Possible role of corticosterone in proteolysis, glycolytic, and amino acid metabolism in primary cultured avian myotubes incubated at high-temperature conditions

Abstract

Excess glucocorticoid secretion induces oxidative damage and muscle proteolysis and modulates glucose and lipid metabolism. It is known that the high-temperature (HT) treatment enhances corticosterone (CORT) secretion, muscle proteolysis, and mitochondrial reactive oxygen species (mtROS) generation in chickens. The present study investigated the co-effects of CORT on proteolysis and mtROS production, together with glucose, fatty acid, and amino acid metabolism in HT-treated cells. Myoblast cells were isolated from the major pectoralis muscle of five 0- or 1-day-old neonatal chicks and were precultured at 37°C/CO₂ conditions for 48 h to reach subconfluent (80%-90%) conditions. Cells were then reseeded onto a 6- or 24-well microplate for the subsequent measurement, followed by the culture under a control temperature (37°C, control) or HT (41°C) conditions for 1 or 6 h. The HT-treated cells were cocultured with physiologically relevant concentrations of CORT (20 ng/mL in dimethyl sulfoxide). The HT treatment decreased cellular protein content (P < 0.05) and increased atrogin-1 mRNA levels and mtROS generation levels compared to the control group (P < 0.05), whereas HT/CORT co-treatment did not induce changes in either parameter. The mRNA level of glucose transporter-1 was decreased in HT-treated cells compared to that in normal cells (P < 0.05), and the decrease was increased in the CORT co-treatment (P < 0.05). While HT treatment did not alter pyruvate dehydrogenase kinase-4 mRNA level, the level was increased in the CORT co-treatment compared to the control and HT-treated cells (P < 0.05). Neither HT nor HT/CORT treatments altered the mRNA levels of fatty acid oxidation-related factors, carnitine palmitoyl transferase-1, and cluster of differentiation 36. The study conducted a metabolic analysis using gas chromatography-mass spectrometry. The results showed that HT/CORT-treated cells had decreased intracellular citrate and α-ketoglutarate levels (P < 0.05) and increased extracellular alanine and amino acid that have gluconeogenic properties, as well as increased aspartate, isoleucine, serine, methionine, and threonine levels (P < 0.05) compared to HT-treated cells. These results suggest that CORT may not affect proteolysis and mtROS production but can suppress pyruvate oxidation and promote alanine production in HT-treated chickens.

Keywords: Atrogin-1; FoxO3; Glucose-alanine cycle; PDK4.