We previously showed that the 12/15-lipoxygenase (12/15-LO) pathway of arachidonate acid metabolism is involved in multiple events related to diabetic nephropathy (DN), including glomerular hypertrophy and extracellular matrix deposition. In this study, we investigated whether in vivo delivery of small interfering RNAs (siRNAs) targeting 12/15-LO can ameliorate renal injury and DN in a streptozotocin (STZ)-injected mouse model of type 1 diabetes. To achieve greater in vivo access and siRNA expression in the kidney, we used double stranded 12/15-LO siRNA oligonucleotides conjugated with cholesterol. Diabetic DBA/2J mice were injected subcutaneously with either cholesterol tagged 12/15-LO siRNA, mismatched control siRNA, or vehicle alone, twice weekly for 7 weeks. Relative to controls, mice that received 12/15-LO siRNA showed significant reduction in albuminuria, kidney/body weight ratios, glomerular mesangial matrix expansion, renal structural damage and monocyte/macrophage infiltration. These effects were associated with lower renal cortical or glomerular levels of pro-fibrotic markers transforming growth factor-beta, connective tissue growth factor, type I and type IV collagens, plasminogen activator inhibitor 1 and fibronectin. Diabetes induced increase in glomerular cyclin dependent kinase inhibitors that are associated with hypertrophy was also prevented by siRNA administration. Our results show for the first time that systemic delivery of cholesterol tagged siRNAs targeting 12/15-LO has reno-protective effects under diabetic conditions and therefore could be a novel therapeutic approach for DN.