Discussion
In this study we investigated the potential sPLA2 inhibitory activity of several plant extracts that are used in traditional medicine in Sri Lanka and found that T. hispida aqueous and butanol soluble fraction had potent sPLA2 inhibitory activities. In fact, it had significantly higher sPLA2 inhibitory activity than the commercial sPLA2 inhibitor CAY10590, when assessed in dengue patient sera. The HPLC analysis showed that T. hispida butanol fraction contained many flavonoid compounds, which have previously shown to inhibit sPLA2 activity 22. Snake venom is known to be a rich source of sPLA2 enzymes and aqueous solution of leaves and roots of Tragia involucrata have been taken orally by certain tribes in Tamil Nadu to treat snake bites23. However, the presence of sPLA2 inhibitory activity of these plant extracts had not been characterized previously.
The phospholipase A2 enzymes have many inflammatory actions in addition to generation of PAF, which has shown to be an important mediator of endothelial dysfunction in dengue 10. sPLA2 is an acute phase protein, with a wide range of inflammatory effects18. Lipopolysaccharide (LPS), hypoxia and cytokines have shown to induce its activity 24,25. We have previously shown that the sPLA2 activity is significantly higher during early illness (72 to 84 hours since onset of illness) in patients who subsequently proceed to develop DHF 9. In addition to its widely known inflammatory effects, sPLA2 has also shown to activate cytoplasmic PLA2 in mast cells, which in turn could contribute to generation of PAF and other arachidonic acid metabolites such as leukotrienes 26. PAF acted synergistically with sPLA2 to induce neutrophil exocytosis, thereby is likely to further contribute to endothelial dysfunction and elevated cytokines seen in dengue27. Leukotrienes (LTE4) levels were shown to be higher in patients who proceeded to develop DHF, and the LTE4 levels continued to rise in patients with DHF19. Since sPLA2 appears to have a wide range of activities in the pathogenesis of endothelial dysfunction and severe dengue, drugs which inhibit sPLA2 could be of potential therapeutic value.
In this study we have shown that the leave extract of T. hispidawhich has been used for centuries to treat fever and inflammatory diseases in traditional medicine has potent sPLA2 inhibitory activity and in the cell cytotoxicity assays, it was shown to have minimum toxicity for mammalian cell lines. However, we could not isolate the exact compound in the extract of T. hispida that had these functions. Since the whole leave extract appears to be safe due to long term consumption and based on the evidence of in vitro studies, it would be important to carry out double-blind placebo controlled clinical trials to evaluate its efficacy in treating patients with acute dengue. In addition, it would be most important to also identify the exact chemical structure of this compound that had sPLA2 inhibitory activity.