Mechanical Stress Helps to Maintain the Differentiation Potency of Mesenchymal Stem Cells

Objectives: Mesenchymal Stem Cells (MSCs) are pluripotent cells that can differentiate into osteoblasts, adipocytes or chondrocytes depending on the culture condition. Long-term culture of MSC is known to lose their differentiation potency. Development of new cell culture method to maintain their stemness is needed for successful application of MSCs in clinical dentistry.
Methods:
Mechanical stress is the physical stimulation that cells can sense as a result of the application of strain or deformation by external forces. Mechanotransduction is involved in the regulation of cellular physiological events. Previously, I reported that the expressions of Nanog, Oct4, and Sox2, which are crucial transcriptional factors for the stemness, were promoted with the mechanical stimulation by low-intensity pulsed ultrasound (LIPUS).
In this study, MSC isolated from subcutaneous adipose tissue of mice and osteo-progenitor cells derived from calvariae of newborn mice were cultured for 1~18 passages with or without the mechanical stimulation by LIPUS, for 120 min/day, and then induced to differentiate in osteogenic differentiation medium. Matrix mineralization was determined by alizarin red staining. The expressions of osteogenic and stemness marker genes were analyzed by real-time RT-PCR. Cell lysates were collected by PLC lysis buffer, and analyzed by western blotting.
Results:
While the cell calcification and mRNA levels of osteogenic marker genes were decreased in long-term cultured of osteo-progenitor cells, LIPUS stimulation maintained their differentiation potency during serial subculture. LIPUS stimulation also recovered the continuous subculture-induced reduction of Nanog, Oct4, and Msx2. Next, I found that spleen tyrosine kinase (Syk) was markedly phosphorylated with LIPUS stimulation in MSCs and osteoblasts. Syk expression levels were gradually decreased in osteogenic and adipogenic differentiation. Notably, pretreatment with a Syk-specific inhibitor significantly suppressed LIPUS-induced induction of Nanog.
Conclusions:
Taken together, our study suggests that mechanical stimulation using LIPUS has the possibility to maintain the differentiation potency of MSCs and osteo-progenitor cells. These LIPUS-induced effects are mediated by Syk and are specific for undifferentiated cells.