Tumor necrosis factor (TNF)-alpha plays a central role in the state of insulin 
resistance associated with obesity. It has previously been shown that one 
important mechanism by which TNF-alpha interferes with insulin signaling is 
through the serine phosphorylation of insulin receptor substrate-1 (IRS-1), 
which can then function as an inhibitor of the tyrosine kinase activity of the 
insulin receptor (IR). However, the receptors and the signaling pathway used by 
TNF-alpha that mediate the inhibition of IR activity are unknown. We show here 
that human TNF-alpha, which binds only to the murine p55 TNF receptor (TNFR), is 
as effective at inhibiting insulin-dependent tyrosine phosphorylation of IR and 
IRS-1 in adipocytes and myeloid 32D cells as murine TNF-alpha, which binds to 
both p55 TNFR and p75 TNFR. Likewise, antibodies that are specific agonists for 
p55 TNFR or p75 TNFR demonstrate that stimulation of p55 TNFR is sufficient to 
inhibit insulin signaling, though a small effect can also be seen with 
antibodies to p75 TNFR. Exogenous sphingomyelinase and ceramides, known to be 
formed by activation of p55 TNFR, inhibit IR and IRS-1 tyrosine phosphorylation 
and convert IRS-1 into an inhibitor of IR tyrosine kinase in vitro. Myeloid 32D 
cells expressing IR and IRS-1 are sensitive to this inhibition, but cells 
expressing IR and IRS-2 are resistant, pointing to an important difference in 
the biological function between IRS-1 and IRS-2. These data strongly suggest 
that TNF-alpha inhibits insulin signaling via stimulation of p55 TNFR and 
sphingomyelinase activity, which results in the production of an inhibitory form 
of IRS-1.