Leishmaniasis is a parasitic disease that is prevalent in approximately 88 countries, and yet no licensed human vaccine exists against it. Towards control of leishmaniasis, we have developed Leishmania major Centrin gene deletion mutant strains (LmCen-/-) as a live attenuated vaccine, which induces a strong Th1 response to provide IFN-γ-mediated protection to the host. However, the immune mechanisms of such protection remain to be understood. Metabolomic reprogramming of the host cells following Leishmania-infection has been shown to play a critical role in pathogenicity and shaping the immune response following infection. Here, we applied untargeted mass spectrometric analysis to study the metabolic changes induced by infection with LmCen-/- and compared with virulent L. major parasite infection to identify the immune mechanism of protection. Our data shows that immunization with LmCen-/- parasites, in contrast to virulent L. major infection, alters tryptophan metabolism to down-regulate Kynurenine-AhR signaling and promote a pro-inflammatory response.

Leishmaniasis is a tropical disease present in more than 90 countries. Presently, there is no approved vaccine for human use. We have previously developed live attenuated L. mexicana Cen-/- (LmexCen-/-) as a vaccine candidate that showed excellent efficacy that was characterized by reduced activation of Th2 responses and enhanced Th1 responses, contrary to wild type L. mexicana (LmexWT) infection. Towards understanding the interplay between immune mechanisms of protection and metabolic reprogramming, we applied untargeted mass spectrometric analysis to LmexCen-/- and compared them with LmexWT infection. Data showed that enriched pentose phosphate pathway (PPP) in ears immunized with LmexCen-/- parasites, compared to naive and LmexWT-infected ears. This pathway is known to promote an M1 phenotype in macrophages, suggesting a switch to a pro-inflammatory phenotype following LmexCen-/- inoculation. Accordingly, inhibition of the PPP in macrophages cultured with LmexCen-/- parasites led to diminished production of nitric oxide, IL-12, and IL-1beta, hallmarks of classical activation. Overall, our study revealed novel immune regulatory mechanisms that may be critical for the induction of protective immunity.

Cutaneous leishmaniasis (CL) is characterized by extensive skin lesions associated with an aggressive inflammatory reaction. Despite the extensive inflammation, CL lesions are usually painless, indicating that Leishmania infection may trigger anti-nociceptive activities in the infected tissues. To this date, the molecular mechanisms responsible for this clinical phenomenon have not been identified. Through an untargeted metabolomic analysis by mass spectrometry, we found enriched anti-nociceptive metabolic pathways in mice infected with Leishmania (L.) mexicana. In particular, endogenous purines were elevated at the lesion site during chronic infection, as well as in vitro in infected macrophages, compared to non-infected mice. These purines have known anti-inflammatory and analgesic properties by acting through adenosine receptors and inhibiting transient receptor potential channels of the vanilloid subtype 1 (TRPV1). Additionally, purine metabolites can promote interleukin (IL)-10 production, with a subsequent decrease in inflammation and pain sensitivity. We also found arachidonic acid metabolism enriched in the ear lesions compared to the non-infected controls. Arachidonic acid is a metabolite of anandamide (AEA) and 2-arachidonoylglycerol (2-AG). These endocannabinoids act on cannabinoid receptors 1 and 2 and TRPV1 channels to exert anti-inflammatory and analgesic effects. Our study provides the first evidence of metabolic pathways upregulated during L. mexicana infection that may mediate anti-nociceptive effects experienced by CL patients and identifies macrophages as a source of these metabolites.