Comparison of chondrogenic differentiation of mesenchymal stromal cells from human amniotic fluid and human adipose-derived tissue in chitosan-xanthan gum scaffolds

Articular cartilage has a low regenerative capacity, and when affected by injuries, it can suffer permanent damage, potentially progressing to diseases such as osteoarthritis (OA). Current treatments for these injuries are palliative. The use of stem cells (SCs) with their proliferative and differentiation capabilities, with or without scaffolds to induce chondrogenesis and promote tissue repair, is under investigation. This study compared the chondrogenesis promoted by human amniotic fluid SCs (AF-hSCs) with that promoted by human adipose tissue SCs (AT-hSCs) cultured in chitosan/xanthan (CX) scaffolds and stimulated by TGF-β3. AF-hSC samples were obtained from women in the second trimester of pregnancy who, for obstetric reasons, underwent amniocentesis. AT-hSC samples were obtained from patients who underwent liposuction through plastic surgery. For AF-hSCs, cells positive for CD117 (ckit) were selected, and SCs from both sources followed the same cell expansion steps, with characterization based on adherence, specific markers in flow cytometry, and differentiation into three mesenchymal lineages: adipogenesis, chondrogenesis, and osteogenesis. SCs were placed in CX scaffolds and cultured under TGF-β3 stimulation for 21 days. To confirm and assess chondrogenesis, scanning electron microscopy (SEM), histology, immunohistochemistry, and immunofluorescence were performed. SCs from both sources demonstrated rapid and high expansion capacity, adherent culture with fibroblast-like morphology, positivity for mesenchymal lineage markers, pluripotency, chondrogenic potential, and negativity for hematopoietic markers. Differentiation into adipogenic, chondrogenic, and osteogenic mesenchymal lineages was also observed. Chondrogenesis was confirmed by histological staining with Hematoxylin and Eosin, Alcian Blue, Picrosirius Red, and Masson's Trichrome, indicating matrix production, and presence of collagen and proteoglycans. Immunohistochemical analysis showed positive labeling for collagen II and aggrecan, as well as in immunofluorescence. SEM analysis revealed an intense collagen fiber network in the scaffold. Cells from both sources proved effective in inducing chondrogenesis.