs including cardiovascular tissues such as heart valves and blood vessels. Although (tropo) elastin production in a tissue-engineered construct has previously been described, the assembly to functional elastic fibers in vitro using human cells has been highly challenging. In the present study, we seeded primary isolated human vascular smooth muscle cells (VSMCs) onto 3D electrospun scaffolds and exposed them to defined laminar shear stress using a customized bioreactor system. Increased elastin expression followed by elastin deposition onto the electrospun scaffolds, as well as on newly formed fibers, was observed after six days. Most interestingly, we identified the successful deposition of elastogenesis-associated proteins, including fibrillin-1 and -2, fibulin-4 and -5, fibronectin, elastin microfibril interface located protein 1 (EMILIN-1) and lysyl oxidase (LOX) within our engineered constructs. Ultrastructural analyses revealed a developing extracellular matrix (ECM) similar to native human fetal tissue, which is composed of collagens, microfibrils and elastin. To conclude, the combination of a novel dynamic flow bioreactor and an electrospun hybrid polymer scaffold allowed the production and assembly of an elastic fiber-containing ECM.