Cytokine-CpG Motif Oligodeoxynucleotide Co-inoculation in the Murine Malaria Model

Abstract

Malaria is a global problem that affects up to 500 million people and kills hundreds of thousands every year making it a serious global health predicament. Currently, there is only a partially efficacious vaccine licensed for use against malaria and available therapeutic control measures continue to be impeded by the emergence of drug-resistant strains of Plasmodia parasites which cause high morbidity and mortality. Current cutting-edge research on anti-malarial mechanisms is intensively focused on biochemical and molecular agents with the potential of improving vaccination and therapeutic strategies against malaria. The outcome of host-pathogen interactions, with respect to Plasmodia parasites, is determined by an extremely delicate balance of various biomolecules, cytokines and host-specific factors. Plasmodia parasites evade immune mechanisms and it is still unclear what exact modulations of the immune system are required in their elimination. Proper understanding of the intricate mechanisms underlying the immunopathogenesis of malaria is an essential component in the development of vaccination and therapeutic interventions. A vast array of immunopotentiating molecules like unmethylated CpG motif oligodeoxynucleotides (ODNs) operate in concert with cytokines and it was recently revealed that cytokine-CpG motif ODNs co-inoculation renders hosts resistant to Leishmania infection. The CpG ODNs exert potent immunostimulatory effects via nexus with dendritic cell Toll-like receptors (TLRs) like TLR 9 and by activating immune cells like B-cells and plasmacytoid dendritic cells (pDCs) and NK cells. The current project investigated cytokine-CpG motif ODN co-inoculation in BALB/c mice infected with P. berghei ANKA strain. Two BALB/c mice groups were infected with virulent P. berghei ANKA strain parasites, followed by five consecutive days of cytokine-involving CpG ODN-based gene therapies. One of the P. berghei-infected mice groups received IL-18- CpG ODN, and another one received IL-12-CpG ODN co-inoculation. Six other control groups with various therapeutic regimens were involved in the study. At ten days postinfection, all mice groups were humanely sacrificed for the extraction of EDTA-treated blood, plasma and splenocyte samples which were used to quantify a plethora of transcription factors (TFs), physiologic biomolecules, haematological and clinical chemistry parameters, chemokines, cytokines, immunoglobulin M (IgM) and splenocyte recall proliferation in a multiplicity of bioassays. Analysis using one-factor ANOVA unraveled cytokine-CpG co-immunotherapy as a strong trigger of antimalarial mechanisms that lead to overall parasitaemia reduction, less dramatic parasitaemia trends, milder clinico-haematological outcomes, and protective patterns in expression of TFs, physiologic biomolecules, chemokines, cytokines, IgM, and antigen-specific splenocyte recall proliferation. Murine recipients of the cytokine-CpG ODN co-inoculation ditherapy manifested with increased levels of NF-kB, NFATc, IRF-5, AhR, KLF and reduced levels of FOX-P3 and STAT-6 TFs. They also had enhanced of anti-Plasmodial activities accompanied by elevations in adiponectin, ANGPT1, NRP-1 and Cox-2 and delevations in Angiogenin, ANGPT2, MMP-8 and MMP-9 physiologic biomolecules. Cytokine-CpG ODN co-injection triggered upregulated concentrations of CCL-2, CCL-5, CXCL-12, CXCL-1, CX3CL-1, recall proliferation SI and downregulated concentrations of CCL-3, CXCL-5, CXCL-10 and CXCL-16. Augmented quantities of IFN-γ, TNF-α, IL-17, IL-23a, and IgM repressed measurements of IL-4, and IL-10 were detected with ramifications in the potential of cytokine-CpG-based DNA therapy in counteracting malaria, other infectious diseases and medical conditions.