Exposure to a nontoxic dose of bacterial endotoxin (lipopolysaccharide [LPS]) potentiates the hepatotoxicity of aflatoxin B(1) (AFB(1)). Because some of the pathophysiologic effects associated with LPS are mediated through tumor necrosis factor alpha (TNF-alpha), this study was conducted to explore the role of TNF-alpha in the AFB(1)/LPS model. Male Sprague-Dawley rats (250-300 g) were treated with either 1 mg AFB(1)/kg, intraperitoneally, or its vehicle (0.5% dimethyl sulfoxide [DMSO]/water), and 4 hours later with either Escherichia coli lipopolysaccharide (7.4 x 10(6)EU/kg, intravenously) or its saline vehicle. LPS administration resulted in a marked rise in TNF-alpha levels at 6 hours, which preceded the onset of liver injury. TNF-alpha messenger RNA (mRNA) in liver was increased by LPS treatment. The mRNA of receptors (R1 and R2) for TNF-alpha was also examined. R1 mRNA levels were not altered; however, R2 mRNA levels were increased by either AFB(1) or LPS administration. To determine if TNF-alpha plays a causal role in the development of liver injury, the increase in TNF-alpha was attenuated by administration of either pentoxifylline or anti-TNF-alpha serum, and liver injury was assessed. Administration of either of these agents resulted in protection. LPS treatment resulted in the upregulation of gene transcription for cyclooxygenase-2 (COX-2). However, administration of the selective COX-2 inhibitor NS-398 did not decrease injury. TNF-alpha and COX-2 inhibitors did not affect hepatic sequestration of neutrophils. Furthermore, it did not appear that TNF-alpha contributed to injury through inhibition of tissue repair. These data support the hypothesis that LPS-induced expression of TNF-alpha underlies the potentiation of AFB(1)-induced hepatotoxicity.