It has long been a matter of debate whether the hormone-sensitive lipase (HSL)-mediated lipolysis in pancreatic beta-cells can affect insulin secretion through the alteration of lipotoxicity. We generated mice lacking both leptin and HSL (Lep(ob/ob)/HSL-/-) and explored the role of HSL in pancreatic beta-cells in the setting of obesity. Lep(ob/ob)/HSL-/- developed elevated blood glucose levels and reduced plasma insulin levels compared with Lep(ob/ob)/HSL+/+ in a fed state, while the deficiency of HSL did not affect glucose homeostasis in Lep(+/+) background. The deficiency of HSL exacerbated the accumulation of triglycerides in Lep(ob/ob) islets, leading to reduced glucose-stimulated insulin secretion. The deficiency of HSL also diminished the islet mass in Lep(ob/ob) mice due to decreased cell proliferation. In conclusion, HSL affects insulin secretary capacity especially in the setting of obesity. (C) 2009 Elsevier Inc. All rights reserved.