One-pot Synthesis of Elastin-like Polypeptide Hydrogels with Grafted VEGF-Mimetic Peptides.
One-pot Synthesis of Elastin-like Polypeptide Hydrogels with Grafted VEGF-Mimetic Peptides.
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DOI：
10.1039/c3bm60293a
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发表时间：
2014-05-01
影响因子：
6.6
通讯作者：
Heilshorn SC
中科院分区：
文献类型：
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作者：
Cai L;Dinh CB;Heilshorn SC
将生长因子固定到聚合物基质上是组织工程支架设计中的常见策略，以促进组织再生，这需要与周围基质发生复杂的细胞信号传导事件。然而，在聚合物支架中使用大的蛋白质生长因子通常受到免疫原性、短的体内半衰期和降低的生物活性的困扰。为了解决这些问题，我们开发了一种单步的，细胞相容的策略，将小的生长因子模拟肽系到具有可调生物材料特性的蛋白质工程水凝胶中。具体来说，我们共价键的QK肽，血管生成肽模仿血管内皮生长因子（VEGF）的受体结合区，可调弹性蛋白样多肽（ELP）水凝胶，包括细胞粘附性的RGD序列。使用细胞相容的胺反应性交联剂，我们进行了一锅法合成，同时封装细胞，同时精确控制ELP水凝胶中的QK接枝密度（10 nM - 100 μM），而不改变其他材料特性。荧光标记的QK肽的荧光分析表明，与ELP水凝胶的缀合效率>75%，并且共价固定有效地消除了所有QK扩散。与原始ELP水凝胶相比，10 nM或1 μM QK固定的ELP水凝胶显著增强了人脐静脉内皮细胞（HUVEC）的增殖。此外，在栓系的QK-ELP水凝胶内包封后，HUVEC球状体保持接近100%的活力，并且与补充有相同浓度的可溶性QK肽的那些相比，显示出显著更多的三维生长。这些结果鼓励进一步开发用生长因子模拟肽修饰的蛋白质工程支架，以使用这种通用的单步合成提供长期的生物信号。
Immobilization of growth factors to polymeric matrices has been a common strategy in the design of tissue engineering scaffolds to promote tissue regeneration, which requires complex cell signaling events with the surrounding matrix. However, the use of large protein growth factors in polymeric scaffolds is often plagued by immunogenicity, short in vivo half-lives, and reduced bioactivity. To address these concerns, we develop a single-step, cell-compatible strategy to tether small, growth-factor-mimetic peptides into a protein-engineered hydrogel with tunable biomaterial properties. Specifically, we covalently immobilize the QK peptide, an angiogenic peptide mimicking the receptor-binding region of vascular endothelial growth factor (VEGF), within tunable elastin-like polypeptide (ELP) hydrogels that include a cell-adhesive RGD sequence. Using a cell-compatible, amine-reactive crosslinker, we conducted a one-pot synthesis to simultaneously encapsulate cells while precisely controlling the QK grafting density (10 nM – 100 μM) in the ELP hydrogels without altering other material properties. Fluorescence analysis of fluor-labeled QK peptides demonstrated that the conjugation efficiency to ELP hydrogels was >75% and that covalent immobilization effectively eliminates all QK diffusion. Compared with pristine ELP hydrogels, human umbilical vein endothelial cell (HUVEC) proliferation was significantly enhanced on ELP hydrogels immobilized with 10 nM or 1 μM QK. Moreover, upon encapsulation within tethered QK-ELP hydrogels, HUVEC spheroids maintained near 100% viability and demonstrated significantly more three-dimensional outgrowth compared to those supplemented with soluble QK peptide at the same concentration. These results encourage the further development of protein-engineered scaffolds decorated with growth-factor-mimetic peptides to provide long-term biological signals using this versatile, single-step synthesis.