Fetal ovine skeletal and cardiac muscle transcriptomics are differentially altered by increased maternal cortisol during gestation

Abstract

We have previously found that in utero exposure to excess maternal cortisol (1 mg/kg/day) in late gestation increases the incidence of stillbirth during labor and produces fetal bradycardia at birth. In the interventricular septum, mitochondrial DNA (mt-DNA) was decreased, and transcriptomics and metabolomics were consistent with altered mitochondrial metabolism. The present study uses transcriptomics to model effects of increased maternal cortisol on fetal biceps femoris. Transcriptomic modeling revealed that pathways related to mitochondrial metabolism were downregulated, whereas pathways for regulation of reactive oxygen species and activation of the apoptotic cascade were upregulated. Mt-DNA and the protein levels of cytochrome C were significantly decreased in the biceps femoris. RT-PCR validation of the pathways confirmed a significant decrease in SLC2A4 mRNA levels and a significant increase in PDK4, TXNIP, ANGPTL4 mRNA levels, suggesting that insulin sensitivity of the biceps femoris muscle may be reduced in cortisol offspring. We also tested for changes in gene expression in diaphragm by rt-PCR. PDK4, TXNIP, and ANGPTL4 mRNA were also increased in the diaphragm, but SLC2A4, cytochrome C protein, and mt-DNA were unchanged. Comparison of the change in gene expression in biceps femoris to that in cardiac interventricular septum and left ventricle showed few common genes and little overlap in specific metabolic or signaling pathways, despite reduction in mt-DNA in both heart and biceps femoris. Our results suggest that glucocorticoid exposure alters expression of nuclear genes important to mitochondrial activity and oxidative stress in both cardiac and skeletal muscle tissues, but that these effects are tissue-specific.

Keywords: cortisol; fetus; heart; metabolism; mt-DNA; skeletal muscle.

Fig. 1.

Heat map of the differentially expressed genes (uncorrected P < 0.05) in biceps femoris of fetuses of control (C1–C6) or cortisol-treated (Cort1–Cort6) ewes. FF, female fetus; FL, female lamb; MF, male fetus; T, twin.

Fig. 2.

Heat map of the differentially expressed genes (uncorrected P < 0.05) in left ventricle of fetuses of control (C1–C5) or cortisol-treated (Cort1–Cort5) ewes. FF, female fetus; FL, female lamb; MF, male fetus.

Fig. 3.

Heat map of the differentially expressed genes (uncorrected P < 0.05) in interventricular septum of fetuses of control (C1–C8) or cortisol-treated (Cort1–Cort7) ewes. FF, female fetus; FL, female lamb; MF, male fetus; T, twin.

Fig. 4.

Venn diagram showing common differentially expressed genes (DEG) in biceps femoris (biceps), cardiac interventricular septum (IVS) and cardiac left ventricular (LV) free wall.

Fig. 5.

Expression of differentially expressed genes (DEG) as quantified by real-time quantitative PCR. A: biceps femoris; B: diaphragm. Mt-DNA copy number corrected for DNA for GADPH (a nuclear encoded gene) in biceps femoris (C) and diaphragm (D). *Genes that are significantly differently expressed, P < 0.05. Black bars, control group; gray bars, cortisol group. TXNIP, thioredoxin-interacting protein: SLC2A4, solute carrier family 2 (facilitated glucose transporter) 4; ANGPTL4, angiopoietin-like 4; BCL2, B cell lymphoma-2; SOCS3, suppressor of cytokine signaling 3; PDK4, pyruvate dehydrogenase kinase 4.