ABSTRACTLack of effective anti‐cardiac hypertrophy drugs creates a major cause for the increasing prevalence of heart failure. In the present study, we determined the anti‐hypertrophy and anti‐fibrosis potential of a natural plant triterpenoid, Cucurbitacin B both in vitro and in vivo. Aortic banding (AB) was performed to induce cardiac hypertrophy. After 1 week of surgery, mice were receive cucurbitacin B treatment (Gavage, 0.2 mg/kg body weight/2 day). After 4 weeks of AB, cucurbitacin B demonstrated a strong anti‐hypertrophy and ‐fibrosis ability as evidenced by decreased of heart weight, myocardial cell cross‐sectional area and interstitial fibrosis, ameliorated of systolic and diastolic abnormalities, normalized in gene expression of hypertrophic and fibrotic markers, reserved microvascular density in pressure overload induced hypertrophic mice. Cucurbitacin B also showed significant hypertrophy inhibitory effect in phenylephrine stimulated cardiomyocytes. The Cucurbitacin B‐mediated mitigated cardiac hypertrophy was attributable to the increasing level of autophagy, which was associated with the blockade of Akt/mTOR/FoxO3a signal pathway, validated by SC79, MK2206, and 3‐MA, the Akt agonist, inhibitor and autophagy inhibitor in vitro. The overexpression of constitutively active Akt completely abolished the Cucurbitacin B‐mediated protection of cardiac hypertrophy in human cardiomyocytes AC16. Collectively, our findings suggest that cucurbitacin B protects against cardiac hypertrophy through increasing the autophagy level in cardiomyocytes, which is associated with the inhibition of Akt/mTOR/FoxO3a signal axis. J. Cell. Biochem. 118: 3899–3910, 2017. © 2017 Wiley Periodicals, Inc.