Mechanisms of apoptosis: UVA1‐induced immediate and UVB‐induced delayed apoptosis in human T cells in vitro†

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Abstract
The decreased number of lymphocytes combined with the induction of apoptosis and necrosis seems to be the key mechanism of many phototherapeutic agents. The purpose of our study was to determine the regulating pathway, time course and dose dependence of UVA1- vs. UVB-induced cell death in human T lymphocytes. In our study we applied an in vitro method using single-laser flow cytometry differentiating between intact (Annexin V-FITC-/PI-), apoptotic (Annexin V-FITC+/PI-) and necrotic T cells (Annexin V-FITC+/PI+) following UVA1 (340-400 nm) or UVB (280-320 nm) irradiation. Additionally, fluorescence microscopy of apoptotic cells was performed using acridine orange and ethidium bromide. Compared to DNA-binding fluorescent microscopy, the flow cytometric method revealed similar, but more precise, results concerning apoptosis and necrosis. Our data indicate that UVB irradiation exerts its effects by the induction of delayed apoptosis within 24-48 h. In contrast, UVA1 irradiation acts via the dose-dependent induction of immediate apoptosis and necrosis within 6 h. Our findings demonstrate that UVA1 irradiation may effect structural and functional modifications leading to immediate initiation of apoptosis followed by early membrane rupture, whereas UVB irradiation leads to DNA damage followed by delayed apoptosis, obviously without initial membrane alteration.