Real-time imaging of viable-apoptotic switch in GSNO-induced mouse thymocyte apoptosis.

Many scientists are focusing on the apoptotic-necrotic or the necrotic-apoptotic switch and its mechanism, but little attention has been paid to the viable-apoptotic switch. Most of the techniques and methods used for detecting apoptosis are performed on fixed samples, yielding static information of specific time points. We have studied the viable-apoptotic switch in S-nitrosoglutathione (GSNO)-induced mouse thymocyte apoptosis in real-time by means of a novel technique, intensified charge coupled device (ICCD)-based real-time fluorescence micro-imaging, coupled with Annexin V-FITC labeling for phosphatidylserine (PS) translocation in cell membrane. We have successfully recorded the initiating time points (mostly at 2 h) of the viable-apoptotic switch in GSNO-initiated apoptosis, as well as shown the real-time differences between living and apoptotic thymocytes. These findings suggest that NO is also a switch molecule for the conversion from viable to apoptotic cell. Thymocytes cotreated by N-G-monomethyl-L-arginine acetate salt (L-NMMA) provide further evidence for this suggestion, as well as for the suggestion that L-NMMA prolongs the early stage of thymocyte apoptosis rather than strongly blocks it.