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Malaria is a common and sometimes fatal disease caused by infection with Plasmodium parasites. Cerebral malaria (CM) is a most severe complication of infection with Plasmodium falciparum parasites which features a complex immunopathology that includes a prominent neuroinflammation. The experimental mouse model of cerebral malaria (ECM) induced by infection with Plasmodium berghei ANKA has been used abundantly to study the role of single genes, proteins and pathways in the pathogenesis of CM, including a possible contribution to neuroinflammation. In this review, we discuss the Plasmodium berghei ANKA infection model to study human CM, and we provide a summary of all host genetic effects (mapped loci, single genes) whose role in CM pathogenesis has been assessed in this model. Taken together, the reviewed studies document the many aspects of the immune system that are required for pathological inflammation in ECM, but also identify novel avenues for potential therapeutic intervention in CM and in diseases which feature neuroinflammation.

Genes and pathways in which inactivation dampens tissue inflammation present new opportunities for understanding the pathogenesis of common human inflammatory diseases, including inflammatory bowel disease, rheumatoid arthritis and multiple sclerosis. We identified a mutation in the gene encoding the deubiquitination enzyme USP15 (Usp15L749R) that protected mice against both experimental cerebral malaria (ECM) induced by Plasmodium berghei and experimental autoimmune encephalomyelitis (EAE). Combining immunophenotyping and RNA sequencing in brain (ECM) and spinal cord (EAE) revealed that Usp15L749R-associated resistance to neuroinflammation was linked to dampened type I interferon responses in situ. In hematopoietic cells and in resident brain cells, USP15 was coexpressed with, and functionally acted together with the E3 ubiquitin ligase TRIM25 to positively regulate type I interferon responses and to promote pathogenesis during neuroinflammation. The USP15-TRIM25 dyad might be a potential target for intervention in acute or chronic states of neuroinflammation.

The polymorphism 677CT (NM_005957.4:c.665CT/p.Ala222Val, rs1801133:CT) in methylenetetrahydrofolate reductase (MTHFR) results in mild enzymatic deficiency and increased risk for several complex traits including adverse reproductive outcomes, birth defects, and heart disease. Despite these deleterious effects, homozygosity is high (5%-15%) in many populations, and among the highest in Mediterranean regions, where malaria was historically endemic and may have conferred a selective advantage for other mutations. We infected Mthfr-deficient (Mthfr(+) (/-) ) and MTHFR overexpressing (MTHFR(Tg) ) mice with Plasmodium berghei ANKA to induce cerebral malaria. Mthfr(+/-) mice survived longer (P 0.02, log-rank test), and MTHFR(Tg) mice died earlier (P 0.05, log-rank test) after infection compared with wild-type littermates. Flow cytometry revealed increased lymphocyte populations and increased CCR4(+) NK cells in spleen of Mthfr(+) (/-) mice; MTHFR(Tg) animals had decreased numbers of these NK cells. Interferon-γ and interleukin-10 immunoreactive proteins were increased and decreased, respectively, in brain of Mthfr(+/-) mice compared with wild-type. We suggest that mild MTHFR deficiency protects against malarial infection and that this phenomenon may have led to the high frequency of the 677CT/c.665CT variant in human populations.

Background The potential emergence and spread of resistance to artemisinins in the Plasmodium falciparum malaria parasite constitutes a major global health threat. Hence, improving the efficacy of artemisinins and of artemisinin-based combination therapy (ACT) represents a major short-term goal in the global fight against malaria. Mice defective in the enzyme pantetheinase (Vnn3) show increased susceptibility to blood-stage malaria (increased parasitaemia, reduced survival), and supplementation of Vnn3 mutants with the reaction product of pantetheinase, cysteamine, corrects in part the malaria-susceptibility phenotype of the mutants. Cysteamine (Cys) is a small, naturally occurring amino-thiol that has very low toxicity in vivo and is approved for clinical use in the life-long treatment of the kidney disorder nephropathic cystinosis. Methods The ability of Cys to improve the anti-plasmodial activity of different clinically used artemisinins was tested. The effect of different CYS/ART combinations on malarial phenotypes (parasite blood-stage replication, overall and survival from lethal infection) was assessed in a series of in vivo experiments using Plasmodium strains that induce either blood-stage (Plasmodium chabaudi AS) or cerebral disease (Plasmodium berghei ANKA). This was also evaluated in an ex vivo experimental protocol that directly assesses the effect of such drug combinations on the viability of Plasmodium parasites, as measured by the ability of tested parasites to induce a productive infection in vivo in otherwise naïve animals. Results Cys is found to potentiate the anti-plasmodial activity of artesunate, artemether, and arteether, towards the blood-stage malaria parasite P. chabaudi AS. Ex vivo experiments, indicate that potentiation of the anti-plasmodial activity of artemisinins by Cys is direct and does not require the presence of host factors. In addition, potentiation occurs at sub-optimal concentrations of artemisinins and Cys that on their own have little or no effect on parasite growth. Cys also dramatically enhances the efficacy and protective effect of artemisinins against cerebral malaria induced by infection with the P. berghei ANKA parasite. Conclusion These findings indicate that inclusion of Cys in current formulations of ACT, or its use as adjunct therapy could improve the anti-plasmodial activity of artemisinin, decrease mortality in cerebral malaria patients, and prevent or delay the development and spread of artemisinin resistance. Electronic supplementary material The online version of this article (doi:10.1186/s12936-016-1317-3) contains supplementary material, which is available to authorized users.

