Our studies in insulin resistant cells and animals, and type 2 diabetic patients showed that the IkappaB kinase, IKKbeta, plays a central role in acquired insulin resistance. In 3T3-L1 adipocytes and Fao hepatoma cells, activation of IKKbeta by treatment of TNF or by transfection of upstream kinase, NIK, or constitutively active IKK, induced cellular insulin resistance-decreased insulin-stimulated Tyr phosphorylation of IR and IRSs. Inhibition of IKKbeta by specific inhibitors- high doses of salicylates (NaS) or aspirin (ASA)- or by transfection of dominant negative IKKbeta reversed TNF-induced insulin resistance. Treatment of high doses of NaS or ASA to type 2 diabetes patients or obese and insulin resistant animals reversed insulin resistance as judged by glucose tolerance test (GTT), insulin tolerance test (ITT) and hyperinsulinemic, euglycemic clamp studies. Hyperglycemia, hyperinsulinemia, and dyslipidemia are normalized by ASA treatment. Insulin signaling studies conducted with tissues isolated from the rodents show that insulin-stimulated Tyr phosphorylation of IR and IRS are increased due to decrease in inhibitory Ser/Thr phosphorylation of IRSs. Heterozygous deletion (IkkBeta +/-) in mice reduced fasting glucose and insulin concentrations, and protected against the development of insulin resistance during high-fat feeding and in obese Lep-ob/ob mice. We also found that obese animals have higher IKK kinase activity than control animals. To study which tissue(s) are important for IKK-mediated insulin resistance and for the reversal of insulin resistance by salicylates, we generated mice expressing constitutively active IKKbeta in fat, muscle and liver with the hypothesis that increasing IKKbeta activity in animal tissues may itself induce insulin resistance. We now have colonies of all three tissue transgenic mice and found that fat- and liver-specific transgenic mice have developed insulin resistance as early as 4 week-old. We will characterize these mice to determine which tissue is responsible for IKKbeta-induced insulin resistance and how activation of IKKbeta in one tissue can induce whole body insulin resistance. These experiments will validate IKKbeta as a major mediator of insulin resistance and as a useful target for the discovery of new drugs to treat type 2 diabetes and insulin resistance.