Tumor necrosis factor-alpha (TNFalpha) is a potential mediator of beta cell destruction in insulin-dependent diabetes mellitus. We have studied TNF-responsive pathways leading to apoptosis in beta cells. Primary beta cells express low levels of the type I TNF receptor (TNFR1) but do not express the type 2 receptor (TNFR2). Evidence for TNFR1 expression on beta cells came from flow cytometry using monoclonal antibodies specific for TNFR1 and TNFR2 and from RT-PCR of beta cell RNA. NIT-1 insulinoma cells similarly expressed TNFR1 (at higher levels than primary beta cells) as detected by flow cytometry and radio-binding studies. TNF induced NF-kappaB activation in both primary islet cells and NIT-1 cells. Apoptosis in response to TNFalpha was observed in NIT-1 cells whereas apoptosis of primary beta cells required both TNFalpha and interferon-gamma (IFNgamma). Apoptosis could be prevented in NIT-1 cells by expression of dominant negative Fas-associating protein with death domain (dnFADD). Apoptosis in NIT-1 cells was increased by coincubation with IFNgamma, which also increased caspase 1 expression. These data show that TNF-activated pathways capable of inducing apoptotic cell death are present in beta cells. Caspase activation is the dominant pathway of TNF-induced cell death in NIT-1 cells and may be an important mechanism of beta cell damage in insulin-dependent diabetes mellitus.