RNA-Binding Protein LIN28a Regulates New Myocyte Formation in the Heart Through Long Noncoding RNA-H19.

BACKGROUND: Developmental cardiac tissue holds remarkable capacity to regenerate after injury and consists of regenerative mononuclear diploid cardiomyocytes. On maturation, mononuclear diploid cardiomyocytes become binucleated or polyploid and exit the cell cycle. Cardiomyocyte metabolism undergoes a profound shift that coincides with cessation of regeneration in the postnatal heart. However, whether reprogramming metabolism promotes persistence of regenerative mononuclear diploid cardiomyocytes enhancing cardiac function and repair after injury is unknown. Here, we identify a novel role for RNA-binding protein LIN28a, a master regulator of cellular metabolism in cardiac repair after injury. METHODS: LIN28a overexpression was tested using mouse transgenesis on postnatal cardiomyocyte numbers, cell cycle, and response to apical resection injury. With the use of neonatal and adult cell culture systems and adult and Mosaic Analysis with Double Markers myocardial injury models in mice, the effect of LIN28a overexpression on cardiomyocyte cell cycle and metabolism was tested. Last, isolated adult cardiomyocytes from LIN28a and wild-type mice 4 days after myocardial injury were used for RNA-immunoprecipitation sequencing. RESULTS: LIN28a was found to be active primarily during cardiac development and rapidly decreases after birth. LIN28a reintroduction at postnatal day (P) 1, P3, P5, and P7 decreased maturation-associated polyploidization, nucleation, and cell size, enhancing cardiomyocyte cell cycle activity in LIN28a transgenic pups compared with wild-type littermates. Moreover, LIN28a overexpression extended cardiomyocyte cell cycle activity beyond P7 concurrent with increased cardiac function 30 days after apical resection. In the adult heart, LIN28a overexpression attenuated cardiomyocyte apoptosis, enhanced cell cycle activity, cardiac function, and survival in mice 12 weeks after myocardial infarction compared with wild-type littermate controls. Instead, LIN28a small molecule inhibitor attenuated the proreparative effects of LIN28a on the heart. Neonatal rat ventricular myocytes overexpressing LIN28a mechanistically showed increased glycolysis, ATP production, and levels of metabolic enzymes compared with control. LIN28a immunoprecipitation followed by RNA-immunoprecipitation sequencing in cardiomyocytes isolated from LIN28a-overexpressing hearts after injury identified long noncoding RNA-H19 as its most significantly altered target. Ablation of long noncoding RNA-H19 blunted LIN28a-induced enhancement on cardiomyocyte metabolism and cell cycle activity. CONCLUSIONS: Collectively, LIN28a reprograms cardiomyocyte metabolism and promotes persistence of mononuclear diploid cardiomyocytes in the injured heart, enhancing proreparative processes, thereby linking cardiomyocyte metabolism to regulation of ploidy/nucleation and repair in the heart.

UCP2 modulates cardiomyocyte cell cycle activity, acetyl-CoA, and histone acetylation in response to moderate hypoxia.

Developmental cardiac tissue is regenerative while operating under low oxygen. After birth, ambient oxygen is associated with cardiomyocyte cell cycle exit and regeneration. Likewise, cardiac metabolism undergoes a shift with cardiac maturation. Whether there are common regulators of cardiomyocyte cell cycle linking metabolism to oxygen tension remains unknown. The objective of the study is to determine whether mitochondrial UCP2 is a metabolic oxygen sensor regulating cardiomyocyte cell cycle. Neonatal rat ventricular myocytes (NRVMs) under moderate hypoxia showed increased cell cycle activity and UCP2 expression. NRVMs exhibited a metabolic shift toward glycolysis, reducing citrate synthase, mtDNA, mitochondrial membrane potential (ΔΨm), and DNA damage/oxidative stress, while loss of UCP2 reversed this phenotype. Next, WT and mice from a global UCP2-KO mouse line (UCP2KO) kept under hypoxia for 4 weeks showed significant decline in cardiac function that was more pronounced in UCP2KO animals. Cardiomyocyte cell cycle activity was reduced, while fibrosis and DNA damage was significantly increased in UCP2KO animals compared with WT under hypoxia. Mechanistically, UCP2 increased acetyl-CoA levels and histone acetylation, and it altered chromatin modifiers linking metabolism to cardiomyocyte cell cycle under hypoxia. Here, we show a potentially novel role for mitochondrial UCP2 as an oxygen sensor regulating cardiomyocyte cell cycle activity, acetyl-CoA levels, and histone acetylation in response to moderate hypoxia.