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Stabilization of Quercetin Paradoxically Reduces Its Proapoptotic Effect on UVB-Irradiated Human Keratinocytes

Authors' Affiliations: ¹ Arizona Cancer Center and ² Department of Cell Biology and Anatomy, University of Arizona, Tucson, Arizona; and ³ Hormel Institute, University of Minnesota, Austin, Minnesota

Requests for reprints: G. Tim Bowden, Arizona Cancer Center, Room 4999, 1515 North Campbell Avenue, Tucson, AZ 85724. Phone: 520-626-6006; Fax: 520-626-4979; E-mail: tbowden{at}azcc.arizona.edu.

UVB light promotes survival of initiated keratinocytes, in part, by the direct activation of the phosphatidylinositol 3-kinase (PI3K) signaling pathway. Novel chemopreventative agents targeting UVB-induced signaling pathways are needed to reduce the incidence of nonmelanoma skin cancer. Quercetin (Qu) is a dietary flavonoid and a known inhibitor of PI3K. We determined that Qu degrades rapidly when diluted in DMEM and incubated under normal cell culture conditions. Degradation was delayed by supplementing the medium with 1 mmol/L ascorbic acid (AA), and as expected, stabilization actually increased the effectiveness of Qu as a PI3K inhibitor because basal and UVB-induced Akt phosphorylation were reduced compared with Qu treatment in the absence of AA. Although AA stabilization increased Qu-induced apoptosis in mock-irradiated HaCaT cells, consistent with it acting as a PI3K inhibitor (13.4% Annexin V–positive cells for AA-stabilized Qu versus 6.3% for Qu), AA stabilization of Qu actually reduced the ability of the compound to induce apoptosis of UVB-irradiated HaCaTs (29.7% of Qu-treated cells versus 15.5% of AA + Qu–treated cells). Similar trends were seen in the analysis of caspase-3 and poly(ADP-ribose) polymerase cleavage. Qu is known to oxidize to form reactive products, and we found that dihydroethidium is oxidized by Qu regardless of whether or not it was stabilized. Although redox cycling occurs even in the presence of AA, stabilization reduces the accumulation of reactive Qu products that contribute to the proapoptotic effect of the compound, and thus reduces the ability of the compound to induce apoptosis of UVB-irradiated HaCaT cells.

Key Words: quercetin • HaCaT • apoptosis • PI3K • ROS