The early stage of diabetic nephropathy (DN) is linked to proteinuria. Transforming growth factor (TGF)-β1 increases glomerular permeability to albumin (Palb), whereas 20-HETE and EETs reduce Palb. To investigate the impact of hyperglycemia and hyperlipidemia on 20-HETE, EETs, and TGF-β1 in the glomeruli, rats were divided into four groups: ND rats were fed a normal diet, HF rats were fed a high-fat diet, STZ rats were treated with 35 mg/kg of streptozotocin, and HF/STZ rats were fed a HF diet and treated with STZ. After 10 wk on these regimens, blood glucose, urinary albumin, serum cholesterol, serum triglyceride levels, and the kidney-to-body weight ratio were significantly elevated in STZ and HF/STZ rats compared with HF and ND rats. STZ and HF/STZ rats had histopathologic changes and abnormal renal hemodynamics. Expression of glomerular CYP4A, enzymes for 20-HETE production, was significantly decreased in STZ rats, whereas expression of glomerular CYP2C and CYP2J, enzymes for EETs production, was significantly decreased in both STZ and HF/STZ rats. Moreover, glomerular TGF-β1 levels were significantly greater in STZ and HF/STZ rats than in HF and ND rats. Five-week treatment of STZ rats with clofibrate induced glomerular CYP4A expression and 20-HETE production, but reduced glomerular TGF-β1 and urinary protein excretion. These results demonstrate that hyperglycemia increases TGF-β1 but decreases 20-HETE and EETs production in the glomeruli, changes that may be important in causing glomerular damage in the early stage of DN. Copyright © 2009 the American Physiological Society.