Large-Scale Production of Secretome from Human Dental Pulp Stem Cells for Articular Cartilage Regeneration

Introduction:Articular cartilage degeneration can lead to a progressive loss of joint function and, eventually, total joint failure. The use of mesenchymal stem cells (MSCs) for cartilage regeneration has proven effective; however, the difficulty of their clinical application has prompted research into other strategies, such as using MSC secretome instead of cells themselves. Our aim was to generate a culture system to obtain large amounts of MSC secretome with a controlled and known composition and the potential to induce articular cartilage regeneration. Methods:Human dental pulp stem cells (hDPSCs) embedded in alginate/agarose beads were cultured in a bioreactor with constant agitation and periodic culture medium exchange. The secretome was collected and characterized according to known chondral markers (TGF-β1, SERPINE1, miR-140, miR-675, miR-23a, miR-204, miR-211, and miR-337-5p). Metabolomic and proteomic analyses were carried out for detailed characterization of the collected secretome. The chondrogenic potential of the collected secretome was tested in an in vitro model developed using human primary chondrocytes cultured in 3D on alginate-agarose scaffolds. Results:Our system allowed us to obtain 1200 mL of secretome from intensive cultures of hDPSCs. The cells cultured on the platform remained viable and in an active anabolic state, secreting large amounts of prochondrogenic mediators. Variability between batches of secretome was low, and the collected secretome induced primary chondrocyte differentiation in vitro. Conclusions:Genomic, metabolomic, and proteomic data indicate that hDPSCs on the platform proliferate and acquire a chondrocyte-like phenotype. These cells secrete mediators that define a microenvironment favorable for chondral regeneration. This is further supported by evidence found in the in vitro differentiation model. Our platform allows the production of large volumes of secretome with controlled composition and chondrogenic induction potential. This is a necessary preclinical study for the subsequent analysis of the secretome obtained using in vivo experimental models.