TNF-α-INDUCED APOPTOSIS IN DAUDI CELLS: MULTIPARAMETRIC ANALYSIS

Abstract

Tumour necrosis factor α (TNF-α) is a cytokine that induces physiological and pathophysiological effects in the immune system. In this study we analyzed its action on a human lymphoma cell line (Daudi cells) after 1 h, 6 h and 24 h of incubation. Using vital DNA stains, DNA gel electrophoresis, in situ nick translation, transmission electron microscopy and flow cytometry we showed that as early as after 6 h of treatment, target cells were able to undergo death by apoptosis. This was associated with cell shrinkage, chromatin condensation, apoptotic bodies in the cytoplasm without the typical DNA fragmentation into low molecular weight nucleosomes. Of interest was the observation of a significant number (60%) of cells positive to the nick translation in specimens treated for 6 h, decreasing to 40% in samples treated for 24 h, when most of the cells were in late apoptosis. In addition, no subdiploid peak was evident in flow cytometry regardless of the time of incubation with TNF. Our study on Daudi cells clearly supports the existence of alternative forms of apoptosis in which DNA degradation does not result only in oligonucleosomal fragmentation.