Abstract
No vaccine exists against visceral leishmaniasis. Toward developing vaccines against VL, we have reported previously on the immunogenicity of live attenuated LdCen−/− parasites in animal models. Immunization with LdCen−/– parasites has been shown to induce durable protective immunity in pre-clinical animal models. Although the innate immune responses favoring a Th1 type immunity are produced following LdCen−/− immunization, the molecular determinants of such responses remain unknown. To identify early biomarkers of immunogenicity associated with live attenuated parasitic vaccines, we infected macrophages derived from healthy human blood donors with LdCen−/− or LdWT parasites ex vivo and compared the early gene expression profiles. In addition to altered expression of immune related genes, we identified several microRNAs that regulate important cytokine genes, significantly altered in LdCen−/– infection compared to LdWT infection. Importantly, we found that LdCen−/– infection suppresses the expression of microRNA-21 (miR-21) in human macrophages, which negatively regulates IL12, compared to LdWT infection. In murine DC experiments, LdCen−/− infection showed a reduced miR-21 expression with a concomitant induction of IL12. Silencing of miR-21 using specific inhibitors resulted in an augmented induction of IL12 in LdWT infected BMDCs, illustrating the role of miR-21 in LdWT mediated suppression of IL12. Further, exosomes isolated from LdCen−/− infected DCs contained significantly reduced levels of miR-21 compared to LdWT infection, that promoted proliferation of CD4+ T cells in vitro. Similar miR-21 mediated IL12 regulation was also observed in ex vivo human macrophage infection experiments indicating that miR-21 plays a role in early IL12 mediated immunity. Our studies demonstrate that LdCen−/− infection suppresses miR-21 expression, enables IL12 mediated induction of adaptive immunity including proliferation of antigen experienced CD4+ T cells and development of a Th1 immunity, and suggest that miR-21 could be an important biomarker for LdCen−/− vaccine immunity in human clinical trials.One Sentence SummaryRole of miR-21 in vaccine induced immunity.
Highlights
Leishmaniasis, caused by the protozoan parasites Leishmania, is a neglected protozoan disease that mainly affects the tropical as well as subtropical countries of the world
RNA sequencing revealed distinct microRNA expression profiles in human macrophages upon LdCen−/− infection compared to LdWT infection (Figure 1A, Supplementary Figure 2)
Of the several microRNAs that are enriched in LdWT infection compared to LdCen−/− infection, miR-21 was selected for further study due to its role in the regulation of IL-12 expression in recent studies [21, 22]
Summary
Leishmaniasis, caused by the protozoan parasites Leishmania, is a neglected protozoan disease that mainly affects the tropical as well as subtropical countries of the world. 200,000–400,000 new cases of visceral leishmaniasis (VL) are reported that results in 20,000–40,000 deaths with 95% of the fatal cases occurring in six countries, namely, India, Bangladesh, Sudan, Nepal, Ethiopia, and Brazil [1]. Most of the cases of leishmaniasis diagnosed in the United States are in people who became infected while traveling or living in other countries, autochthonous cases of cutaneous leishmaniasis are reported in southern United States [https://www.cdc.gov/parasites/leishmaniasis/gen_ info/faqs.html, [5]]. These factors pose a serious risk of spread of the disease in the US by these blood-borne pathogens. There is an urgent need for developing effective vaccines against human leishmaniasis
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