Mechanisms of apple polyphenols-induced proliferation inhibiting and apoptosis in a metastatic oral adenoid cystic carcinoma cell line.

Adenoid cystic carcinoma (ACC) is characterized by intensive local invasion and high incidence of distant metastases. Conventional chemotherapy for ACC produces a poor result. We aimed to evaluate the effect of apple polyphenols (APs), a novel nutraceutical agent, on the proliferation and apoptosis levels in a metastatic oral ACC cell line. A metastatic ACC (ACC-M) cell line and control cells (MRC-5 cells derived from normal lung tissue) were treated with APs at different concentrations. MTT assay was used to determine the in vitro cytotoxicity. The cell cycle distribution and apoptosis levels were measured by flow cytometry. To evaluate the mechanism of APs, vascular endothelial growth factor receptor-2 (VEGFR-2) and caspase-3 messenger ribonucleic acid (mRNA) and protein levels were evaluated by reverse transcription-polymerase chain reaction and Western blots, respectively. After cells were cultured for 24 hours or 48 hours, the critical concentration of cytotoxicity of APs in MRC-5 cells was found to be 250 μg/mL. In contrast, in the concentration range of 100-250 μg/mL, the cytotoxicity of APs in ACC-M cells was time- and dose-dependent: ACC-M cell proliferation declined at 100 μg/mL when cultured for 48 hours, whereas growth was not inhibited at the concentrations of APs below 200 μg/mL when cultured for 24 hours. In selected time and dose patterns (ACC-M cells cultured at the concentrations of 150 and 250 μg/mL for 48 hours), the flow cytometry performance showed that apoptosis and necrosis occurred in APs-treated ACC-M cells. Also, in these patterns, VEGFR-2 mRNA and protein levels decreased whereas the levels of caspase-3 increased. In summary, APs could inhibit proliferation and induce apoptosis in ACC-M cells in vitro. These effects may be related to the downregulation of VEGFR-2 expression and the activation of caspase-3 expression.