Noninvasive estimation of cell cycle phase and proliferation rate of human mesenchymal stem cells by phase-shifting laser microscopy

The simultaneous determination of the cell cycle phase of individual adherent mesenchymal stem cells (MSCs) using a fluorescence microscope after staining with 4′,6-diamidine-2′-phenylindole dihydrochloride and bromodeoxyuridine and the laser phase shift by phase-shifting laser microscopy (PLM) revealed that the laser phase shift of cells in the G₂/M phase was markedly higher than that of cells in the G₀/G₁ phase. Even in the synchronous cultures to G₀/G₁ and G₂/M cell cycle phases, the laser phase shift of the cells in the G₂/M phase was markedly higher than that of the cells in the G₀/G₁ phase. The analysis of the cultures of MSCs from different donors with the addition of FGF2 at different concentrations revealed that there was a marked negative correlation between the average phase shift and mean generation time. In conclusion, it is possible to estimate noninvasively the proliferation activity of MSCs population by measuring the phase shift using PLM.