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Duchi Serena,Francis Sam L.,Onofrillo Carmine,O’Connell Cathal D.,Choong Peter,Di Bella Claudia 한국조직공학과 재생의학회 2023 조직공학과 재생의학 Vol.20 No.1
BACKGROUND: Articular cartilage repair using implantable photocrosslinkable hydrogels laden with chondrogenic cells, represents a promising in situ cartilage engineering approach for surgical treatment. The development of a surgical procedure requires a minimal viable product optimized for the clinical scenario. In our previous work we demonstrated how gelatin based photocrosslinkable hydrogels in combination with infrapatellar derived stem cells allow the production of neocartilage in vitro. In this study, we aim to optimize the critical facets of the in situ cartilage engineering therapy: the cell source, the cell isolation methodology, the cell expansion protocol, the cell number, and the delivery approach. METHODS: We evaluated the impact of the critical facets of the cell-laden hydrogel therapy in vitro to define an optimized protocol that was then used in a rabbit model of cartilage repair. We performed cells counting and immunophenotype analyses, chondrogenic potential evaluation via immunostaining and gene expression, extrusion test analysis of the photocrosslinkable hydrogel, and clinical assessment of cartilage repair using macroscopic and microscopic scores. RESULTS: We identified the adipose derived stem cells as the most chondrogenic cells source within the knee joint. We then devised a minimally manipulated stem cell isolation procedure that allows a chondrogenic population to be obtained in only 85 minutes. We found that cell expansion prior to chondrogenesis can be reduced to 5 days after the isolation procedure. We characterized that at least 5 million of cells/ml is needed in the photocrosslinkable hydrogel to successfully trigger the production of neocartilage. The maximum repairable defect was calculated based on the correlation between the number of cells retrievable with the rapid isolation followed by 5-day non-passaged expansion phase, and the minimum chondrogenic concentration in photocrosslinkable hydrogel. We next optimized the delivery parameters of the cell-laden hydrogel therapy. Finally, using the optimized procedure for in situ tissue engineering, we scored superior cartilage repair when compared to the gold standard microfracture approach. CONCLUSION: This study demonstrates the possibility to repair a critical size articular cartilage defect by means of a surgical streamlined procedure with optimized conditions.
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