Evaluation of periodontal ligament stem cell viability and proliferation on native collagen membranes: An in vitro study

Background: Periodontal ligament stem cells (PDLSCs) are mesenchymal stem cells with potential for periodontal regeneration. Native collagen membranes are widely used scaffolds due to their biocompatibility and support for cell growth. This study aimed to evaluate the viability, proliferation, and characterization of PDLSCs cultured on native collagen membranes. Materials and Methods: Human PDLSCs were isolated from healthy premolars and cultured using explant techniques. Morphology was assessed using phase-contrast microscopy, while viability and population doubling time were measured using trypan blue exclusion. Clonogenic potential was evaluated using a colony-forming unit assay. Immunophenotyping was performed by flow cytometry for mesenchymal markers (CD73, CD90, and CD105) and negative hematopoietic markers (CD34 and CD45). Multilineage differentiation was tested using osteogenic and adipogenic media. The biocompatibility of collagen membrane eluates and scaffolds was evaluated using water-soluble tetrazolium salt-1 and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. Results: Four PDLSC lines exhibited high viability (>99%) and retained fibroblast-like morphology. Flow cytometry confirmed mesenchymal marker expression. Cells showed successful osteogenic and adipogenic differentiation. Both direct seeding and eluate exposure from collagen membranes supported PDLSC viability and proliferation, particularly at 24-h eluate intervals. Conclusion: Native collagen membranes are biocompatible scaffolds that support PDLSC proliferation, offering promise for future periodontal regenerative therapies.