The impact of microstructure and extracellular matrix suspension on the proliferation of bone marrow-derived mesenchymal stem cells for osteochondral defect repair

Osteochondral defects are a challenge in orthopaedic surgery due to the complexity and function of cartilage. Within this scenario, this study aimed to develop/characterize bioactive porous supports based on oxidized polyvinyl alcohol (OxPVA), with/without human cartilage-derived decellularized ECM (dECM), as platforms for HM1-SV40 cell adhesion and proliferation. OxPVA scaffolds were fabricated using a particle-leaching technique (gelatin concentrations: 10%, 15% and 25% w/w); Scanning Electron Microscopy (SEM) was used to examine the ultrastructure, and a morphometric study assessed pores number, size and porosity percentage. Fluorescence Recovery after Photobleaching (FRAP) was used to evaluate the interconnectivity of the scaffold pores. To enhance the bioactivity of OxPVA, dECM (25% w/w) was incorporated into the scaffolds; thus, the expression of genes related to collagen synthesis and cartilage differentiation/remodelling in seeded HM1-SV40 cells was analyzed by quantitative PCR; relative protein expression levels of SOX9, ACAN and COMP were also assessed. Composite scaffolds biocompatibility was proved by subcutaneous implantation in Sprague-Dawley. As for bone, 3D-printed polylactic acid (PLA)-based scaffolds with varying geometries (67%, 53% and 40% porosity; 600-1400 mu m pores size) were fabricated and tested in vitro. Lower gelatin concentrations led to numerous superficial pores, whereas higher concentrations produced larger, coalescing ones. HM1-SV40 cells showed better adhesion to scaffolds prepared with 25% gelatin. The OxPVA+dECM scaffolds exhibited a homogeneous matrix distribution, further promoting cell interaction, with a reduction in mean pore size versus matrix-free scaffolds. Moreover, OxPVA supports prepared with 25% gelatin + dECM provided a favorable environment supporting chondrogenic differentiation and cartilage matrix deposition. No inflammatory response to the implants was observed in vivo. All PLA supports showed good cell viability; SEM higlighted full-thickness HM1-SV40 cell distribution on and within PLA scaffolds, indicating complete colonization. Further studies are needed to evaluate stem cell differentiation, but bioactive OxPVA and 3D-printed PLA scaffolds show potential for osteochondral regeneration.