Directing human mesenchymal stem cells differentiation towards hypertrophic chondrocytes using fiber-reinforced bone dECM hydrogel scaffolds

Jun 29, 2022, 2:30 PM
Room: S4 A

Room: S4 A


Idaszek, Joanna (Warsaw University of Technology)


Intramembranous ossification is the most common pathway used in tissue engineering (TE) of bone. However, therapeutic effects in case of large bone defects are suboptimal due to hindered vascularization. On the other hand, TE grafts generated via the endochondral ossification pathway, where the bone formation occurs through cartilage intermediaries, become vascularized and form bone upon implantation. In order to build bone, the chondrocytes have to become hypertrophic. The goal of this study was to develop fiber-reinforced bone decellularized extracellular matrix (bdECM)-based hydrogels for enhanced hypertrophic differentiation of chondrogenicly primed human bone marrow MSC (hMSC). </div>

<div>The hydrogels were composed of either 10 mg/ml type I collagen or bdECM mixed with alginate (30 mg/ml). As the reinforcement, 3D fibrous scaffolds were fabricated using PCL and PCL-based composites containing 25 wt% of β-TCP and FDM technology. HMSC were encapsulated in the pre-gel at density of 20x106/ml, infused into the 3D scaffolds (50 μl of cell suspension/ construct) and cultured in chondrogenic medium for 28 days. To measure cell viability, live/dead staining and MTS assay were performed. Secretion of cartilaginous matrix was evaluated by staining for safranin O, collagen type I, II and X. Moreover, quantification of GAGs was performed using 1,9-dimethylmethylene blue assay and collagen using total collagen assay. Additionally, alkaline phosphatase (ALP) activity was measured using colorimetric assay with para-nitrophenyl phosphate as a substrate. </div> <div>The cell metabolic activity did not decrease upon culture, although the overall viability measured by live/dead staining decreased, suggesting cell proliferation. A synergistic effect of the dECM and β-TCP on hypertrophic differentiation of hMSC was measured my means increased ALP activity and visualized by more intense staining against type X collagen. The aforementioned finding suggests that bone dECM hydrogels reinforced with β-TCP/PCL scaffolds are suitable materials for chondrogenic priming of MSC and bone TE via osteochondral pathway. 52354583529

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