curcumin and diacetyldichlorofluorescein

Ultraviolet (UV) light is a strong apoptotic trigger that induces caspase-dependent biochemical changes in cells. Previously we showed that UV irradiation can activate caspase-3, and the subsequent cleavage and activation of p21(Cdc42/Rac)-activated kinase 2 (PAK2) in human epidermoid carcinoma A431 cells. In this study we demonstrate that curcumin (Cur), the yellow pigment of Curcuma longa with known anti-oxidant and anti-inflammatory properties, can prevent UV irradiation-induced apoptotic changes, including c-Jun N-terminal kinase (JNK) activation, loss of mitochondrial membrane potential (MMP), mitochondrial release of cytochrome C, caspase-3 activation, and cleavage/activation of PAK2 in A431 cells. Flow cytometric analysis using the cell permeable dye 2',7'-dichlorofluorescin diacetate (DCF-DA) as an indicator of reactive oxygen species (ROS) generation revealed that the increase in intracellular oxidative stress caused by UV irradiation could be abolished by Cur. In addition, we found that SP600125, a JNK-specific inhibitor, reduced UV irradiation-induced JNK activation as well as caspase-3 activation, indicating that JNK activity is required for UV irradiation-induced caspase activation. Collectively, our results demonstrate that Cur significantly attenuates UV irradiation-induced ROS formation, and suggest that ROS triggers JNK activation, which in turn causes MMP change, cytochrome C release, caspase activation, and subsequent apoptotic biochemical changes.

Topics: Antineoplastic Agents; Apoptosis; Carcinoma, Squamous Cell; Caspase 3; Caspases; Curcumin; Cytochromes c; Enzyme Activation; Enzyme Inhibitors; Flow Cytometry; Fluoresceins; Humans; JNK Mitogen-Activated Protein Kinases; MAP Kinase Kinase 4; Membrane Potentials; Mitochondria; Mitogen-Activated Protein Kinase Kinases; Oxidative Stress; p21-Activated Kinases; Protein Serine-Threonine Kinases; Reactive Oxygen Species; Tumor Cells, Cultured; Ultraviolet Rays