Effect of Expansive Force on Mesenchymal Stem Cells Isolated From the Mid-Palatal Suture of Mice
Cyclic Tensile Force
Understanding the mechanisms by which craniofacial sutures respond to mechanical force is essential for improving orthodontic treatment strategies. However, the innate ability to regenerate bone from calvarial stem cells is still unknown. Therefore, we have initiated a study to isolate cells from the mid-palatal suture of mice. The aim of our study is to evaluate mesenchymal stem cell (MSC) characteristics of cells isolated from the mid-palatal suture and their ability for osteogenic differentiation when subjected to cyclic tensile force in vitro. A total of 10, 6-week old male C57BL/6 mice were used to obtain mid-palatal suture cells. Cultured cells were evaluated for MSC markers using flow cytometry and their potential for multi-lineage differentiation was evaluated using Alizarin Red S , Oil Red O, and Toluidine Blue staining. Cultured cells were subjected to cyclic tensile force with 15% elongation and a frequency of 0.5 Hz for 2 hours on the Flexcell-FX5000 tension system. Both stretched and control cells were cultured in osteogenic medium and the effect of tensile force on osteogenic differentiation was evaluated using Western Blot analysis and Alizarin Red S staining. Our results showed that mid-palatal suture cells formed colony-forming-units (CFU-F), expressed MSC-markers CD73, CD90, CD105, and Sca-1, and were negative for hematopoietic markers CD34 and CD45. In addition, these cells showed multi-lineage differentiation to osteogenic, chondrogenic, and adipogenic cell lines. Western blot analysis showed an upregulation in expression of osteoblastic markers, including ALP, OCN, and RUNX2 in the stretch group compared to control which was confirmed by a marked increase in extracellular matrix deposit in the stretched group using Alizarin Red S staining. In summary, our findings show that cells isolated from the mid-palatal suture of mice have MSC characteristics in vitro and that cyclic mechanical tensile strain can promote osteogenic differentiation through an upregulation of osteogenic markers and an increase in production of mineralized matrix.