Abstract
Protozoan parasites of Leishmania genus are able to successfully infect their host macrophage due to multiple virulence strategies that result in its deactivation. Recent studies suggest Leishmania GP63 to be a critical virulence factor in modulation of many macrophage molecules, including protein tyrosine phosphatases (PTPs) and transcription factors (TFs). Additionally, we and others recently reported that Leishmania-released exosomes can participate in pathogenesis. Exosomes are 40–100 nm vesicles that are freed by many eukaryotic cells. To better understand the GP63-dependent immune modulation of the macrophage by Leishmania parasites and their exosomes, we compared the immunomodulatory properties of Leishmania major (WT) and L. major gp63−/− (KO) as well as their exosomes in vitro and in vivo. Importantly, we observed that Leishmania exosomes can modulate macrophage PTPs and TFs in a GP63-dependent manner. In addition, our qRT-PCR analyses showed that WT parasites were able to downregulate multiple genes involved in the immune response, especially cytokines and pattern recognition receptors. KO parasites showed a strongly reduced modulatory capacity compared to WT parasites. Furthermore, comparison of WT versus KO exosomes also showed divergences in alteration of gene expression, especially of chemokine receptors. In parallel, studying the in vivo inflammatory recruitment using a murine air pouch model, we found that exosomes have stronger proinflammatory properties than parasites and preferentially induce the recruitment of neutrophils. Finally, comparative proteomics of WT and KO exosomes surprisingly revealed major differences in their protein content, suggesting a role for GP63 in Leishmania exosomal protein sorting. Collectively our data clearly establish the crucial role of GP63 in dampening the innate inflammatory response during early Leishmania infection, and also provides new insights in regard to the role and biology of exosomes in Leishmania host-parasite interactions.
Highlights
Leishmaniasis is a spectrum of diseases caused by the protozoan parasites of genus Leishmania
We evaluated the purity of the exosomes by transmission electron microscopy and observed that vesicles with the same size and morphology of exosomes have been purified from the Conditioned Medium (CM) of both wild type (WT) and KO parasites (Figure 1D and E)
We further showed that GP63-bearing Leishmania exosomes have inflammatory properties and are capable of macrophage immune modulation at both signaling and gene expression levels
Summary
Leishmaniasis is a spectrum of diseases caused by the protozoan parasites of genus Leishmania. This disease ranges from self-healing lesions of cutaneous leishmaniasis (caused mainly by Leishmania mexicana and Leishmania major) to potentially lethal visceral leishmaniasis (caused mainly by Leishmania donovani and Leishmania infantum). When the sandfly takes a bloodmeal, the parasites are subcutaneously injected into the mammalian host. There, they are taken up by phagocytic cells and transform into amastigotes in the phagolysosome of the macrophages, their definitive mammalian host cell. The cycle is completed when amastigotes and infected cells are picked up in another bloodmeal [1]
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