Sustained release of polyethylene glycol-modified hydrogel-packaged growth factors and endothelial progenitor cells

doi:10.3969/j.issn.2095-4344.2014.30.015

Abstract

Abstract:

BACKGROUND: Many studies have demonstrated that in order to achieve the endothelialization of the polymer materials, surface loading of bioactive factors is an important means and the introduction of human endothelial cells on the surface of the polymer materials will contribute to increase the biocompatibility of materials.
OBJECTIVE: To synthesize the polyethylene glycol hydrogel packaging vascular endothelial growth factor, basic fibroblast growth factor and endothelial progenitor cells, and to observe the sustained release of growth factors and culture status of endothelial progenitor cells.
METHODS: The polyethylene glycol-modified hydrogel containing arginine-glycine-aspartic acid peptide was synthesized, and then, rat endothelial progenitor cells, basic fibroblast growth factor, vascular endothelial growth factor, matrix metalloproteinases were successively added. The hydrogel were immersed in PBS, and ELISA was used to test the levels of growth factors in supernatants every 12 hours. Matrix metalloproteinases-2 (100, 1 000 ng) and matrix metalloproteinases-9 (100, 1 000 ng) at different mass were added into the PBS containing hydrogel after 72 hours. ELISA was also used to test the levels of growth factors in supernatants every 12 hours. The hydrogel encapsulating endothelial progenitor cells was cultured in a culture medium for 5 days, and the number of survival cells was counted by a flow cytometer after digestion.
RESULTS AND CONCLUSION: Within 12-72 hours, the release percentage of basic fibroblast growth factor and vascular endothelial growth factor remained at about 41%. After 72 hours, the release percentage of both growth factors was found to grow steadily when matrix metalloproteinases-2 (100, 1000 ng) and matrix metalloproteinases-9 (100, 1 000 ng) at different mass were added, which reached 95%. The release percentage was increased with the increasing mass of matrix metalloproteinases. After 5 days of culture, 88.17% cells still survived in the hydrogel. These findings indicate that sell-assembly polyethylene glycol-modified hydrogel can realize the controlled release of basic fibroblast growth factor and vascular endothelial growth factor, and it can also support the proliferative growth of endothelial progenitor cells.

中国组织工程研究杂志出版内容重点：生物材料；骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性；组织工程

全文链接：

Key words: polyethylene glycols, hydrogel, fibroblast growth factors, vascular endothelial growth factors

CLC Number:

R318

Han Yan-jiu, Liu Guo-hui, Ouyang Liu. Sustained release of polyethylene glycol-modified hydrogel-packaged growth factors and endothelial progenitor cells[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(30): 4851-4856.

Figures/Tables 2

2.1 RT-PCR 法检测内皮祖细胞特异性分子标志内皮祖细胞特异性分子标志vWF、Flk-1、Tie-2、CD34、CD133、eNOS mRNA均呈阳性表达(图1)。 2.2 基质金属蛋白酶介导的改性聚乙二醇水凝胶碱性成纤维细胞生长因子、血管内皮生长因子体外释放性能 2.2.1 不同基质金属蛋白酶对改性聚乙二醇水凝胶中碱性成纤维细胞生长因子体外释放的影响基质金属蛋白酶的质量对改性聚乙二醇水凝胶中碱性成纤维细胞生长因子释放曲线及最大释放量的影响不明显。72 h前，基质金属蛋白酶存在与否及基质金属蛋白酶的质量对改性水凝胶中碱性成纤维细胞生长因子的释放均不明显；72 h后，基质金属蛋白酶2及基质金属蛋白酶9均能明显增加水凝胶内碱性成纤维细胞生长因子的释放，148 h后基本达释放的平台期。改性水凝胶内碱性成纤维细胞生长因子的释放对基质金属蛋白酶有一定的时间依赖性(图2，3)。 2.2.2 不同基质金属蛋白酶对改性聚乙二醇水凝胶中血管内皮生长因子体外释放的影响通过绘制细胞因子体外缓释曲线，可以观察基质金属蛋白酶2同基质金属蛋白酶9相比使改性聚乙二醇水凝胶内血管内皮生长因子释放较为平缓，但是基质金属蛋白酶的质量对改性聚乙二醇水凝胶内血管内皮生长因子释放的影响不明显。72 h前，基质金属蛋白酶存在与否及基质金属蛋白酶的质量对改性水凝胶内血管内皮生长因子的释放亦均不明显；72 h后，基质金属蛋白酶2及基质金属蛋白酶9亦均能明显增加水凝胶内血管内皮生长因子的释放，148 h后基本达释放的平台期，说明改性水凝胶内血管内皮生长因子的释放对基质金属蛋白酶亦存在一定的时间依赖性(图4，5)。 2.3 改性聚乙二醇水凝胶封装内皮祖细胞后对细胞活性的影响通过Annexin V-FITC/PI双染，流式细胞仪检测存活细胞数的方法。正常对照组细胞凋亡细胞比例为(5.36± 3.24)%，活细胞比例为(95.983±4.76)%；水凝胶封装后细胞凋亡比例为(9.13±4.13)%，活细胞比例为(88.17±5.17)%。经组间t 检验分析，实验组和正常对照组间凋亡细胞比例和活细胞比例差异均无显著性意义(P > 0.05)，说明改性水凝胶对内皮祖细胞的活性无明显不利影响(图6)。"

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