Biomedical Engineering Department, Northwestern University, Evanston, IL 60208, USA.
Acta Biomater. 2010 Jan;6(1):3-11. doi: 10.1016/j.actbio.2009.07.031. Epub 2009 Jul 25.
There is great clinical interest in cell-based therapies for ischemic tissue repair in cardiovascular disease. However, the regenerative potential of these therapies is limited due to poor cell viability and minimal retention following application. We report here the development of bioactive peptide amphiphile nanofibers displaying the fibronectin-derived RGDS cell adhesion epitope as a scaffold for therapeutic delivery of bone marrow derived stem and progenitor cells. When grown on flat substrates, a binary peptide amphiphile system consisting of 10 wt.% RGDS-containing molecules and 90wt.% negatively charged diluent molecules was found to promote optimal cell adhesion. This binary system enhanced adhesion 1.4-fold relative to substrates composed of only the non-bioactive diluent. Additionally, no enhancement was found upon scrambling the epitope and adhesion was no longer enhanced upon adding soluble RGDS to the cell media, indicating RGDS-specific adhesion. When encapsulated within self-assembled scaffolds of the binary RGDS nanofibers in vitro, cells were found to be viable and proliferative, increasing in number by 5.5 times after only 5 days, an effect again lost upon adding soluble RGDS. Cells encapsulated within a non-bioactive scaffold and those within a binary scaffold with scrambled epitope showed minimal viability and no proliferation. Cells encapsulated within this RGDS nanofiber gel also increase in endothelial character, evident by a decrease in the expression of CD34 paired with an increase in the expression of endothelial-specific markers VE-Cadherin, VEGFR2 and eNOS after 5 days. In an in vivo study, nanofibers and luciferase-expressing cells were co-injected subcutaneously in a mouse model. The binary RGDS material supported these cells in vivo, evident by a 3.2-fold increase in bioluminescent signal attributable to viable cells; this suggests the material has an anti-apoptotic and/or proliferative effect on the transplanted bone marrow cells. We conclude that the binary RGDS-presenting nanofibers developed here demonstrate enhanced viability, proliferation and adhesion of associated bone marrow derived stem and progenitor cells. This study suggests potential for this material as a scaffold to overcome current limitations of stem cell therapies for ischemic diseases.
人们对细胞疗法在心血管疾病缺血组织修复中的应用具有浓厚的临床兴趣。然而，由于细胞活力差和应用后保留率低，这些疗法的再生潜力有限。我们在此报告了一种生物活性肽两亲纳米纤维的开发，该纤维展示了纤维连接蛋白衍生的 RGDS 细胞黏附表位，可用作骨髓来源的干细胞和祖细胞治疗递送的支架。在平面基底上，发现由 10wt%含有 RGDS 的分子和 90wt%带负电荷的稀释剂分子组成的二元肽两亲体系促进了最佳的细胞黏附。与仅由非生物活性稀释剂组成的基底相比，该二元体系使黏附增强了 1.4 倍。此外，当对表位进行乱序时，没有发现增强作用，并且当将 RGDS 添加到细胞培养基中时，黏附不再增强，表明 RGDS 具有特异性黏附。在体外，将细胞包封在二元 RGDS 纳米纤维的自组装支架内时，发现细胞保持活力和增殖，仅在 5 天后数量增加了 5.5 倍，当添加可溶性 RGDS 时，这种作用再次丧失。包封在非生物活性支架内的细胞和包封在具有乱序表位的二元支架内的细胞显示出最小的活力并且没有增殖。包封在这种 RGDS 纳米纤维凝胶中的细胞也增加了内皮特性，这表现为 CD34 的表达减少，同时内皮特异性标志物 VE-Cadherin、VEGFR2 和 eNOS 的表达增加，5 天后达到 5 天。在体内研究中，将纳米纤维和表达荧光素酶的细胞在小鼠模型中皮下共注射。二元 RGDS 材料在体内支持这些细胞，这表现为归因于存活细胞的生物发光信号增加了 3.2 倍；这表明该材料对移植的骨髓细胞具有抗凋亡和/或增殖作用。我们得出结论，这里开发的二元 RGDS 呈现纳米纤维增强了相关骨髓来源的干细胞和祖细胞的活力、增殖和黏附。这项研究表明，该材料作为支架具有克服缺血性疾病中干细胞治疗当前局限性的潜力。
