The Effect of Fibrinogen, Collagen Type I, and Fibronectin on Mesenchymal Stem Cell Growth and Differentiation into Osteoblasts
The Effect of Fibrinogen, Collagen Type I, and Fibronectin on Mesenchymal Stem Cell Growth and Differentiation into Osteoblasts
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DOI：
10.1089/ten.tea.2012.0523
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发表时间：
2013-06-01
影响因子：
4.1
通讯作者：
Tawil, Bill
中科院分区：
文献类型：
--
作者：
Linsley, Chase;Wu, Benjamin;Tawil, Bill
我们已经证明，当种植在三维(3-D)纤维蛋白结构中时，人间充质干细胞(HMSCs)具有分化为骨的潜力。纤维蛋白结构的内源性蛋白质和3D结构中细胞分泌的蛋白质提供了促进hMSCs分化的线索，以及对细胞生长和迁移的机械支持。在这项研究中，我们决定退一步，检测不同的细胞外基质(ECM)蛋白-纤维蛋白原、纤维连接蛋白和I型胶原-在二维(2-D)单层培养中对hMSC成骨分化的影响。简单地说，24孔组织培养板预涂有纤维蛋白原(10 mg/mL)、纤维连接蛋白(10 mU g/m L)或I型胶原(1 mg/m L)，接种25,000个细胞/孔，在正常生长培养基或成骨诱导培养基中培养。在培养的第1、7、14、21和30天，使用AlamarBlue(R)和碱性磷酸酶及Von Kossa染色的成骨指示剂评估细胞生长。结果表明，I型胶原在成骨培养液和标准培养液中的钙沉积阳性染色在30d时达到最大。21d时纤维蛋白原在成骨基质中的染色最强，21d时纤维连接蛋白是唯一促进钙沉积的底物。这些结果表明，hMSCs的成骨分化既受培养条件的影响，又受基质的影响，两者具有协同作用。通过了解三维纤维蛋白结构中的ECM蛋白在促进hMSCs成骨分化方面的作用，将改进复杂的仿生模型的构建，以操纵hMSC向成骨细胞系分化。
We have shown that human mesenchymal stem cells (hMSCs) have the potential to differentiate into bone when seeded within three-dimensional (3-D) fibrin constructs. Proteins endogenous to the fibrin construct and those secreted by cells in the 3-Dconstructs provide cues that can promote differentiation of hMSCs along with mechanical support for cell growth and migration. In this study, we decided to take a step back and examine the effect different extracellular matrix (ECM) proteins-fibrinogen, fibronectin, and collagen type I-had on hMSC osteogenic differentiation on two-dimensional (2-D) monolayer cultures. Briefly, 24-well tissue culture plates pre-coated with either fibrinogen (10 mg/mL), fibronectin (10 mu g/mL), or collagen type I (1 mg/mL) were seeded with 25,000 cells/well and cultured in normal growth medium or in osteogenic induction medium. At days 1, 7, 14, 21, and 30, cultures were assessed for cell growth using alamarBlue (R) and osteogenic indicators using alkaline phosphatase and Von Kossa staining. The results show that collagen type I stained positive for calcium deposition the greatest by day 30 in both osteogenic medium and standard culture medium. However, fibrinogen had the greatest staining in osteogenic medium at day 21 and fibronectin was the only substrate to promote calcium deposition in standard culture medium at day 21. These results suggest that the osteogenic differentiation of hMSCs is influenced by both culturing conditions and substrate and that together they have a synergistic effect. By knowing the effect ECM proteins in 3-D fibrin construct have on promoting osteogenic differentiation of hMSCs, the fabrication of complex, biomimetic models designed to manipulate hMSC differentiation toward an osteoblastic lineage will be improved.