Paclitaxel-induced apoptosis in human keratinocytes is enhanced by IFN-γ and is modulated by activation of caspase-8 and 3 with a concomitant down regulation of the AKT survival pathway Free

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Purpose: We have investigated the molecular effects of paclitaxel and interferon (IFN)- γ on cultured human keratinocyte cells, HaCaT assessing the induction of both the apoptotic pathway and cell survival signals. Methods: Cellular cytotoxcity assays were performed by MTT dye assay. Caspase 8, 3 and AKT (Ser473 and Ser308 residues) were assessed by Western Blot analysis. Morphological characteristics were examined by Wright stain analysis. Results: Paclitaxel reduced keratinocyte growth in a 3-day bioassay with an effective ED₅₀ of 6-600ng/ml. A large variation in ED₅₀ can be attributed to the asynchronous population of cells. Paclitaxel treatment induced activation of the AKT survival pathway in a time dependent manner. The downregulation of AKT signal was preceded by the subsequent activation of caspase-8 and 3 leading to apoptosis as evidenced by morphological assessment. These results indicate that paclitaxel activates both the PI3-AKT cell survival pathway followed by induction of apoptotic signals in cultured human keratinocytes. The induction of apoptosis in paclitaxel treated cells was enhanced by the co-administration of IFN-γ. The synergistic effect of these two agents on HaCaT cells relies on a pathway involving increased activities of caspase-8 and 3. Conclusion: Collectively, our data indicates that (a) paclitaxel-induced apoptosis is enhanced by IFN- γ, (b) the downregulation of PI3/AKT survival pathway may help potentiate the apoptotic effects of paclitaxel and (c) the apoptotic signaling pathways are initiated with the activation of caspase-8 and 3 activities.