Recent reports have implicated a possible but undefined role for reactive oxygen species in the i... more Recent reports have implicated a possible but undefined role for reactive oxygen species in the induction and mediation of apoptosis. In the present study, the role of free radicals and metal ions in apoptosis induced in rat thymocytes by dexamethasone and etoposide was examined. Copper chelators, but not iron specific chelators, inhibited apoptosis induced by both these stimuli. Several antioxidants also possessed potent inhibitory effects. We therefore propose that diverse agents may induce apoptosis in thymocytes by a common mechanism involving a copper mediated Fenton reaction, generating site specific hydroxyl radicals, possibly as a result of activation of the redox sensitive transcription factor NF-KB.
The analysis, by slab gel electrophoresis, of internucleosomal DNA cleavage or laddering, charact... more The analysis, by slab gel electrophoresis, of internucleosomal DNA cleavage or laddering, characteristic of apoptosis in many cell systems, is labour intensive, difficult to automate and at best only semi-quantitative. In this report we show that CE, using dilute solutions of hydroxyethyicellulose as a replaceable sieving matrix, can be applied to the relatively rapid analysis of DNA laddering in whole digests of apoptotic rat thymocytes. Also, using the sensitivity of laser-induced fluorescence detection and the highly sensitive nucleic acid stain YO-PRO-1, the CE method reported here can use 1000-2000 fold fewer cells than needed for traditional slab gel methods.
Recent reports have implicated a possible but undefined role for reactive oxygen species in the i... more Recent reports have implicated a possible but undefined role for reactive oxygen species in the induction and mediation of apoptosis. In the present study, the role of free radicals and metal ions in apoptosis induced in rat thymocytes by dexamethasone and etoposide was examined. Copper chelators, but not iron specific chelators, inhibited apoptosis induced by both these stimuli. Several antioxidants also possessed potent inhibitory effects. We therefore propose that diverse agents may induce apoptosis in thymocytes by a common mechanism involving a copper mediated Fenton reaction, generating site specific hydroxyl radicals, possibly as a result of activation of the redox sensitive transcription factor NF-KB.
The analysis, by slab gel electrophoresis, of internucleosomal DNA cleavage or laddering, charact... more The analysis, by slab gel electrophoresis, of internucleosomal DNA cleavage or laddering, characteristic of apoptosis in many cell systems, is labour intensive, difficult to automate and at best only semi-quantitative. In this report we show that CE, using dilute solutions of hydroxyethyicellulose as a replaceable sieving matrix, can be applied to the relatively rapid analysis of DNA laddering in whole digests of apoptotic rat thymocytes. Also, using the sensitivity of laser-induced fluorescence detection and the highly sensitive nucleic acid stain YO-PRO-1, the CE method reported here can use 1000-2000 fold fewer cells than needed for traditional slab gel methods.
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Papers by James Wolfe