Short- and longer-term regulation of very-low-density lipoprotein secretion by insulin, dexamethasone and lipogenic substrates in cultured hepatocytes. A biphasic effect of insulin

Abstract

1. The precise effects of insulin, dexamethasone and lipogenic precursors on the secretion of very-low-density lipoprotein (VLDL) cholesterol and triacylglycerol were dependent on the age of the culture and the duration of treatment. 2. The rates of secretion of triacylglycerol and cholesterol gradually declined with the age of the culture, although there was no detectable decrease within a given 24 h period. 3. Between 4 h and 24 h after cell preparation, insulin inhibited VLDL secretion. Inhibition was maximal between 6 and 12 h after addition of insulin. Longer-term treatment (24-48 h) with insulin resulted in a stimulation of VLDL secretion. This effect was less apparent when dexamethasone was simultaneously present. The secretion of triacylglycerol and cholesteryl ester was more sensitive to insulin than was that of non-esterified cholesterol. 4. Dexamethasone alone stimulated the secretion of VLDL to an extent which increased with the age of the culture. In young cultures (up to 24 h old) dexamethasone protected against inhibition by insulin, but was ineffective in older cultures. 5. In young cultures the stimulatory effect of lipogenic precursors (lactate and pyruvate) on the secretion of triacylglycerol and cholesterol was more pronounced in the presence of dexamethasone. In cultures older than 24 h, the secretion of these components was less sensitive to short-term stimulation by lactate and pyruvate.

Hepatic secretion of very-low-density lipoprotein apolipoprotein B-100 studied with a stable isotope technique in men with visceral obesity

“…By contrast, studies using rat livers VLDL apoB secretion in men with visceral obesity FM Riches et al have shown that in the long-term, insulin stimulates VLDL apoB secretion. 45,46 This may however, have been due to development of insulin resistance via insulin receptor downregulation. Our ®ndings suggest that in visceral obesity there is increased secretion of VLDL apoB which may in part be due to loss of sensitivity to the normal insulin-mediated suppression of VLDL apoB secretion.…”

Role of insulin in hepatic fatty acid partitioning: emerging concepts

“…Culture of hepatocytes with insulin for 6-48 h results in the inhibition of secretion, whereas more prolonged culture (up to 72 h) results in stimulation of secretion (over a diminishing control value) [142][143][144]. Although this phenomenon has not always been observed [6,145], it has been interpreted as indicating that insulin acts acutely on the liver to inhibit TAG secretion, whereas, when the liver is chronically exposed to high levels of insulin, it becomes resistant to the hormone and the effect is lost.…”

Glucose Phosphorylation Is Essential for the Turnover of Neutral Lipid and the Second Stage Assembly of Triacylglycerol-Rich ApoB-Containing Lipoproteins in Primary Hepatocyte Cultures

“…Hepatocytes were prepared under sterile conditions from rats weighing between 220 and 280 g at 10 AM (2 hours into the light phase of the cycle), as described previously. 13 The isolated hepatocytes were suspended at a concentration of 0.65ϫ10 6 cells/mL in Waymouth's medium MB752/1 containing glucose (25 mmol/L), methionine (0.33 mmol/L), fetal bovine serum (10% v/v), penicillin (90 000 U/L), streptomycin (90 000 g/L), glutamine (3.6 mmol/L), alanine (0.36 mmol/L), and serine (0.45 mmol/L). The cell suspension (3.0 mL) was plated out onto collagen-coated dishes, 39 and the cells were allowed to form a monolayer (3 to 4 hours).…”

“…This medium is subsequently referred to as supplemented medium. 13,14 At this stage, glucose was added to give initial concentrations of 0, 5, 10, 15, 20, or 25 mmol/L. Hepatocytes were cultured for 24 hours, the medium was removed, and the cells were harvested.…”

“…11 We have previously shown that high rates of VLDL output from primary hepatocytes can be maintained only when the culture medium is supplemented with certain amino acids and fatty acid precursors, which provide an environment conducive to vigorous de novo lipogenesis. [12][13][14] It has also been observed in primary hepatocyte cultures that apoB output requires efficient de novo fatty acid synthesis. 15 Glucose stimulates fatty acid synthesis in hepatocytes partly by a mechanism that involves increased expression of the lipogenic genes acetylcoenzyme A carboxylase and fatty acid synthase.…”