Ethyl acetate extract from Glycosmis parva leaf induces apoptosis and cell-cycle arrest by decreasing expression of COX-2 and altering BCL-2 family gene expression in human colorectal cancer HT-29 cells

Abstract

Context: Glycosmis parva Craib (Rutaceae) is reported to have cytotoxic and anti-inflammatory activities by decreasing COX-2 expression.Objective: To investigate the effect of G. parva on human colorectal cancer cells expressing COX-2, HT-29 cells.Materials and methods: HT-29 cells were treated with ethyl acetate extract from the leaves of G. parva (GPE 6.25–100 µg/ml) for 24–72 h. Cell viability was evaluated by the resuzurin reduction assay. An apoptotic study was performed using annexinV/FITC-PI staining. The cell-cycle pattern was investigated by PI staining. The expression of BCL-2 family genes was analyzed by quantitative RT-PCR and expression of cyclins and COX-2 were done by RT-PCR.Results: GPE at 6.25–100 µg/ml reduced HT-29 cell viability with IC50 values of 69.49, 55.89, and 48.94 µg/ml at 24, 48, and 72 h, respectively. HT-29 apoptosis was induced by 18.23% at 100 µg/ml. Cells in S phase decreased by 5.22% and 13.28% at 50 and 100 µg/ml, respectively, causing G0/G1 (10.6% at 50 µg/ml) and G2/M (15.67% at 100 µg/ml) accumulation. GPE at 50 µg/ml downregulated cyclin A (11.46%), cyclin E (17.98%), BCL-2 (0.32-fold), and COX-2 (29.06%) expression with an increased BAK expression (1.79-fold).Discussion and conclusion: GPE reduced HT-29 cell viability, inhibited cell proliferation, induced apoptosis, and arrested the cell cycle. Underlying mechanisms may involve decreases in COX-2, cyclin A, and cyclin E expression in addition to changes in BCL-2 family gene expression. Fundamental knowledge of GPE anticancer effects found in this study could lead to future use of this compound for colorectal cancer treatment.