Abstract
In Leishmania species, Protein disulfide isomerase (PDI) - a redox chaperone, is reported to be involved in its virulence and survival. This protein has also been identified, through proteomics, as a Th1 stimulatory protein in the soluble lysate of a clinical isolate of Leishmania donovani (LdPDI). In the present study, the molecular characterization of LdPDI was carried out and the immunogenicity of recombinant LdPDI (rLdPDI) was assessed by lymphocyte proliferation assay (LTT), nitric oxide (NO) production, estimation of Th1 cytokines (IFN-γ and IL-12) as well as IL-10 in PBMCs of cured/endemic/infected Leishmania patients and cured L. donovani infected hamsters. A significantly higher proliferative response against rLdPDI as well as elevated levels of IFN-γ and IL-12 were observed. The level of IL-10 was found to be highly down regulated in response to rLdPDI. A significant increase in the level of NO production in stimulated hamster macrophages as well as IgG2 antibody and a low level of IgG1 in cured patient's serum was observed. Higher level of IgG2 antibody indicated its Th1 stimulatory potential. The efficacy of pcDNA-LdPDI construct was further evaluated for its prophylactic potential. Vaccination with this construct conferred remarkably good prophylactic efficacy (∼90%) and generated a robust cellular immune response with significant increases in the levels of iNOS transcript as well as TNF-α, IFN-γ and IL-12 cytokines. This was further supported by the high level of IgG2 antibody in vaccinated animals. The in vitro as well as in vivo results thus indicate that LdPDI may be exploited as a potential vaccine candidate against visceral Leishmaniasis (VL).
Highlights
Visceral Leishmaniasis (VL) is the most severe systemic disease among the three main categories of leishmaniasis and affects 500,000 people every year [1,2]
Because the expression of recombinant proteins by mammalian cells transfected with bacterial plasmid DNA is a critical condition for the stimulation of the immune system, the expression of the recombinant LdPDI (rLdPDI) protein was initially assessed in HEK-293T cells transfected with the pcDNA-LdPDI construct
Reported that the 52-kDa Protein disulfide isomerase (PDI) is linked to L. major virulence [35] and that there is a correlation between B cell activation/Ab production and lesion progression in Leishmania-infected patients and experimental animals [52,53]
Summary
Visceral Leishmaniasis (VL) is the most severe systemic disease among the three main categories of leishmaniasis and affects 500,000 people every year [1,2]. There is no effective vaccine against leishmaniasis and control of the disease is almost confined to chemotherapy. Increasing incidence of drugresistant strains has hampered the control of the disease even by chemotherapy [8,9,10,11]. Attention has been shifted towards the development of effective vaccines. Induction of lifelong protection against re-infection in individuals who recovered from the disease, demonstrates that a protective vaccine can be achieved, an effective vaccine against human leishmaniasis has yet to be discovered [12]
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