Blue laser irradiation enhances extracellular calcification of primary mesenchymal stem cells.
OBJECTIVE
Mesenchymal stem cells (MSCs) are multipotent cells present in adult bone marrow that replicate as undifferentiated cells and can differentiate to lineages of mesenchymal tissues. Homeostatic control of bone remodeling maintains bone mass by ensuring that bone resorption and bone formation occur sequentially and in a balanced manner. As most homeostatic functions occur in a circadian manner, a circadian clock could control bone mass. Here we show that laser irradiation can direct the extracellular calcification of mouse MSCs by altering the intracellular localization of the circadian rhythm protein cryptochrome 1 (CRY1).


METHODS
MSCs were irradiated with a blue laser (wavelength 405 nm) for 180 sec via a fiber attached to the bottom of the culture dish. After laser irradiation, the MSCs were incubated in osteogenic differentiation medium for 5 d. After laser irradiation, circadian rhythm protein CRY1 was immunostained and histochemical staining for extracellular calcification was observed.


RESULTS
Laser irradiation promoted extracellular calcification of MSCs, induced the translocation of CRY1 protein from the cytoplasm to the nucleus, and decreased CRY1 mRNA levels quantified by real-time PCR. Since the timing of nuclear accumulation of clock proteins constitutes an important step in the transcription-translation feedback loop driving the circadian core oscillator, laser irradiation could provide a simple and effective technology for clock protein localization and turnover. Our results also indicate that CRY1 is a master regulator of circadian rhythm that regulates the extracellular calcification of MSCs.


CONCLUSIONS
Laser irradiation could provide a simple and effective means of controlling the fate of MSCs as a therapeutic strategy, and act as a "molecular switch" of regulatory proteins by suppressing CRY transcription. Furthermore, this model system may be useful for exploring the cross-talk between circadian rhythm and cell function.
Kushibiki
Toshihiro
T
Frontier Research Base for Global Young Researchers, Frontier Research Center, Graduate School of Engineering, Osaka University, Osaka, Japan. kushibiki@see.eng.osaka-u.ac.jp
Awazu
Kunio
K
D000818:Animals; D002113:Calcification, Physiologic / Q000502:physiology*; D002478:Cells, Cultured; D002940:Circadian Rhythm / Q000502:physiology; D056931:Cryptochromes; D005420:Flavoproteins / Q000502:physiology*; D007834:Lasers; D008297:Male; D059630:Mesenchymal Stem Cells / Q000528:radiation effects*; D051379:Mice; D008807:Mice, Inbred BALB C; D010012:Osteogenesis; D016133:Polymerase Chain Reaction
C538964:Cry1 protein, mouse; D056931:Cryptochromes; D005420:Flavoproteins