Background Uncoupling protein (UCP) 2 is related to the dysfunction of beta cells induced by fatty acids. However, whether UCP2 has similar effects on alpha cell is still not clear. This study aimed to investigate the effects of UCP2 and its possible mechanisms in lipotoxicity-induced dysfunction of pancreatic alpha cells. Methods The alpha TC1-6 cells were used in this study to evaluate the effects of palmitate and/or UCP2 inhibit factors on the glucagon secretory function, glucagon content, the glucagon mRNA level and the nitrotyrosine level in the supernatant. Meantime, the expression levels of UCP2 and peroxisome proliferator-activated receptor-gamma coactivator-1 alpha (PGC-1 alpha) were measured by real-time reverse transcription polymerase chain reaction (RT-PCR) and Western blotting. Furthermore, the possible relationship between UCP2 and insulin signal transduction pathway was analyzed. Results Palmitate stimulated alpha cell glucagon secretion and the expression of UCP2 and PGC-1 alpha, which could be partially decreased by the inhibition of UCP2. Palmitate increased nitrotyrosine level and suppressed insulin signal transduction pathway in alpha cells. Inhibition of UCP2 influenced the effects of free fatty acid on alpha cells and may relate to glucagon secretion. Conclusion UCP2 played an important role on alpha cell dysfunction induced by free fatty acid in vitro, which may be related to its effects on oxidative stress and insulin signal transduction pathway. Chin Med J 2010;123(17):2416-2423