Effects of inhibition of pan-caspase and poly ADP-ribose polymerase (PARP) on apoptosis counts and

Joe Ogg,1 Bala Venkatesh,2 Thomas J Morgan,3 Glenda Gobe,4 Mark Jones 5

1 Tweed Hospital, Tweed Heads, NSW (primary author)

2 Princess Alexandra and Wesley Hospitals, Brisbane, QLD (animal model)

3 Mater Hospitals, Brisbane, QLD (animal model)

4 University of Queensland, Brisbane, QLD (histological analysis)

5 University of Queensland, Brisbane, QLD (statistical analysis)

Objective

To test whether broad-spectrum caspase inhibition, poly ADP-ribose polymerase (PARP) inhibition, or both together have an effect on apoptosis counts or tissue lactate concentrations following haemorrhagic shock injury.

Method

Our established model uses anaesthetised rats, with shock produced by blood withdrawal, followed by re-infusion 15 minutes later. Sixty minutes later, one of four treatments was given: normal saline (control), a PARP inhibitor(PJ34), a pancaspase inhibitor (zVAD-fmk), or both the PARP and pancaspase inhibitors.Skin and small bowel tissue was collected at four predetermined times for histological assessment of apoptosis activity and lactate assay.

Results

Comparison between time-points at baseline and after haemorrhage, but before treatment was given, showed increases in gut lactate concentrations (P=0.02), skin lactate concentrations (P=0.003) and villi apoptosis counts (P=0.004). There was no change in crypt apoptosis counts (P=0.30), and only a suggestion that skin apoptosis counts increased over time (P=0.09).No suppression of apoptosis or lactate concentrations was found with the inhibitor treatment(s). This supports the existence of a recently described pathway that is independent of caspase or PARP systems. There was some evidence (P=0.06) that rats receiving the caspase inhibitor had an increase in skin apoptosis counts in the final time period. There may be a putative caspasedependent inhibitor of late apoptosis, which was derepressed by caspase inhibition, in this study.

Conclusions

There was no reduction in apoptosis or lactate concentrations with any of the treatments. This lends support to the presence of a recently described apoptotic pathway independent of the caspase and PARP systems. Evidence (atP=0.06 level) of an association between caspase inhibition and an increase in late apoptosis suggests the presence of a caspase-dependent inhibitor of late apoptosis. Further research is required to elucidate details of such a system.