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노용권,Ping Du,김인걸,Jaehoon Ko,Seong Who Kim,Kwideok Park 한국생체재료학회 2016 생체재료학회지 Vol.20 No.2
Background: Tissue-engineered scaffold should mimic the structure and biological function of the extracellular matrix and have mechanically supportive properties for tissue regeneration. In this study, we utilized a PLGA/PLA mesh scaffold, coated with cell-derived extracellular matrix (CDM) and assessed its potential as an osteogenic microenvironment for human umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs). CDM was obtained by decellularization of in vitro-cultured type I collagen overexpressing (Col I -293 T-DK) cells. Test groups are mesh itself (control), fibronectin-coated (FN-mesh), and CDM-coated mesh scaffold (CDM-mesh). CDM was then solubilized and used for scaffold coating. Results: CDM was successfully collected and applied to mesh scaffolds. The presence of CDM was confirmed via SEM and FN immunofluorescence. After then, UCB-MSCs were seeded into the scaffolds and subjected to the induction of osteogenic differentiation for 21 days in vitro. We found that the seeded cells were viable and have better proliferation activity on CDM-mesh scaffold. In addition, when osteogenic differentiation of UCB-MSCs was examined for up to 21 days, alkaline phosphatase (ALP) activity and osteogenic marker (COL I, ALP, osteocalcin, bone sialoprotein) expression were significantly improved with UCB-MSCs when cultured in the CDM-mesh scaffold compared to the control and FN-mesh. Conclusion: Polymer mesh scaffold incorporated with CDM can provide UCB-MSCs with a better microenvironment for osteogenesis in vitro.
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노용권,Ping Du,Avelino Dos Santos Da Costa,박귀덕 한국생체재료학회 2018 생체재료학회지 Vol.22 No.2
Background: Formation of mature and functional articular cartilage is still challenging in cartilage tissue engineering. This study investigates the potential of using heparin-grafted decellularized extracellular matrix (ECM) as a novel growth factor delivery platform towards human placenta-derived mesenchymal stem cells (hPMSCs) chondrogenic differentiation. Human fibroblast-derived extracellular matrix (hFDM) is naturally obtained from in vitro-cultured human lung fibroblasts via a mild decellularization process. hFDM was then conjugated with heparin via N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDC) chemistry and subject to transforming growth factor (TGF)-β1 immobilization. Once heparin graftedhFDM (hFDM-hep) and hPMSCs were co-embedded into collagen gel, they were examined for in vitro and in vivo chondrogenesis of hPMSCs for 4 weeks. Results: We identified heparin moieties on hFDM via toluidine blue O assay and Fourier transform infrared spectroscopy, respectively. We found out that collagen spheroids containing hFDM-hep and TGF-β1 exhibited a sustained release of growth factor for 28 days in vitro. Chondrogenesis of hPMSCs in vitro was supported by accumulated glycosaminoglycan (GAG) content and upregulated chondrogenic specific markers (collagen II, aggrecan, Sox9). Meanwhile, PKH26 - labeled hPMSCs incorporated collagen with either hFDM or hFDM-hep was pre-conditioned in a chondrogenic media for 3 days and subcutaneously implanted in the back of nude mice for 4 weeks. The implanted collagen spheroids containing both hPMSCs and hFDM-hep retained more viable hPMSCs and showed higher level of chondrogenic differentiation, based on immunostaining of collagen type II over collagen alone or Col/hFDM group. In addition, histological examination showed more positive signals of GAG via Safranin-O staining. Conclusion: TGF-β1-immobilized hFDM-hep can provide an appropriate microenvironment for chondrogenic differentiation of hPMSCs in 3D collagen spheroid.
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