Cytochrome P450 1B1 protects against angiotensin II (Ang II)–induced hypertension and associated cardiovascular changes in female mice, most likely via production of 2-methoxyestradiol. This study was conducted to determine whether 2-methoxyestradiol ameliorates Ang II–induced hypertension, renal dysfunction, and end-organ damage in intact Cyp1b1−/−, ovariectomized female, and Cyp1b1+/+ male mice. Ang II or vehicle was infused for 2 weeks and administered concurrently with 2-methoxyestradiol. Mice were placed in metabolic cages on day 12 of Ang II infusion for urine collection for 24 hours. 2-Methoxyestradiol reduced Ang II–induced increases in systolic blood pressure, water consumption, urine output, and proteinuria in intact female Cyp1b1−/− and ovariectomized mice. 2-Methoxyestradiol also reduced Ang II–induced increase in blood pressure, water intake, urine output, and proteinuria in Cyp1b1+/+ male mice. Treatment with 2-methoxyestradiol attenuated Ang II–induced end-organ damage in intact Cyp1b1−/− and ovariectomized Cyp1b1+/+ and Cyp1b1−/− female mice and Cyp1b1+/+ male mice. 2-Methoxyestradiol mitigated Ang II–induced increase in urinary excretion of angiotensinogen in intact Cyp1b1−/− and ovariectomized Cyp1b1+/+ and Cyp1b1−/− female mice but not in Cyp1b1+/+ male mice. The G protein–coupled estrogen receptor 1 antagonist G-15 failed to alter Ang II–induced increases in blood pressure and renal function in Cyp1b1+/+ female mice. These data suggest that 2-methoxyestradiol reduces Ang II–induced hypertension and associated end-organ damage in intact Cyp1b1−/−, ovariectomized Cyp1b1+/+ and Cyp1b1−/− female mice, and Cyp1b1+/+ male mice independent of G protein–coupled estrogen receptor 1. Therefore, 2-methoxyestradiol could serve as a therapeutic agent for treating hypertension and associated pathogenesis in postmenopausal females, and in males.