Combined Effect of Osteopontin and BMP-2 Derived Peptides Grafted to an Adhesive Hydrogel on Osteogenic and Vasculogenic Differentiation of Marrow Stromal Cells
Combined Effect of Osteopontin and BMP-2 Derived Peptides Grafted to an Adhesive Hydrogel on Osteogenic and Vasculogenic Differentiation of Marrow Stromal Cells
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DOI：
10.1021/la205005h
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发表时间：
2012-03-27
期刊：
影响因子：
3.9
通讯作者：
Jabbari, Esmaiel
中科院分区：
文献类型：
--
作者：
He, Xuezhong;Yang, Xiaoming;Jabbari, Esmaiel
本工作的目的是研究将骨桥蛋白（OPD肽）的氨基酸残基162-168对应的肽和骨形态发生蛋白-2（BMP肽）的氨基酸残基73-92对应的肽移植到RGD缀合的惰性水凝胶上对骨髓基质（BMS）细胞的成骨和血管生成分化的联合作用。通过熔融沉积成型快速成型系统制备牺牲蜡模，制备具有明确圆柱形孔几何形状的RGD缀合的三维（3D）多孔水凝胶支架。将丙烯酸炔丙酯和4-戊烯醛分别通过点击反应和肟连接法偶联到水凝胶上，用于BMP和OPD肽的正交接枝。通过氨氧基-mPEG-OPD（mPEG =微型聚（乙二醇））的氨氧基部分与水凝胶中的醛部分之间的反应来接枝OPD肽。通过Az-mPEG-BMP的叠氮化物部分与水凝胶中的炔丙基部分之间的反应来接枝BMP肽。通过生物化学、免疫细胞化学和mRNA分析表征接种有BMS细胞的水凝胶。组包括RGD对照水凝胶（RGD）、不含OPD的RGD和BMP肽（RGD+BMP）、具有突变OPD的RGD和BMP肽（RGD+BMP+mOPD）以及具有OPD的RGD和BMP肽（RGD+BMP+OPD）接枝的水凝胶。28天后，RGD、RGD+BMP、RGD+BMP+mOPD和RGD+BMP+OPD组的矿化程度分别为650 +/- 70、990 +/- 30、850 +/- 30和1150 +/- 40 mg/（mg DNA），表明BMP和OPD肽增强了BMS细胞的成骨分化。接种在RGD+BMP +OPD移植水凝胶上的BMS细胞对血管生成标志物α-SMA、PECAM-1和VE-钙粘蛋白染色呈阳性，而没有OPD肽的组（RGD+BMP和RGD+BMP+mOPD）仅对α-SMA染色，而不对PECAM-1或VE-钙粘蛋白染色。这些结果与在21和28天后，与没有OPD的组相比，RGD+BMP+OPD组的显著更高的PECAM-1 mRNA表达一致。这些发现表明，RGD+BMP+OPD肽为骨髓来源的祖细胞的同时成骨和血管生成分化提供了有利的微环境。
The objective of this work was to investigate the combined effect of grafting the peptide corresponding to amino acid residues 162-168 of osteopontin (OPD peptide) and the peptide corresponding to amino acid residues 73-92 of bone morphogenetic protein-2 (BMP peptide) to an RGD-conjugated inert hydrogel on osteogenic and vasculogenic differentiation of bone marrow stromal (BMS) cells. RGD-conjugated three-dimensional (3D) porous hydrogel scaffolds with well-defined cylindrical pore geometry were produced from sacrificial wax molds fabricated by fused deposition modeling rapid prototyping system. Propargyl acrylate and 4-pentenal were conjugated to the hydrogel for orthogonal grafting of BMP and OPD peptides by click reaction and oxime ligation, respectively. The OPD peptide was grafted by the reaction between aminooxy moiety of aminooxy-mPEG-OPD (mPEG = mini-poly(ethylene glycol)) and the aldehyde moiety in the hydrogel. The BMP peptide was grafted by the reaction between the azide moiety of Az-mPEG-BMP and the propargyl moiety in the hydrogel. The hydrogels seeded with BMS cells were characterized by biochemical, immunocytochemical, and mRNA analyses. Groups included RGD control hydrogel (RGD), RGD and BMP peptides without OPD (RGD+BMP), RGD and BMP peptides with mutant OPD (RGD+BMP+mOPD), and RGD and BMP peptides with OPD (RGD+BMP+OPD) grafted hydrogels. The extent of mineralization of RGD, RGD+BMP, RGD+BMP+mOPD, and RGD+BMP+OPD groups after 28 days was 650 +/- 70, 990 +/- 30, 850 +/- 30, and 1150 +/- 40 mg/(mg of DNA), respectively, indicating that the BMP and OPD peptides enhanced osteogenic differentiation of the BMS cells. The BMS cells seeded on RGD+BMP +OPD grafted hydrogels stained positive for vasculogenic markers alpha-SMA, PECAM-1, and VE-cadherin while the groups without OPD peptide (RGD+BMP and RGD+BMP+mOPD) stained only for alpha-SMA but not PECAM-1 or VE-cadherin. These results were consistent with the significantly higher PECAM-1 mRNA expression for RGD+BMP+OPD group after 21 and 28 days, compared to the groups without OPD. These findings suggest that the RGD+BMP+OPD peptides provide a favorable microenvironment for concurrent osteogenic and vasculogenic differentiation of progenitor marrow-derived cells.