In vitro osteogenic differentiation of human mesenchymal stem cells photoencapsulated in PEG hydrogels

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Abstract
Much research has focused on the differentiation of human mesenchymal stem cells (hMSCs) in monolayer culture; however, little is known about their differentiation potential in three-dimensional culture conditions. In this research, hMSCs were encapsulated in a photocrosslinkable, injectable scaffolding system based on poly(ethylene glycol) (PEG) hydrogels. To demonstrate the ability of hMSCs to differentiate in PEG hydrogels, cell/polymer constructs were cultured in osteogenic differentiation media to elicit an osteoblastic response. First, viability of encapsulated hMSCs up to 4 weeks in culture was investigated using a membrane integrity assay. Second, gene expression of encapsulated cells was determined with reverse transcription polymerase chain reaction (RT-PCR) as a function of media composition. After 1 week in osteogenic differentiation media, encapsulated hMSCs expressed osteonectin, osteopontin, and alkaline phosphatase, which are all characteristic of osteoblasts. Finally, von Kossa staining was used to evaluate mineralization of the PEG gels. Results support the hypothesis that hMSCs photoencapsulated in PEG hydrogels and cultured in the presence of osteogenic differentiation media are able to differentiate to osteoblasts inside the gel and mineralize the matrix. These experiments demonstrate the feasibility of using a PEG-based, photocrosslinkable system to culture and deliver human mesenchymal stem cells for bone tissue regeneration and repair. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 68A: 773–782, 2004