Bone marrow and adipose tissue human mesenchymal stem cells were seeded in highly performing 3D gelatin-chitosan hybrid hydrogels of varying chitosan content in the presence of human platelet lysate and evaluated for their proliferation and osteogenic differentiation. Both bone marrow and adipose tissue human mesenchymal stem cells in gelatin-chitosan hybrid hydrogel 1 (chitosan content 8.1%) or gelatin-chitosan hybrid hydrogel 2 (chitosan 14.9%) showed high levels of viability (80%-90%), and their proliferation and osteogenic differentiation was significantly higher with human platelet lysate compared to fetal bovine serum, particularly in gelatin-chitosan hybrid hydrogel 1. Mineralization was detected early, after 21 days of culture, when human platelet lysate was used in the presence of osteogenic stimuli. Proteomic characterization of human platelet lysate highlighted 59 proteins mainly involved in functions related to cell adhesion, cellular repairing mechanisms, and regulation of cell differentiation. In conclusion, the combination of our gelatin-chitosan hybrid hydrogels with hPL represents a promising strategy for bone regenerative medicine using human mesenchymal stem cells.

Biomedical Center Faculty of Medicine in Pilsen Charles University Pilsen Czech Republic

Centre for the Cellular Microenvironment Division of Biomedical Engineering School of Engineering University of Glasgow Glasgow UK

Centro di Ricerca Emato Oncologica AIL ASST Spedali Civili Brescia Italy

Dental Materials Science Discipline of Applied Oral Sciences Faculty of Dentistry The University of Hong Kong Hong Kong

Department of Clinical and Experimental Sciences University of Brescia Bone Marrow Transplant Unit ASST Spedali Civili Brescia Italy

Department of Clinical and Experimental Sciences University of Brescia Brescia Italy

Department of Information Engineering University of Brescia Brescia Italy

Department of Mechanical and Industrial Engineering University of Brescia Brescia Italy

Department of Medicine and Surgery Clinical Proteomics and Metabolomics Unit University of Milano Bicocca Vedano al Lambro Italy

Department of Orthodontics Dental School University of Brescia Brescia Italy

Department of Transfusion Medicine Laboratory for Stem Cells Manipulation and Cryopreservation ASST Spedali Civili Brescia Italy

Interdipartimental University Center of Research Adaptation and Regeneration of Tissues and Organs University of Brescia Brescia Italy

IRCCS Istituto Ortopedico Rizzoli Laboratorio di Immunoreumatologia e Rigenerazione Tissutale IRCCS Istituto Ortopedico Rizzoli Bologna Italy

Orthopedics and Traumatology Unit ASST Spedali Civili Brescia Italy

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