Diaz Quiroz Juan Felipe, Rodriguez Patricia Diaz, Erndt-Marino Josh D, Guiza Viviana, Balouch Bailey, Graf Tyler, Reichert William M, Russell Brooke, Höök Magnus, Hahn Mariah S
Department of Biomedical Engineering, Rensselaer Polytechnic Institute, Troy, New York 12180, United States.
Department of Biomedical Engineering, Duke University, Durham, North Carolina 27708, United States.
ACS Biomater Sci Eng. 2018 Aug 13;4(8):2934-2942. doi: 10.1021/acsbiomaterials.8b00070. Epub 2018 Jun 29.
Achieving graft endothelialization following implantation continues to be a challenge in the development of "off-the-shelf," small-caliber, arterial prostheses. Coating grafts with biomolecules to support the retention, migration, and differentiation of adherent endothelial precursor cells (EPCs) is a promising approach toward improving graft endothelialization. esigner ollagen Scl2- with integrin binding site per strand (DC2-1X) is a -derived, collagen-like protein that has previously been evaluated as a graft coating due to its ability to resist platelet aggregation and to promote attachment and migration of "late outgrowth" EPCs (EOCs). However, these prior assessments were performed in the absence of physiological shear. In addition, although DC2-1X coatings supported increased migration rates relative to native collagen coatings, EOC attachment and spreading remained inferior to collagen controls at all DC2-1X concentrations assayed. Thus, the objectives of the present work were the following: (1) to improve EOC attachment on DC2 coatings by modulating the number and spacing of DC2 integrin binding sites (IBS) and (2) to evaluate the retention, migration, and differentiation of adherent EOCs under physiological shear stress. Using single point mutations, three novel DC2 variants were generated containing either two IBS (DC2-2X) or three IBS (DC2-3X1 and DC2-3X2) per strand. After initial evaluation of the potential of each DC2 variant to support increased EOC attachment relative to DC2-1X, DC2-2X and DC2-3X1 coatings were further assessed under physiological shear for their capacity to promote EOC retention, migration, and differentiation relative to DC2-1X and collagen controls. An increase in the number of IBS from 1 to 3 significantly improved EOC retention on DC2 coatings while also supporting increased average migration rates. Moreover, EOCs on DC2-3X1 coatings showed increased gene-level expression of intermediate endothelial cell differentiation markers relative to collagen. Overall, the current results suggest that DC2-3X1 warrants further investigation as a vascular graft coating.
在“现货供应”的小口径动脉假体的研发过程中，实现植入后移植物的内皮化仍然是一项挑战。用生物分子包覆移植物以支持黏附的内皮祖细胞（EPC）的保留、迁移和分化，是改善移植物内皮化的一种有前景的方法。每条链带有一个整合素结合位点的设计型胶原蛋白Scl2-（DC2-1X）是一种源自胶原蛋白的类胶原蛋白，因其具有抵抗血小板聚集以及促进“晚期生长型”EPC（EOC）黏附与迁移的能力，此前已被评估用作移植物涂层。然而，这些先前的评估是在无生理剪切力的情况下进行的。此外，尽管相对于天然胶原蛋白涂层，DC2-1X涂层能支持更高的迁移率，但在所有检测的DC2-1X浓度下，EOC的黏附与铺展仍逊于胶原蛋白对照。因此，本研究的目的如下：（1）通过调节DC2整合素结合位点（IBS）的数量和间距来改善EOC在DC2涂层上的黏附，以及（2）评估在生理剪切应力下黏附的EOC的保留、迁移和分化情况。利用单点突变，生成了三种新型DC2变体，每条链分别含有两个IBS（DC2-2X）或三个IBS（DC2-3X1和DC2-3X2）。在初步评估每个DC2变体相对于DC2-1X支持EOC黏附增加的潜力后，进一步在生理剪切条件下评估DC2-2X和DC2-3X1涂层相对于DC2-1X和胶原蛋白对照促进EOC保留、迁移和分化的能力。IBS数量从1增加到3显著改善了EOC在DC2涂层上的保留，同时也支持平均迁移率的提高。此外，相对于胶原蛋白，DC2-3X1涂层上的EOC显示出中间内皮细胞分化标志物基因水平表达的增加。总体而言，目前的结果表明DC2-3X1作为血管移植物涂层值得进一步研究。
