Photosensitization induced reactive oxygen species and oxidative damage in human erythrocytes

Generation of reactive oxygen species by photosensitization is the corner stone of photodynamic therapy of tumors. Cell damage may be mediated by free radical species and lipid peroxidation of their membranes. The effects of oxygen active species (.OH and O(2)(.-) radicals) photogenerated by the novel photosensitizer m-chloroperbenzoic acid (m-CPBA) on human erythrocyte integrity and stability were studied. The biological toxicity of the reactive oxygen species on human red blood cells (RBCs) was evident by increased osmotic fragility, spherocytosis and haemolysis. The haemolysis was increased in concentration and time dependent manner. The lipid peroxidation product thiobarbituric acid reactive substances (TBARS) was elevated in m-CPBA photosensitized RBCs indicating increased oxidative stress. This was accompanied with a depletion of erythrocyte glutathione (GSH). These effects were blunted by hydroxyl radical scavengers, thiourea and mannitol, which might indicate the production of (.)OH radical by photosensitization with m-CPBA. The antioxidant enzyme activities such as superoxide dismutase (SOD), catalase (CAT), peroxidase (Px) and glutathione peroxidase (GSH-Px) were elevated in RBCs treated with m-CPBA in the presence and absence of hydroxyl radical scavengers, mannitol and thiourea. These results suggested that the main oxygen radical photogenerated from m-CPBA is O(2)(&z.rad;-) radical, which is transformed to (.)OH radical probably by hydrogen abstraction. This is probably the main damaging oxygen species and played an essential role in oxidative haemolysis mediated by peroxidation of membrane lipids of human erythrocytes. This study provides an investigational promising data for photodynamic therapy.