Regulation of Cardiomyocyte Proliferation and Myocardial Growth During Development by FOXO Transcription Factors

Abstract

Abstract-Cardiomyocytes actively proliferate during embryogenesis and withdraw from the cell cycle during neonatal stages. FOXO (Forkhead O) transcription factors are a direct target of phosphatidylinositol-3 kinase/AKT signaling in skeletal and smooth muscle and regulate expression of the Cip/Kip family of cyclin kinase inhibitors in other cell types; however, the interaction of phosphatidylinositol-3 kinase/AKT signaling, FOXO transcription factors, and cyclin kinase inhibitor expression has not been reported for the developing heart. Here, we show that FOXO1 and FOXO3 are expressed in the developing myocardium concomitant with increased cyclin kinase inhibitor expression from embryonic to neonatal stages. Cell culture studies show that embryonic cardiomyocytes are responsive to insulin-like growth factor 1 stimulation, which results in the induction of the phosphatidylinositol-3 kinase/AKT pathway, cytoplasmic localization of FOXO proteins, and increased myocyte proliferation. Likewise, adenoviral-mediated expression of AKT promotes cardiomyocyte proliferation and cytoplasmic localization of FOXO. In contrast, increased expression of FOXO1 negatively affects myocyte proliferation. In vivo myocyte-specific transgenic expression of FOXO1 during heart development causes embryonic lethality at embryonic day 10.5 because of severe myocardial defects that coincide with premature activation of p21 cip1 , p27 kip1 , and p57 kip2 and decreased myocyte proliferation. Transgenic expression of dominant negative FOXO1 in cardiomyocytes does not obviously affect heart development at embryonic day 10.5, but results in abnormal morphology of the myocardium by embryonic day 18.5 along with decreased cyclin kinase inhibitor expression and increased myocyte proliferation. These data support FOXO transcription factors as negative regulators of cardiomyocyte proliferation and promoters of neonatal cell cycle withdrawal during heart development. (Circ Res. 2008;102:686-694.)Key Words: heart development Ⅲ FOXO1 (FKHR) Ⅲ Cip/Kip cyclin kinase inhibitors N ormal heart morphogenesis and development are dependent on highly controlled differential regulation of cell proliferation in specific populations of cardiomyocytes during embryonic, fetal, and neonatal stages. 1 Embryonic cardiomyocytes throughout the primitive heart tube rapidly proliferate to provide sufficient cell numbers to build the working myocardium. 2 At fetal stages, proper formation of the ventricular trabeculae, compact zone, and interventricular septum is dependent on more localized temporal and spatial regulation of cardiomyocyte proliferation. 3,4 Immediately before birth, cardiomyocytes throughout the myocardium undergo a hyperplastic to hypertrophic transition in which cell division slows and cell growth increases. 5 After birth, neonatal cardiomyocytes withdraw from the cell cycle, and growth of the myocardium into adulthood occurs primarily by hypertrophy. 1 The molecular mechanisms that control cardiomyocyte maturation, proliferation, and resultant myocardia...

FoxO Transcription Factors Promote Autophagy in Cardiomyocytes

“…In some experiments, the inhibitor of autophagosome formation, 3-methyladenine (3MA, 10 M), was added to determine the requirement for autophagy in starvation-induced cardiomyocyte cell size reduction. Adenoviral infections were performed as described previously (27) with wild-type (WT), constitutively active (CA) FoxO1 or dominant negative FoxO1 (⌬256) obtained from D. Accili (35) and wild-type (WT) or CA-FoxO3 obtained from K. Walsh (26). Infection with cytomegalovirus ␤-galactosidase virus was used as a control for all experiments.…”

“…The FoxO subfamily of transcription factors consists of FoxO1, FoxO3, and FoxO4, which are subject to inhibition by growth factors, including insulin and insulin-like growth factor-1 (25). During growth factor limitation, the inhibition of FoxO activity by phosphatidylinositol 3-kinase/AKT is relieved, and the dephosphorylated FoxO transcription factors are activated and become localized to the nucleus (23,26,27). Sirt1, a mammalian ortholog of NAD ϩ -dependent protein deacetylase, Sir2, promotes the activity of FoxO transcription factors by deacety-lation under conditions of oxidative stress (24).…”

“…Sirt1, a mammalian ortholog of NAD ϩ -dependent protein deacetylase, Sir2, promotes the activity of FoxO transcription factors by deacety-lation under conditions of oxidative stress (24). FoxO target genes are involved in atrophy in skeletal and cardiac myocytes (26,28), autophagy in skeletal myocytes (29,30), differentiation in skeletal and smooth muscle (31), or inhibition of cell cycle progression in neonatal cardiomyocytes (27). FoxO1 null mice die at E10.5 from vascular defects, and FoxO3 null mice develop cardiac hypertrophy as adults, suggesting a possible role for FoxO3 in regulating cardiomyocyte cell size (32,33).…”

Activation of AMPK inhibits cardiomyocyte hypertrophy by modulating of the FOXO1/MuRF1 signaling pathway in vitro

“…The Forkhead box O (FOXO) family of transcription factors plays an important role in cellular metabolism, differentiation, and muscle atrophy in mammals [14,15] . The FOXO1 transcription factor has been suggested to directly modulate the expression of the MuRF1 protein during muscle atrophy [16,17] .…”

“…The FOXO1 transcription factor has been suggested to directly modulate the expression of the MuRF1 protein during muscle atrophy [16,17] . Recent studies have indicated that FOXO1 can suppress cardiomyocyte proliferation and blunt cardiac hypertrophy [15,18] . Thus, atrophy-related FOXO1/MuRF1 pathways may serve as a target for reversing remodeling of the hypertrophied heart.…”