Pentoxifylline inhibits tumor necrosis factor-alpha-mediated cytotoxicity and cytostasis in L929 murine fibrosarcoma cells.
Tumor necrosis factor-alpha (TNF alpha) is recognized as a principal mediator of a variety of inflammatory conditions. In animal models, pentoxifylline attenuates the morbidity and mortality of bacterial sepsis, an effect which has been attributed to its ability to suppress the induction of TNF alpha. To determine whether pentoxifylline also directly inhibits the effects of TNF alpha, the ability to inhibit cytotoxicity on the TNF alpha-sensitive murine fibrosarcoma cell line, L929, was examined. Cell viability was assessed by crystal violet staining and cell proliferation was assessed by [3H]-thymidine uptake assay. TNF alpha induced dose-dependent cytotoxicity. At concentrations of TNF alpha of 1000 U/ml, viability at 3 days was approximately 35% of control. When L929 cells were co-incubated with TNF alpha (1000 U/ml) and pentoxifylline (1 mM), cell viability increased to approximately 75% of control (P = 0.001). At concentrations of TNF alpha of 10,000 U/ml, cell viability which was 11% of control with TNF alpha alone increased to 53% in the presence of pentoxifylline (P = 0.002). TNF alpha at 1000 and 10,000 U/ml concentrations decreased [3H]-thymidine uptake to approximately 5% of control values. Co-incubation with pentoxifylline significantly increased uptake to 13% of control at both TNF alpha concentrations (P = 0.002). Pentoxifylline did not affect the level of type I TNF alpha receptor--ligand cross-link product. However, in TNF alpha receptor binding assays, incubation with pentoxifylline 1 mM for 4 h was associated with an increase in the receptor affinity (control: KD = 0.42 nM vs pentoxifylline-treated: KD = 0.21 nM, P = 0.006), without significant change in number of type I TNF alpha receptors, suggesting that pentoxifylline affects post-receptor signalling events. We have observed that pentoxifylline prevents the TNF alpha-mediated activation of sn-2 arachidonic acid-specific cytosolic phospholipase A2, an important component of the signal transduction pathway of TNF alpha cytotoxicity. Because pentoxifylline does not inhibit all activities mediated by the type I TNF alpha receptor, its selective inhibition of post-receptor signalling may facilitate further study into the mechanisms underlying the diverse effects of TNF alpha.
Takahashi
G W
GW
Medical and Pathology Services, Seattle Veterans Affairs Medical Center, Washington.
Montgomery
R B
RB
Stahl
W L
WL
Crittenden
C A
CA
Valentine
M A
MA
Thorning
D R
DR
Andrews
D F
DF
Lilly
M B
MB
D000818:Animals; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D005354:Fibrosarcoma / Q000473:pathology; D051379:Mice; D010431:Pentoxifylline / Q000494:pharmacology*; D010741:Phospholipases A / Q000187:drug effects; D054467:Phospholipases A2; D018124:Receptors, Tumor Necrosis Factor / Q000187:drug effects / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism / Q000494:pharmacology*
D018124:Receptors, Tumor Necrosis Factor; D014409:Tumor Necrosis Factor-alpha; D010741:Phospholipases A; D054467:Phospholipases A2; D010431:Pentoxifylline