Production of tumor necrosis factor by rat mesangial cells in response to bacterial lipopolysaccharide. Tumor necrosis factor (TNF) is a cytokine which is produced by mononuclear phagocytes upon activation by bacterial lipopolysaccharide (LPS) and various other stimuli. In immune-mediated glomerulonephritis, infiltration of glomeruli by monocytes-macrophages is associated with production of TNF. The purpose of the present experiments was to determine whether mesangial cells could also contribute to glomerular TNF synthesis. TNF activity has been determined in the culture medium of rat mesangial cells using a L-929 fibroblast lytic assay. This activity was detectable only when the cells were exposed to LPS (0.1 to 10 µg/ml) and for periods longer than one hour. The cytotoxic factor was identified as TNF since: (1) the lytic activity was completely inhibited by an anti-mouse TNF polyclonal antibody and was associated with suppression of lipoprotein lipase activity in adipocytes; (2) its molecular weight (110,000 daltons) corresponded to that observed for murine TNF under non-denaturing conditions; and (3) mRNA encoding TNF was expressed by mesangial cells two hours after addition of LPS. To assess the mechanisms whereby TNF production was regulated, the role of prostaglandin E₂ (PGE₂) was determined. LPS caused a dose-dependent increase of PGE₂ synthesis by mesangial cells. Treatment by indomethacin promoted a suppression of PGE₂ production together with an increase of TNF synthesis, indicating that PGE₂ acted in a negative feedback manner to regulate the production of TNF. Addition of PGE₂ (0.1 to 300 nM) or 8-bromo cyclic AMP (0.1 to 100 µM) induced similar dose-dependent reductions of TNF synthesis. Thus the inhibitory effect of PGE₂ probably required in part cyclic AMP accumulation. It is concluded that TNF is released by mesangial cells upon stimulation by LPS, and that its synthesis is regulated by both PGE₂ and cyclic AMP. Production of TNF could participate in the mechanism of endotoxin-induced glomerular injury.