BACKGROUND Diabetic nephropathy (DN) is a progressive microvascular complication of diabetes mellitus (DM), driven largely by podocyte apoptosis. The cysteine protease Calpain 10 is known to augment apoptosis and necrosis, and is a potential therapeutic target in DN. METHODS Type 2 diabetes was induced in SD rats by high-fat diet (HFD) feeding and streptozotocin (STZ) injections, and simulated in vitro by culturing conditionally immortalized mouse podocytes in hyperlipidemic (PA, 100 μM) conditions. The rate of apoptosis in the renal tissues and cultured podocytes was determined by TUNEL assay. The expression of Calpain 10 and its biological effects were assayed by real-time PCR, Western blotting, immunofluorescence and electron microscopy. RESULTS Calpain 10 was up-regulated in the kidneys of DN rats, as well as immortalized mouse podocytes. High levels of Calpain 10 was associated with renal dysfunction and tissue destruction, and podocyte injury and apoptosis. Knockdown of Calpain 10 protected podocytes by decreasing apoptosis rate, and upregulated nephrin. CONCLUSIONS Calpain 10 is a pro-apoptotic factor in DN, and can be targeted for treating glomerular diseases.