Withania somnifera chemotype NMITLI 101R significantly increases the efficacy of antileishmanial drugs by generating strong IFN-γ and IL-12 mediated immune responses in Leishmania donovani infected hamsters

Abstract

Background: Withania somnifera (L.) Dunal (Solanaceae), commonly known as Ashwagandha, is one of the most important medicinal plant in the traditional Indian medical systems. Pharmacological studies have established that root extracts of W. somnifera contain several bioactive constituents called withanolides. The plant has long been used for its several beneficial properties and recently as an immunomodulator. Hypothesis/Purpose: A combination therapy including a potential and safe immunostimulant with lower doses of effective drug, which can reduce the parasitic burden and simultaneously can produce an enhancement of adaptive immunity, has proven to be significantly a more effective approach than immunotherapy or drug therapy alone. Study Design: Evaluation of the immunostimulatory effect of W. somnifera chemotype NMITLI 101R when used in combination with ED₅₀ doses of antileishmanial drugs in Leishmania donovani infected hamsters. Methods: Infected animals were administered with chemotype 101R(30 mg/kg × 15 days) either alone or in combination with ED₅₀ doses of miltefosine (10 mg/kg × 5 days), paromomycin (30 mg/kg × 5 days) or amphotericin B (0.5 mg/kg × 5 days). The treated animals were euthanized on days 30 and 60 post-treatment (p.t.) and checked for parasite clearance, delayed type hypersensitivity (DTH) response, cytokine and inducible nitric oxide synthase levels by real-time PCR, nitric oxide (NO) production, reactive oxygen species (ROS) generation, lymphoproliferative and antibody responses. Results: The group of animals that received 101R and ED₅₀ dose of miltefosine showed optimum inhibition of parasite multiplication (∼98%) by day 60 p.t. followed by the group that received 101R plus paromomycin (∼94%) and 101R plus amphotericin B (∼93%). The efficacy was well supported by the increased inducible NO synthase mRNA transcript, strong IFN-γand IL-12 mediated Th1 immune responses and significantly suppressed levels of Th2 cytokines (IL-4, IL-10 and TGF-β). Additionally, same therapy also induced significant increase in the level of NO production, ROS generation, Leishmania specific IgG2 antibody along with profound DTH and strong T-cell responses as compared with all the other treated groups. Conclusion: Our results suggest that combination of chemotype 101R with ED₅₀ doses of antileishmanial drugs may provide a promising alternative for the cure of visceral leishmaniasis with significant restoration of the host immune response.