Abstract 15822: Novel Method ‘Fucci’ Elucidated the Cardiomyocyte Cell Cycle Dynamics in Various Life Stages
Background: Although accumulating evidences have shown that cardiomyocyte have the potential of cell cycle progression, their experimental methods relied chiefly on unphysiological procedures, such as immunostaining for cell cycle markers and nucleotide analogous incorporation. Against this background, we established a novel technique to visualize the cell cycle. This novel method named ‘Fucci’ (the fluorescent ubiquitination-based cell cycle indicator), effectively labeled individual G1 phase nuclei red, S/G2/M phase green and G1 /S transition phase yellow in living cells, by interlocking fluorescent probes with antiphase oscillating proteins that mark cell cycle transitions. Therefore, the purpose of this study was to reevaluate the cardiomyocyte cell cycle dynamics in various life stages, by using our novel method Fucci.
Methods and results: We evaluated the cell cycle dynamics of Fucci transgenic mouse in each life stage. Interestingly, immunohistochemistry for cardiac markers GATA4 and α-actinin revealed that the embryonic stage has a relatively lower population of S/G2/M phase cardiomyocyte (16.6%-27.1% at E14.5), compared to the early neonatal stage (11.0%-84.4% at P1). Subsequent analysis showed that the S/G2/M phase cardiomyocyte population once drops after birth, around post natal day 3(14.6%-16.9% at P3), and again reaches a peak in post natal day 5 (27.3%-50.6% at P5). To confirm that the Fucci system reflects the cell cycle progression in cardiomyocyte, we performed live imaging of a cardiac slice culture from E14.5 Fucci transgenic mouse. Time-lapse imaging of cardiac slice culture showed green and red fluorescent colors brighten up and disappear temporally, which indicated that the Fucci system reflects cell cycle progression in cardiomyocyte.
Conclusions: In conclusion, our data based on novel method Fucci revealed that embryonic stage has a relatively lower population of S/G2/M phase cardiomyocyte compared to the early neonatal stage. Then the population of S/G2/M phase cardiomyocyte once drops around P3 and reascends in P5. Live imaging demonstrated that Fucci system functions in cardiomyocyte. Further analysis based on this novel method will give us a fresh dimension to the research in cardiomyocyte cell cycle.
- © 2010 by American Heart Association, Inc.