Glycyrrhizic acid suppresses Cox-2-mediated anti-inflammatory responses during Leishmania donovani infection

The aim of the present study was to characterize glycyrrhizic acid (GA) and assess its immunomodulatory potential in a model of experimental visceral leishmaniasis. The antileishmanial activity of GA was tested in an amastigotemacrophage model and its non-cytotoxic dose was measured by a cell viability assay. To understand the effector mechanism of GA-treated macrophages against leishmanial parasites, real-time PCR analysis of inducible nitric oxide synthase 2 (iNOS2) was carried out followed by measurement of nitric oxide generation by Griess reagent. The effect of GA on the production of cytokines, such as interleukin (IL)-12, tumour necrosis factor (TNF)-, IL-10 and transforming growth factor (TGF)-, was measured by ELISA (protein) and real-time PCR. The expression of iNOS2 and cyclooxygenase-2 (Cox-2) was studied by western blotting. The parasite burden of the liver and spleen following GA treatment was determined by the stamp-smear method, and T cell proliferation was assessed via [H-3]thymidine uptake, measured by a liquid scintillation counter. Results showed that GA treatment caused an enhanced expression of iNOS2 along with inhibition of Cox-2 in Leishmania donovani-infected macrophages. GA treatment in infected macrophages enhanced the expression of IL-12 and TNF-, concomitant with a down-regulation of IL-10 and TGF-. GA increased macrophage effector responses via inhibition of Cox-2-mediated prostaglandin E2 release in L. donovani-infected macrophages. GA also decreased hepatic and splenic parasite burden and increased T cell proliferation in Leishmania-infected BALB/c mice. These results provide a mechanistic understanding of GA-mediated protection against leishmanial parasites within the host.