Tumor Necrosis Factor-alpha Suppresses Adipocyte-specific Genes and Activates Expression of Preadipocyte Genes in 3T3-L1 Adipocytes: Nuclear Factor-kappaB Activation by TNF-alpha is Obligatory

Tumor necrosis factor-alpha (TNF-alpha) is a contributing cause of the insulin resistance seen in obesity and obesity-linked type 2 diabetes, but the mechanism(s) by which TNF-alpha induces insulin resistance is not understood. By using 3T3-L1 adipocytes and oligonucleotide microarrays, we identified 142 known genes reproducibly upregulated by at least threefold after 4 h and/or 24 h of TNF-alpha treatment, and 78 known genes downregulated by at least twofold after 24 h of TNF-alpha incubation. TNF-alpha-induced genes include transcription factors implicated in preadipocyte gene expression or NF-kappaB activation, cytokines and cytokine-induced proteins, growth factors, enzymes, and signaling molecules. Importantly, a number of adipocyte-abundant genes, including GLUT4, hormone sensitive lipase, long-chain fatty acyl-CoA synthase, adipocyte complement-related protein of 30 kDa, and transcription factors CCAAT/enhancer binding protein-alpha, receptor retinoid X receptor-alpha, and peroxisome profilerator-activated receptor gamma were significantly downregulated by TNF-alpha treatment. Correspondingly, 24-h exposure of 3T3-L1 adipocytes to TNF-alpha resulted in reduced protein levels of GLUT4 and several insulin signaling proteins, including the insulin receptor, insulin receptor substrate 1 (IRS-1), and protein kinase B (AKT). Nuclear factor-kappaB (NF-kappaB) was activated within 15 min of TNF-alpha addition. 3T3-L1 adipocytes expressing IkappaBalpha-DN, a nondegradable NF-kappaB inhibitor, exhibited normal morphology, global gene expression, and insulin responses. However, absence of NF-kappaB activation abolished suppression of >98% of the genes normally suppressed by TNF-alpha and induction of 60-70% of the genes normally induced by TNF-alpha. Moreover, extensive cell death occurred in IkappaBalpha-DN-expressing adipocytes after 2 h of TNF-alpha treatment. Thus the changes in adipocyte gene expression induced by TNF-alpha could lead to insulin resistance. Further, NF-kappaB is an obligatory mediator of most of these TNF-alpha responses.