Osteogenic Differentiation Effect of Human Periodontal Ligament Stem-Cell Initial Cell Density on Autologous Cells and Human Bone Marrow Stromal Cells

Stem cells have differentiation and regulation functions. Here, we discussed the impact of cell culture density on stem cell proliferation, osteoblastogenesis, and regulation. To discuss the effect of the initial culture density of human periodontal ligament stem cells (hPDLSCs) on the osteogenic differentiation of autologous cells, we found that the hPDLSC proliferation rate decreased with an increase in the initial plating density (0.5-8 x 10(4) cells/cm(2)) for the 48 h culture cycle. After hPDLSCs induced osteogenic differentiation for 14 days with different initial cell culture densities, the expression of osteoprotegerin (OPG) and runt-related transcription factor 2(RUNX2) and the OPG/ Receptor Activator of Nuclear Factor-kappa B Ligand (RANKL) ratio were the highest in the hPDLSCs initially plated at a density of 2 x 10(4) cells/cm(2), and the average cell calcium concentration was also the highest. To study hPDLSCs regulating the osteoblastic differentiation of other cells, we used 50 mu g/mL of secreted exosomes derived from hPDLSCs cultured using different initial cell densities to induce human bone marrow stromal cell (hBMSC) osteogenesis. After 14 days, the results indicated that the gene expression of OPG, Osteocalcin(OCN,)RUNX2, and osterix and the OPG/RANKL ratio were the highest in the 2 x 10(4) cells/cm(2) initial cell density group, and the average calcium concentration was also the highest. This provides a new idea for the clinical application of stem cell osteogenesis.

Automated summary

This study investigated the impact of cell culture density on stem cell proliferation, osteoblastogenesis, and regulation. The results showed that the proliferation rate of human periodontal ligament stem cells (hPDLSCs) decreased with an increase in initial plating density. After osteogenic differentiation, hPDLSCs initially plated at a density of 2 x 10(4) cells/cm(2) showed the highest expression of OPG and RUNX2, as well as the highest OPG/RANKL ratio and cell calcium concentration. Furthermore, exosomes derived from hPDLSCs cultured at different initial cell densities were found to regulate osteoblastic differentiation of human bone marrow stromal cells (hBMSC), with the 2 x 10(4) cells/cm(2) initial cell density group showing the highest expression of OPG, OCN, RUNX2, and osterix, as well as the highest average calcium concentration. This provides a new perspective for the clinical application of stem cell osteogenesis.