Notch signaling controls cellular differentiation and proliferation. Recent studies have shown that Notch signaling plays an important role in the carcinogenesis and progression of a growing number of malignant tumors. We investigated the effect of Notch1 activation on human hepatocellular carcinoma (HCC). In five human HCC cell lines, it was found that SMMC7721 had relatively high while HepG2 relatively low expression of Notch1 and the activity of Notch signaling. Notch1 activation by transfection of active intracellular region of Notch1 (ICN1) into HCC HepG2 cells enhanced cell growth and proliferation, including in vitro single cell colony formation, anchorage-independent proliferation, and in vivo tumorigenicity. Notch1 activation also promoted HepG2 cell cycle progression. Suppression of Notch1 activation by RNAi of Notch1 or by γ-secretase inhibitor (GSI) in HCC SMMC7721 cells decreased cell growth capability and blocked cell cycle progression. Moreover, it was found that suppression of Notch1 activation induced SMMC7721 cell apoptosis, as demonstrated by apoptosis assays. These findings indicate that Notch1 activation promotes human HCC cell growth and proliferation, which may contribute to the progression of this type of malignant carcinoma.