Consuming a Western diet for two weeks suppresses fetal genes in mouse hearts

Abstract

Diets high in sugar and saturated fat (Western diet) contribute to obesity and pathophysiology of metabolic syndrome. A common physiological response to obesity is hypertension, which induces cardiac remodeling and hypertrophy. Hypertrophy is regulated at the level of chromatin by repressor element 1-silencing transcription factor (REST), and pathological hypertrophy is associated with reexpression of a fetal cardiac gene program. Reactivation of fetal genes is commonly observed in hypertension-induced hypertrophy; however, this response is blunted in diabetic hearts, partially due to upregulation of the posttranslational modification O-linked-β-N-acetylglucosamine (O-GlcNAc) to proteins by O-GlcNAc transferase (OGT). OGT and O-GlcNAc are found in chromatin-modifying complexes, but it is unknown whether they play a role in Western diet-induced hypertrophic remodeling. Therefore, we investigated the interactions between O-GlcNAc, OGT, and the fetal gene-regulating transcription factor complex REST/mammalian switch-independent 3A/histone deacetylase (HDAC). Five-week-old male C57BL/6 mice were fed a Western (n = 12) or control diet (n = 12) for 2 wk to examine the early hypertrophic response. Western diet-fed mice exhibited fasting hyperglycemia and increased body weight (P < 0.05). As expected for this short duration of feeding, cardiac hypertrophy was not yet evident. We found that REST is O-GlcNAcylated and physically interacts with OGT in mouse hearts. Western blot analysis showed that HDAC protein levels were not different between groups; however, relative to controls, Western diet hearts showed increased REST and decreased ANP and skeletal α-actin. Transcript levels of HDAC2 and cardiac α-actin were decreased in Western diet hearts. These data suggest that REST coordinates regulation of diet-induced hypertrophy at the level of chromatin.

Keywords: O-GlcNAc; cardiac hypertrophy; chromatin; fetal genes.

Fig. 1.

A: effects of the Western (WES) diet on markers of cardiac hypertrophy compared with control (CON) hearts. Heart weight to tibia length (HW/TL), cardiomyocyte cross-sectional area (CSA), and representative images of cardiomyocyte size. Scale bar: 20 μm. Background and contrast were adjusted using ImageJ software (National Institutes of Health, Bethesda, MD). B: left ventricular homogenates from WES-fed mice exhibited higher protein levels of repressor element 1-silencing transcription factor (REST), α-sk Actin, and ANP vs. CON. Calsequestrin was the internal loading control. n = 6 per group except for CSA; n = 3 per group, *P < 0.05 vs. CON.

Fig. 2.

Nuclear and cytosolic distribution of O-GlcNAc transferase (OGT), O-GlcNAcase (OGA), and O-GlcNAc. Densitometric analysis of cytosolic and nuclear proteins are presented as relative to GAPDH and TATA-BP, respectively. n = 6 per group. *P < 0.05.

Fig. 3.

qRT-PCR showing that the Western diet induced increased mRNA levels of cardiac α-actin and histone deacetylase 2 (HDAC2) but no other genes within the O-GlcNAc pathway or REST complex. n = 5 per group, *P < 0.05; = 0.08.

Fig. 4.

Coimmunoprecipitation (IP) of O-GlcNAc (A) and OGT (B) followed by immunoblots targeting protein involved in the O-GlcNAc pathway and REST complex. The immunoblots presented are representative images from a total of n = 6 per group. C, Control; W, Western.

Fig. 5.

Coimmunoprecipitation of mSin3A (A) and REST (B), immunoblotted for mSin3A, REST, OGT, O-GlcNAc, HDAC1, HDAC2, HDAC4, and HDAC5. The position from which additional lanes were removed is indicated by the lined box and white spaces; immunoblots presented are representative images from a total of n = 6 per group.

Fig. 6.

H9c2 cardiomyoblasts were subjected to treatments that increase protein O-GlcNAcylation. Glu, glucose; GlcN, glucosamine; TG, Thiamet-G. n = 3 per group.