Introduction
Dengue viral infections is one of the most rapidly emerging mosquitos borne infections which is estimated to infect 390 million individuals annually 1. Although the age standardized infection rates, disability adjusted life years and mortality rates have increased over the last 30 years 2, there is no specific treatment for dengue. Due to climate change and rapid urbanization, the incidence of dengue is predicted to further increase in future3. Many dengue endemic countries experience seasonal outbreaks every year with the health care facilities becoming overwhelmed with the large number of dengue patients. Although most individuals infected with the dengue virus (DENV) develop mild illness, a significant proportion develop complications such as dengue haemorrhagic fever (DHF), organ dysfunction and bleeding4. However, as there are no prognostic markers to predict who is likely to develop severe disease and due to the non-availability of specific treatment, all dengue infected patients are serially monitored for early detection of complications for timely fluid management. Therefore, there is an urgent need to development of therapeutics for dengue.
Vascular leak is the hallmark of severe dengue, which leads to plasma leakage with fluid accumulation in pleural and peritoneal cavities, hypotension leading to shock and poor organ perfusion that contributes to organ dysfunction 5. Endothelial dysfunction that leads to vascular leakage has shown to occur due to viral factors such as the secretory protein NS1 directly acting on the endothelial glycocalyx and activating immune cells to produce inflammatory mediators and inflammatory lipid mediators such as secretory phospholipase A2 (sPLA2s) and cytoplasmic phospholipase A2 enzymes 6-8. In addition, a dysfunctional host innate immune response and pre-existing poorly neutralizing DENV specific antibodies also lead to endothelial dysfunction by acting on many different immune cells such as mast cells, monocytes and neutrophils. This in turn induces production of chymase, tryptase, leukotrienes, platelet activating factor (PAF) and sPLA2, which have shown to induce vascular leak 9-12. There are several clinical trials that have been completed and several ongoing trials that have used repurposed drugs to inhibit PAF and those that stabilize mast cells 13,14. Apart from the above completed and ongoing trials, many patients in dengue endemic countries use herbal medicines to treat patients who present with fever.
Tragia hispida, which belongs to the plant family Euphorbiaceae, is a medicinal plant (known as “Welkahambiliya” in Sinhala) is used in the Sri Lankan traditional medicine to treat fever. Although the properties of Tragia hispida have not been characterized, a similar plant in the same family (Tragia involucrate ) was shown to inhibit prostaglandin induced pain, have hepatoprotective effects against chemical induced hepatotoxicity in rats and antibacterial activity against many opportunistic bacteria 15.Justicia adathoda is another plant used for treatment of fever and many inflammation related diseases in Sri Lanka and many other countries. This plant has shown to reduce carrageenan-induced inflammation in rats and had anti-pyrexic properties16. Cyperus rotundus too is included in many traditional medicines which is used to treat pain and many gynecological problems. This plant too has shown to have many anti-inflammatory and anti-pyretic properties 17. However, the use and potential action of these plant extracts in treating patients with dengue has not been explored previously.
As many plant extracts are given in the aqueous form traditional medicine in Sri Lanka and many countries for treatment of fever, we sought to explore if any of these plant extracts were able to inhibit inflammatory mediators that associate with endothelial dysfunction in dengue. We previously showed that PAF was an important cause of vascular leakage and that phospholipase enzymes are important in the generation of PAF 18. It was shown that the activity of the inflammatory lipid enzyme sPLA2 was significantly increased during the early illness in those who progressed to develop DHF9,10. DENV NS1 protein was shown to induce sPLA2 activity and cPLA2 activity 6. These phospholipase A2 enzymes hydrolyze membrane phospholipids thereby generating fatty acids, lysolipds and generate PAF. Therefore, we proceeded to explore if some of the plant extracts used in traditional medicine had any sPLA2 inhibitory activity.