4-phenylbutyrate exerts stage-specific effects on cardiac differentiation via HDAC inhibition.

4-phenylbutyrate (4-PBA), a terminal aromatic substituted fatty acid, is used widely to specifically attenuate endoplasmic reticulum (ER) stress and inhibit histone deacetylases (HDACs). In this study, we investigated the effect of 4-PBA on cardiac differentiation of mouse embryonic stem (ES) cells. Herein, we found that 4-PBA regulated cardiac differentiation in a stage-specific manner just like trichostatin A (TSA), a well-known HDAC inhibitor. 4-PBA and TSA favored the early-stage differentiation, but inhibited the late-stage cardiac differentiation via acetylation. Mechanistic studies suggested that HDACs exhibited a temporal expression profiling during cardiomyogenesis. Hdac1 expression underwent a decrease at the early stage, while was upregulated at the late stage of cardiac induction. During the early stage of cardiac differentiation, acetylation favored the induction of Isl1 and Nkx2.5, two transcription factors of cardiac progenitors. During the late stage, histone acetylation induced by 4-PBA or TSA interrupted the gene silence of Oct4, a key determinant of self-renewal and pluripotency. Thereby, 4-PBA and TSA at the late stage hindered the exit from pluripotency, and attenuated the expression of cardiac-specific contractile proteins. Overexpression of HDAC1 and p300 exerted different effects at the distinct stages of cardiac induction. Collectively, our study shows that timely manipulation of HDACs exhibits distinct effects on cardiac differentiation. And the context-dependent effects of HDAC inhibitors depend on cell differentiation states marked by the temporal expression of pluripotency-associated genes.

Wnt3a upregulation is involved in TGFß1-induced cardiac hypertrophy.

Pathological cardiac hypertrophy, characterized by enlarged cell size and fetal gene reactivation, ultimately leads to cardiac dysfunction and heart failure. The expression of transforming growth factor beta 1 (TGFß1) is often elevated in experimental models of cardiac hypertrophy. In the present study, we observed the activation of Wnt/ß-catenin signaling in TGFß1-induced cardiac hypertrophy. TGFß1 stimulation decreased the phosphorylation levels of ß-catenin and triggered the nuclear accumulation of ß-catenin. In turn, TGFß1 enhanced the expression of c-Myc, which is a transcriptional target of canonical Wnt/ß-catenin pathway. Knockdown of ß-catenin completely blocked TGFß1-induced c-Myc upregulation. Wnt3a is an important Wnt ligand associated with cardiac fibrosis and hypertrophy. Further investigation revealed that TGFß1 can upregulate Wnt3a expression in an ALK5-Smad2/3-dependent manner. A consensus Smad binding sequence is located within the Wnt3a promoter, and TGFß1 stimulation enhanced recruitment of Smad2/3 onto the Wnt3a promoter. Meanwhile, Wnt3a overexpression also stimulated TGFß1 expression. Chemical inhibition of Wnt/ß-catenin signaling partially attenuated TGFß1-induced hypertrophic responses. These findings suggest crosstalk between TGFß1 and canonical Wnt/ß-catenin pathways in cardiac hypertrophy.