Upregulation of low density lipoprotein receptor activity by tumor necrosis factor, a process independent of tumor necrosis factor‐induced lipid synthesis and secretion

Abstract

It has been shown that tumor necrosis factor (TNF) rapidly upregulates expression of the low density lipoprotein (LDL) receptors on Hep G2 cells and acutely stimulates hepatic lipid synthesis and secretion in vivo. It may thus be possible that TNF‐induced expression of LDL receptors is secondary to a decrease in cellular cholesterol content caused by TNF‐stimulated lipid secretion. In order to know whether TNF upregulates LDL receptors by depletion of the cellular cholesterol content, the present experiments were designed to study the temporal relationship between TNF‐stimulated expression of LDL receptor activity and TNF‐induced changes in lipid synthesis and secretion in an in vitro setting by using Hep G2 cells (a highly differentiated human hepatoma cell line) as a hepatocyte model. Hep G2 cells were incubated with TNF (usually 2.5 nmol/L) for certain periods, and LDL receptor activity was evaluated by measuring [125 I]LDL binding at 4°C; lipid synthesis and secretion were assayed by measuring [3 H]glycerol incorporation into triglycerides and phospholipids as well as [14 C]acetate incorporation into cholesterol. We found that a 30‐h exposure of the cells to TNF was needed for the effect of TNF to be seen on lipid synthesis and secretion as measured by incorporation of [3 H]glycerol into triglycerides and phospholipids, whereas TNF rapidly (in several hours) upregulated LDL receptor activity. TNF stimulated triglyceride synthesis, but did not stimulate phospholipid synthesis. On the other hand, TNF stimulated phospholipid secretion, but did not stimulate triglyceride secretion. Exposure of the cells to TNF for 16 or 24 h neither decreased cholesterol synthesis nor stimulated cholesterol secretion as measured by [14 C]acetate incorporation into cholesterol. Upregulation of LDL receptor activity through inhibition of cellular cholesterol synthesis with compactin (a competitive inhibitor of the 3‐hydroxyl‐3‐methylglutaryl‐CoA reductase) was augmented by TNF, whereas downregulation of LDL receptor activity through stimulation of cellular cholesterol synthesis with mevalonolactone almost completely blocked the upregulatory effect of TNF. In conclusion, TNF‐stimulated expression of LDL receptor activity is not secondary to a depletion of cellular cholesterol content through TNF‐stimulated lipid secretion or inhibition of cholesterol synthesis.