Arterial stiffness and compliance are major predictors of adverse cardiovascular events and are influenced by female sex hormones, including estrogen and progesterone. The aim of this longitudinal study was to evaluate the effect of the menstrual cycle, normal pregnancy, and preeclampsia on central and systemic arterial stiffness. Ten healthy nulliparous women with regular menses were studied in the early and midfollicular, periovulatory, and luteal phases of a single menstrual cycle. Twenty-two primigravida pregnant women were studied throughout pregnancy at 16, 24, 32, and 37 weeks gestation and at 7 weeks postpartum. Fifteen primigravida women with preeclampsia were studied at diagnosis and 7 weeks postpartum. Augmentation index and carotid-radial and carotid-femoral pulse wave velocities were measured using applanation tonometry. Augmentation index fell during the luteal phase of the menstrual cycle (luteal phase versus periovulatory phase; P<0.05). In normal pregnancy, pulse wave velocity and augmentation index increased from 24 weeks over the third trimester (P≤0.01 for both). All of the measures were increased in women with preeclampsia (P≤0.01), with augmentation index and carotid-femoral pulse wave velocity remaining elevated 7 weeks postpartum (P≤0.02). We conclude that systemic arterial stiffness undergoes major changes during the menstrual cycle and pregnancy and that preeclampsia is associated with greater and more prolonged increases in arterial stiffness. These effects may contribute to adverse cardiovascular outcomes of pregnancy and preeclampsia.