Heme oxygenase-1 (HO-1) overexpression using gene transfer protects rat livers against ischemia/reperfusion (I/R) injury. This study evaluates the effects of Ad-HO-1 gene transfer in a rat renal isograft model. Donor LEW kidneys were perfused with Ad-HO-1, Ad-beta-gal, or PBS, stored at 4 degrees C for 24 h, and transplanted orthotopically into LEW recipients, followed by contralateral native nephrectomy. Serum creatinine, urine protein/creatinine ratios, severity of histologic changes, HO-1 mRNA/protein expression, and HO enzymatic activity were analyzed. Ad-HO-1 gene transfer conferred a survival advantage when compared with PBS- and Ad-beta-gal-treated controls, with median survival of 100, 7, and 7 d, respectively (P < 0.01). Serum creatinine levels were elevated at day 7 in all groups (range, 2.2 to 5.8 mg/dl) but recovered to 1.0 mg/dl by day 14 (P < 0.01) in Ad-HO-1 group, which was sustained thereafter. Urine protein/creatinine ratio at day 7 was elevated in both PBS and Ad-beta-gal, as compared with the Ad-HO-1 group (12.0 and 9.8 versus 5.0; P < 0.005); histologically, ATN and glomerulosclerosis was more severe in Ad-beta-gal group at all time points. Reverse transcriptase-PCR-based HO-1 gene expression was significantly increased before reperfusion (P < 0.001) and remained increased in the Ad-HO-1-treated group for 3 d after transplantation. Concomitantly, HO enzymatic activity was increased at transplantation and at 3 d posttransplant in the Ad-HO-1 group, compared with Ad-beta-gal controls (P < 0.05); tubular HO-1 expression was discernible early posttransplant in the Ad-HO-1 group alone. These findings are consistent with protective effects of HO-1 overexpression using a gene transfer approach against severe renal I/R injury, with reduced mortality and attenuation of tissue injury.