Time-lapse imaging of cell cycle dynamics during development in living cardiomyocyte.
Mammalian cardiomyocytes withdraw from the cell cycle shortly after birth, although it remains unclear how cardiomyocyte cell cycles behave during development. Compared to conventional immunohistochemistry in static observation, time-lapse imaging can reveal comprehensive data in hard-to-understand biological phenomenon. However, there are no reports of an established protocol of successful time-lapse imaging in mammalian heart. Thus, it is valuable to establish a time-lapse imaging system to enable the observation of cell cycle dynamics in living murine cardiomyocytes. This study sought to establish time-lapse imaging of murine heart to study cardiomyocyte cell cycle behavior. The Fucci (fluorescent ubiquitination-based cell cycle indicator) system can effectively label individual G1, S/G2/M, and G1/S-transition phase nuclei red, green and yellow, respectively, in living mammalian cells, and could therefore be useful to visualize the real-time cell cycle transitions in living murine heart. To establish a similar system for time-lapse imaging of murine heart, we first developed an ex vivo culture system, with the culture conditions determined in terms of sample state, serum concentration, and oxygen concentration. The optimal condition (slice culture, oxygen concentration 20%, serum concentration 10%) successfully mimicked physiological cardiomyocyte proliferation in vivo. Time-lapse imaging of cardiac slices from E11.5, E14.5, E18.5, and P1 Fucci-expressing transgenic mice revealed an elongated S/G2/M phase in cardiomyocytes during development. Our time-lapse imaging of murine heart revealed a gradual elongation of the S/G2/M phase during development in living cardiomyocytes.
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Hisayuki
H
Hashimoto
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan. Electronic address: yuasa@a8.keio.jp.
Shinsuke
S
Yuasa
Department of Anatomy, Keio University School of Medicine, Tokyo 160-8582, Japan.
Hidenori
H
Tabata
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Shugo
S
Tohyama
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Nozomi
N
Hayashiji
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Fumiyuki
F
Hattori
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Naoto
N
Muraoka
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Toru
T
Egashira
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Shinichiro
S
Okata
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Kojiro
K
Yae
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Tomohisa
T
Seki
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Takahiko
T
Nishiyama
Department of Anatomy, Keio University School of Medicine, Tokyo 160-8582, Japan.
Kazunori
K
Nakajima
Life Function and Dynamics, ERATO, JST, 2-1 Hirosawa Wako-city, Saitama 351-0198, Japan; Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-city, Saitama 351-0198, Japan.
Asako
A
Sakaue-Sawano
Life Function and Dynamics, ERATO, JST, 2-1 Hirosawa Wako-city, Saitama 351-0198, Japan; Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-city, Saitama 351-0198, Japan.
Atsushi
A
Miyawaki
Department of Cardiology, Keio University School of Medicine, Tokyo 160-8582, Japan.
Keiichi
K
Fukuda
D000818:Animals; D002453:Cell Cycle / Q000502:physiology*; D049109:Cell Proliferation; D004622:Embryo, Mammalian; D047108:Embryonic Development / Q000502:physiology*; D005260:Female; D005456:Fluorescent Dyes; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence / Q000379:methods; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000502:physiology; D011247:Pregnancy; D059008:Time-Lapse Imaging; D046509:Tissue Culture Techniques; D054875:Ubiquitination
D005456:Fluorescent Dyes
Cardiac development; Cardiac regeneration; Cell culture; Cell cycle; Imaging