Effects of non-esterified fatty acids on the gluconeogenesis in bovine hepatocytes

Abstract

Dairy cows experience an increased demand for glucose to support milk production. However, negative energy balance is a common condition in peripartum cows. In response, fat mobilization provides non-esterified fatty acids (NEFAs) for oxidation in the liver to generate ATP. To investigate the effects of NEFAs on gluconeogenesis, the expression and enzyme activity of pyruvate carboxylase (PC) and phosphoenolpyruvate carboxylase (PEPCK) in cultured bovine hepatocytes were evaluated by quantitative polymerase chain reaction and spectrophotometry, respectively. The results showed that PC and PEPCK mRNA levels were marked decreased when the NEFAs concentrations exceeded 0.5 and 1.5 mmol/l, respectively. The PC and PEPCK enzyme activity showed significantly decreased when the NEFAs concentrations exceeded 1.5 and 0.5 mmol/l, respectively. These findings indicate that high circulating levels of NEFAs inhibit hepatocyte gluconeogenesis, thereby promoting negative energy balance.

Effect of Non-Esterified Fatty Acids on Fatty Acid Metabolism-Related Genes in Calf Hepatocytes Culturedin Vitro

“…NEFA are mainly oxidised in the liver to generate ATP for cows. In addition to a metabolic substrate, NEFA also act as signalling molecules involved in many metabolic processes, such as glucose metabolism and insulin resistance [8,11,12]. However, the effect of NEFA on fatty acid metabolism in dairy cows was unclear, especially at high NEFA concentrations.…”

“…Hepatocytes were cultured using the collagenase perfusion method described previously [8,9]. Briefly, the liver (caudate lobe) was obtained under sterile conditions from a Holstein newborn calf anesthetised with thiamylal sodium.…”

“…The preparation of the NEFA solution was performed as described previously by Li et al [8]. Oleic acid (2.175 mmol), linoleic acid (0.245 mmol), palmitic acid (1.595 mmol), stearic acid (0.72 mmol), and palmitoleic acid (0.265 mmol) were dissolved in 100 mL of a potassium hydroxide solution (1 mol/L) at 60°C.…”

SREBP-1c Gene Silencing can Decrease Lipid Deposits in Bovine Hepatocytes Cultured <b><i>in Vitro</i></b>

“…The caudate lobe of the liver was obtained through liver excision by surgery on a newborn calf. Hepatocytes were isolated by a modified two-step collagenase perfusion method established by our laboratory [8]. The hepatocytes density was adjusted to 2×10 6 cells/mL with adherent culture medium.…”

Increase of Fatty Acid Oxidation and VLDL Assembly and Secretion Overexpression of PTEN in Cultured Hepatocytes of Newborn Calf

“…The calf was nursed care, and recovered finally. Calf hepatocytes were isolated by a modified two-step collagenase perfusion method established by our laboratory [20]. The cells were maintained in culture in a humidified atmosphere of 5% CO 2 at 37°C.…”