Induction of apoptosis in HeLa cells by 3beta-hydroxyurs-12-en-27-oic acid.

3Beta-hydroxyurs-12-en-27-oic acid (1), a pentacyclic triterpenoid isolated from the rhizomes of Astilbe chinensis, was structurally very similar to ursolic acid, with the only difference being the interchange of the COOH and Me group at C(14) and C(17). Ursane-type triterpene with a COOH group at C(14) is present in a limited number of natural resources. Compound 1 was found to exhibit more distinctive cytotoxicity toward human cervical squamous carcinoma (HeLa) cells than ursolic acid, suggesting that the position of the COOH group significantly affects the cytotoxicity of ursane-type pentacyclic triterpenes with a COOH group. To elucidate the underlying biological mechanism responsible for the cytotoxicity of 1, we investigated its growth-inhibitory and apoptosis-inducing effect on HeLa cells. Compound 1 induced a marked concentration- and time-dependent inhibition of cell proliferation with an IC50 value of 6.80+/-0.88 microg/ml following 48 h incubation. The drug-treated HeLa cells displayed typical morphological apoptotic characteristics and formation of DNA ladders in agarose-gel electrophoresis. Flow cytometric analysis showed that the cell cycle was arrested in G0/G1 phase by 1, and the apoptotic rate of HeLa cells treated for 48 h with 20 microg/ml of 1 was 21.08+/-2.14%. Also, 1 increased and decreased the expression of Bax and Bcl-2 proteins, respectively, and lowered the mitochondrial transmembrane potential (delta psi(m)). The peptidic caspase-3 inhibitor DEVD-CHO (NH2-Asp-Glu-Val-Asp-CHO, at 2 microM) could increase the viability of HeLa cells previously treated with 1. These results indicate that 1 induces efficient cell apoptosis through down-regulating Bcl-2 expression, up-regulating Bax expression, lowering delta psi(m), and by activating the caspase-3 pathway.

Induction of apoptosis in HeLa cells by 3beta-hydroxy-12-oleanen-27-oic acid from the rhizomes of Astilbe chinensis.

3beta-Hydroxy-12-oleanen-27-oic acid (ATA) was an antitumor active oleanane-type triterpenoid from the rhizomes of Astilbe chinensis. ATA was structurally very similar to oleanolic acid, with the only difference being interchange of the carboxyl and methyl group at the C-14 and C-17 positions. Oleanane-type triterpene with a carboxyl group at C-14 is present in a limited number of natural resources. ATA was found to exhibit more distinctive cytotoxicity toward human cervical squamous carcinoma HeLa cells than oleanolic acid, which suggested that the position of the carboxyl group significantly affects the cytotoxicity of oleanane-type pentacyclic triterpenes with a carboxyl group. The biological mechanism responsible for the cytotoxicity of ATA is not yet well understood. In this study, we investigated the induction of apoptosis in HeLa cells by ATA and the putative pathways of its actions. ATA induced a marked concentration- and time-dependent inhibition of HeLa cell proliferation, and reduced the protein content in HeLa cells. ATA-treated HeLa displayed typical morphological apoptotic characteristics and formation of DNA ladders in agarose gel electrophoresis. Flow cytometric analysis showed that the HeLa cell cycle was arrested in the G(0)/G(1) phase by ATA, and the apoptotic rate of HeLa cells treated with ATA 20 microg/mL for 48 h was 22.63 +/- 1.65%. Meanwhile, ATA increased the expression of Bax, decreased the expression of Bcl-2, and lowered the DeltaPsi(m). DEVD-CHO 2 microM could increase the viability of ATA-treated HeLa cells. These results indicate that ATA could significantly induce cell apoptosis through down-regulating Bcl-2 expression, up-regulating Bax expression, lowering DeltaPsi(m), and activating the caspase-3 pathway, and should be useful in the search for new potential anti-tumor agents and for developing semisynthetic oleanane-type triterpene derivatives with anti-tumor activity.