Apigenin, a common dietary flavonoid, has been shown to induce cell cycle arrest in both epidermal and fibroblast cells and inhibit skin tumorigenesis in murine models. The present study assessed the influence of apigenin on cell growth and the cell cycle in the human colon carcinoma cell lines SW480, HT‐29, and Caco‐2. Treatment of each cell line with apigenin (0–80 μM) resulted in a dose‐dependent reduction in both cell number and cellular protein content, compared with untreated control cultures. DNA flow cytometric analysis indicated that treatment with apigenin resulted in G2/M arrest in all three cell lines in a time‐ and dose‐dependent manner. Apigenin treatment (80 μM) for 48 h produced maximum G2/M arrest of 64%, 42%, and 26% in SW480 cells, HT‐29 cells, and Caco‐2 cells, respectively, in comparison with control cells (15%). The proportion of S‐phase cells was not altered by apigenin treatment in each of the three cell lines. The G2/M arrest was reversible after 48 h of apigenin treatment in the most sensitive cell line SW480. The degree of G2/M arrest by apigenin was inversely correlated with the corresponding inhibition of cell growth measurements in all three cell lines (r = −0.626 to −0.917, P≤0.005). Moreover, an immune complex kinase assay demonstrated an inhibition of p34cdc2 kinase activity, a critical enzyme in G2/M transition, in each cell line after treatment with apigenin (50–80 μM). Western blot analyses indicated that both p34cdc2 and cyclin B1 proteins were also decreased after apigenin treatment. These results indicate that apigenin inhibits colon carcinoma cell growth by inducing a reversible G2/M arrest and that this arrest is associated, at least in part, with inhibited activity of p34cdc2 kinase and reduced accumulation of p34cdc2 and cyclin B1 proteins. Differences in induction of G2/M arrest by apigenin in the three colon carcinoma cell lines suggest that dietary apigenin may be differentially effective against tumors with specific mutational spectra. Mol. Carcinog. 28:102–110, 2000. © 2000 Wiley‐Liss, Inc.

Apigenin, a common dietary flavonoid, has been shown to induce cell cycle arrest in both epidermal and fibroblast cells and inhibit skin tumorigenesis in murine models. The present study assessed the influence of apigenin on cell growth and the cell cycle in the human colon carcinoma cell lines SW480, HT-29, and Caco-2. Treatment of each cell line with apigenin (0-80 microM) resulted in a dose-dependent reduction in both cell number and cellular protein content, compared with untreated control cultures. DNA flow cytometric analysis indicated that treatment with apigenin resulted in G2/M arrest in all three cell lines in a time- and dose-dependent manner. Apigenin treatment (80 microM) for 48 h produced maximum G2/M arrest of 64%, 42%, and 26% in SW480 cells, HT-29 cells, and Caco-2 cells, respectively, in comparison with control cells (15%). The proportion of S-phase cells was not altered by apigenin treatment in each of the three cell lines. The G2/M arrest was reversible after 48 h of apigenin treatment in the most sensitive cell line SW480. The degree of G2/M arrest by apigenin was inversely correlated with the corresponding inhibition of cell growth measurements in all three cell lines (r = -0.626 to -0.917, P

This research assessed the importance of the adenomatous polyposis coli (APC) tumor suppressor mutation in the ability of apigenin to induce cell cycle arrest using HT29-APC cells, which contain inducible wild-type APC under the metallothionein promoter. HT29-GAL cells, containing beta-galactosidase (GAL), were included as control. Treatment with apigenin (0, 20, 40, 60, and 80 microM) for 48 h resulted in reduction in the cell number (P < 0.05) concurrent with flow cytometry results showing a dose-dependent accumulation of cells in the G2/M phase in both HT29-APC and HT29-GAL cells without ZnCl(2) treatment. Flow cytometric analysis showed an increase (P < 0.05) in the percentage of cells in G2/M when HT29-APC cells were treated with 80 microM apigenin for 120 h. This increase was not present in HT29-APC cells when treated with both 80 microM apigenin and 100 microM ZnCl(2) for 120 h. Western blot analysis verified the induction of APC protein expression in ZnCl(2)-treated HT29-APC cells but not in ZnCl(2)-treated HT29-GAL cells. Apigenin plus ZnCl(2) treatment increased the expression of APC protein in HT29-APC cells by 50 fold above expression observed with ZnCl(2) alone. Upon induction of the APC gene with ZnCl(2) in HT29-APC cells, the percentage of apoptotic cells increased significantly (P< 0.05) after 120-h treatment. Additionally, apigenin treatment (80 microM) further increased the percentage of apopototic HT29-APC following ZnCl(2) treatment to induce wild-type APC expression. These results suggest that APC dysfunction may be critical for apigenin to induce cell cycle arrest in human colon cancer cell lines and furthermore, apigenin enhances APC expression and apoptosis in cells with wild-type APC.

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