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Formation of apoptotic bodies is associated with internucleosomal DNA fragmentation during drug-induced apoptosis
1Childrens Leukaemia and Cancer Research Centre, University of New South Wales, Prince of Wales Children's Hospital, Randwick, Sydney, Australia.
Abstract:
The onset of apoptosis is often coincident with internucleosomal DNA fragmentation or ladders which are considered a hallmark of the process. However, several studies have indicated that MOLT-4 human lymphoblastoid cells exposed to various agents, including VP16, display some apoptotic characteristics in the absence of either internucleosomal ladders or production of apoptotic bodies. The present study records that, in the presence of aurintricarboxylic acid (ATA), internucleosomal ladders were detected in DNA isolated from VP16-treated MOLT-4 cells; a paradoxical result in view of inhibition by ATA of nuclease activity in cell free preparations. The activity of ATA in mediating genomic fragmentation was dose- and time-dependent. Moreover, addition of ATA to VP16-treated MOLT-4 cells also resulted in production of apoptotic bodies, this effect being quantified by morphological examination and flow cytometry. Detection of ladders and apoptotic bodies after addition of ATA was not attributable to increased toxicity in cells exposed to the combined treatment relative to VP16 alone. A similar response, that is the appearance of both internucleosomal fragmentation and apoptotic bodies, occurred after exposure of MOLT-4 cells to the mitotic inhibitor podophyllotoxin. The consistent association between internucleosomal fragmentation of DNA and formation of apoptotic bodies exhibited during death of MOLT-4 cells, insofar as both characteristics are either present or absent following different agents, suggests interdependence.
Insights
Aurintricarboxylic acid (ATA) induced DNA fragmentation and apoptotic bodies in VP16-treated MOLT-4 cells, suggesting interdependence between these apoptosis hallmarks. This finding challenges previous observations of apoptosis without these features.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Apoptosis, or programmed cell death, is typically marked by DNA fragmentation and apoptotic bodies.
- However, some cell types, like MOLT-4 cells, can exhibit apoptosis without these classic signs when treated with certain agents.
- This suggests alternative or incomplete apoptotic pathways may exist.
Purpose of the Study:
- To investigate the effect of aurintricarboxylic acid (ATA) on apoptosis induction in VP16-treated MOLT-4 cells.
- To determine if ATA can restore or induce classic apoptotic features (DNA fragmentation and apoptotic bodies) in cells not showing them.
- To explore the potential interdependence of DNA fragmentation and apoptotic body formation during cell death.
Main Methods:
- Treatment of MOLT-4 human lymphoblastoid cells with VP16 and/or aurintricarboxylic acid (ATA).
- Analysis of DNA for internucleosomal fragmentation (DNA ladders) using gel electrophoresis.
- Quantification of apoptotic bodies via morphological examination and flow cytometry.
- Assessment of cellular toxicity to differentiate treatment effects from inherent toxicity.
Main Results:
- Aurintricarboxylic acid (ATA) treatment restored internucleosomal DNA fragmentation in VP16-treated MOLT-4 cells, despite ATA's known nuclease inhibitory activity in cell-free systems.
- ATA also induced the formation of apoptotic bodies in these cells, a dose- and time-dependent effect.
- Similar induction of both DNA fragmentation and apoptotic bodies was observed when cells were treated with the mitotic inhibitor podophyllotoxin.
- The combined VP16 and ATA treatment did not increase overall cell toxicity compared to VP16 alone.
Conclusions:
- The induction of both DNA fragmentation and apoptotic bodies by ATA in VP16-treated cells suggests these events are interdependent during apoptosis.
- This study challenges the notion that apoptosis strictly requires these hallmarks, showing they can be modulated.
- The findings highlight the complex regulation of programmed cell death pathways and the potential for specific agents to influence distinct apoptotic features.