We identify an N -ethyl- N -nitrosourea (ENU)-induced I23N mutation in the THEMIS protein that causes protection against experimental cerebral malaria (ECM) caused by infection with Plasmodium berghei ANKA. Themis I23N homozygous mice show reduced CD4 + and CD8 + T lymphocyte numbers. ECM resistance in P. berghei ANKA-infected Themis I23N mice is associated with decreased cerebral cellular infiltration, retention of blood-brain barrier integrity, and reduced proinflammatory cytokine production. THEMIS I23N protein expression is absent from mutant mice, concurrent with the decreased THEMIS I23N stability observed in vitro . Biochemical studies in vitro and functional complementation in vivo in Themis I23N/ + : Lck −/ + doubly heterozygous mice demonstrate that functional coupling of THEMIS to LCK tyrosine kinase is required for ECM pathogenesis. Damping of proinflammatory responses in Themis I23N mice causes susceptibility to pulmonary tuberculosis. Thus, THEMIS is required for the development and ultimately the function of proinflammatory T cells. Themis I23N mice can be used to study the newly discovered association of THEMIS (6p22.33) with inflammatory bowel disease and multiple sclerosis.

In humans, cerebral malaria is a rare but often lethal complication of infection with Plasmodium parasites, the occurrence of which is influenced by complex genetic factors of the host. We used a mouse model of experimental cerebral malaria (ECM) with Plasmodium berghei ANKA to study genetic factors regulating appearance of neurological symptoms and associated lethality. In a genome-wide screen of N-ethyl-N-nitrosourea-mutagenized mice derived from C57BL/6J (B6) and 129S1/SvImJ (129) mouse strains, we detected a strong interaction between the genetic backgrounds of these strains, which modulates ECM resistance. We have mapped a major gene locus to central chromosome 4 (log of the odds (LOD) 6.7; 79.6-97.3 Mb), which we designate Berr8. [corrected]. B6 alleles at Berr6 are associated with resistance, and are inherited in a co-dominant fashion. In mice heterozygous for Berr6 B6/129 alleles, resistance to ECM is strongly modulated by a second locus, Berr7, that maps to the proximal portion of chromosome 1 (LOD 4.03; 41.4 Mb). 129 alleles at Berr7 are associated with ECM resistance in a dosage-dependent fashion. Results are discussed in view of the possible role of this two-locus system in susceptibility to unrelated inflammatory conditions in mice and humans.

Cerebral malaria (CM) is a lethal neurological complication of malaria. We implemented a genome-wide screen in mutagenized mice to identify host proteins involved in CM pathogenesis and whose inhibition may be of therapeutic value. One pedigree (P48) segregated a resistance trait whose CM-protective effect was fully penetrant, mapped to chromosome 8, and identified by genome sequencing as homozygosity for a mis-sense mutation (W81R) in the FERM domain of Janus-associated kinase 3 (Jak3). The causative effect of Jak3(W81R) was verified by complementation testing in Jak3(W81R/-) double heterozygotes that were fully protected against CM. Jak3(W81R) homozygotes showed defects in thymic development with depletion of CD8(+) T cell, B cell, and NK cell compartments, and defective T cell-dependent production of IFN-γ. Adoptive transfer of normal splenocytes abrogates CM resistance in Jak3(W81R) homozygotes, an effect attributed to the CD8(+) T cells. Jak3(W81R) behaves as a dominant negative variant, with significant CM resistance of Jak3(W81R/+) heterozygotes, compared to CM-susceptible Jak3(+/+) and Jak3(+/-) controls. CM resistance in Jak3(W81R/+) heterozygotes occurs in presence of normal T, B and NK cell numbers. These findings highlight the pathological role of CD8(+) T cells and Jak3-dependent IFN-γ-mediated Th1 responses in CM pathogenesis.

The E2 protein of human papillomavirus (HPV) binds to specific sites in the viral genome to regulate its transcription, replication, and maintenance in infected cells. Like most regulatory proteins, E2 is rapidly turned over. A high-throughput assay was developed to quantify the expression and stability of E2 in vivo, based on its fusion toRenillaluciferase (RLuc). The steady-state levels of Rluc-E2 were quantified by measuring the amounts of associated luciferase activity, and its degradation was measured by monitoring the decrease in enzymatic activity occurring after a block of translation with cycloheximide. Using this assay, the E2 proteins from a low-risk (HPV11) and a high-risk (HPV31) human papillomavirus (HPV) type were found to have short half-lives of 60 min in C33A cervical carcinoma cells and to be ubiquitinated and degraded by the proteasome. Analysis of mutant proteins showed that the instability of E2 is independent of its DNA-binding and transcriptional activities but is encoded within its transactivation domain, the region that binds to the cellular chromatin factor bromodomain-containing protein 4 (Brd4) to regulate viral gene transcription. Overexpression of Brd4, or of its C-terminal E2-interaction domain, was found to increase the steady-state levels and stability of wild-type E2 but not of E2 mutants defective for binding Brd4. These results indicate that the stability of E2 is increased upon complex formation with Brd4 and highlight the value of the luciferase assay for the study of E2 degradation.