Michel Hérin

Tip-DC Development during Parasitic Infection Is Regulated by IL-10 and Requires CCL2/CCR2, IFN-γ and MyD88 Signaling

The development of classically activated monocytic cells (M1) is a prerequisite for effective elimination of parasites, including African trypanosomes. However, persistent activation of M1 that produce pathogenic molecules such as TNF and NO contributes to the development of trypanosome infection-associated tissue injury including liver cell necrosis in experimental mouse models. Aiming to identify mechanisms involved in regulation of M1 activity, we have recently documented that during Trypanosoma brucei infection, CD11b+Ly6C+CD11c+ TNF and iNOS producing DCs (Tip-DCs) represent the major pathogenic M1 liver subpopulation. By using gene expression analyses, KO mice and cytokine neutralizing antibodies, we show here that the conversion of CD11b+Ly6C+ monocytic cells to pathogenic Tip-DCs in the liver of T. brucei infected mice consists of a three-step process including (i) a CCR2-dependent but CCR5- and Mif-independent step crucial for emigration of CD11b+Ly6C+ monocytic cells from the bone marrow but dispensable for their blood to liver migration; (ii) a differentiation step of liver CD11b+Ly6C+ monocytic cells to immature inflammatory DCs (CD11c+ but CD80/CD86/MHC-IIlow) which is IFN-γ and MyD88 signaling independent; and (iii) a maturation step of inflammatory DCs to functional (CD80/CD86/MHC-IIhigh) TNF and NO producing Tip-DCs which is IFN-γ and MyD88 signaling dependent. Moreover, IL-10 could limit CCR2-mediated egression of CD11b+Ly6C+ monocytic cells from the bone marrow by limiting Ccl2 expression by liver monocytic cells, as well as their differentiation and maturation to Tip-DCs in the liver, showing that IL-10 works at multiple levels to dampen Tip-DC mediated pathogenicity during T. brucei infection. A wide spectrum of liver diseases associates with alteration of monocyte recruitment, phenotype or function, which could be modulated by IL-10. Therefore, investigating the contribution of recruited monocytes to African trypanosome induced liver injury could potentially identify new targets to treat hepatic inflammation in general, and during parasite infection in particular.

IL-10 Dampens TNF/Inducible Nitric Oxide Synthase-Producing Dendritic Cell-Mediated Pathogenicity during Parasitic Infection

Antiparasite responses are associated with the recruitment of monocytes that differentiate to macrophages and dendritic cells at the site of infection. Although classically activated monocytic cells are assumed to be the major source of TNF and NO during Trypanosoma brucei brucei infection, their cellular origin remains unclear. In this study, we show that bone marrow-derived monocytes accumulate and differentiate to TNF/inducible NO synthase-producing dendritic cells (TIP-DCs) in the spleen, liver, and lymph nodes of T. brucei brucei-infected mice. Although TIP-DCs have been shown to play a beneficial role in the elimination of several intracellular pathogens, we report that TIP-DCs, as a major source of TNF and NO in inflamed organs, could contribute actively to tissue damage during the chronic stage of T. brucei brucei infection. In addition, the absence of IL-10 leads to enhanced differentiation of monocytes to TIP-DCs, resulting in exacerbated pathogenicity and early death of the host. Finally, we demonstrate that sustained production of IL-10 following IL-10 gene delivery treatment with an adeno-associated viral vector to chronically infected mice limits the differentiation of monocytes to TIP-DCs and protects the host from tissue damage.

Alternatively Activated Myeloid Cells Limit Pathogenicity Associated with African Trypanosomiasis through the IL-10 Inducible Gene Selenoprotein P

Uncontrolled inflammation is a major cause of tissue injury/pathogenicity often resulting in death of a host infected with African trypanosomes. Thus, comparing the immune response in hosts that develop different degrees of disease severity represents a promising approach to discover processes contributing to trypanosomiasis control. It is known that limitation of pathogenicity requires a transition in the course of infection, from an IFN-γ-dependent response resulting in the development of classically activated myeloid cells (M1), to a counterbalancing IL-10-dependent response associated with alternatively activated myeloid cells (M2). Herein, mechanisms and downstream effectors by which M2 contribute to lower the pathogenicity and the associated susceptibility to African trypanosomiasis have been explored. Gene expression analysis in IL-10 knockout and wild-type mice, that are susceptible and relatively resistant to Trypanosoma congolense infection, respectively, revealed a number of IL-10-inducible genes expressed by M2, including Sepp1 coding for selenoprotein P. Functional analyses confirm that selenoprotein P contributes to limit disease severity through anti-oxidant activity. Indeed, Sepp1 knockout mice, but not Sepp1Δ240-361 mice retaining the anti-oxidant motif but lacking the selenium transporter domain of selenoprotein P, exhibited increased tissue injury that associated with increased production of reactive oxygen species and increased apoptosis in the liver immune cells, reduced parasite clearance capacity of myeloid cells, and decreased survival. These data validate M2-associated molecules as functioning in reducing the impact of parasite infection on the host.

African Trypanosomiasis: Naturally Occurring Regulatory T Cells Favor Trypanotolerance by Limiting Pathology Associated with Sustained Type 1 Inflammation

Tolerance to African trypanosomes requires the production of IFN-γ in the early stage of infection that triggers the development of classically activated macrophages controlling parasite growth. However, once the first peak of parasitemia has been controlled, down-regulation of the type 1 immune response has been described. In this study, we have evaluated whether regulatory T cells (Tregs) contribute to the limitation of the immune response occurring during Trypanosoma congolense infection and hereby influence the outcome of the disease in trypanotolerant C57BL/6 host. Our data show that Foxp3+ Tregs originating from the naturally occurring Treg pool expanded in the spleen and the liver of infected mice. These cells produced IL-10 and limited the production of IFN-γ by CD4+ and CD8+ effector T cells. Tregs also down-regulated classical activation of macrophages resulting in reduced TNF-α production. The Treg-mediated suppression of the type 1 inflammatory immune response did not hamper parasite clearance, but was beneficial for the host survival by limiting the tissue damages, including liver injury. Collectively, these data suggest a cardinal role for naturally occurring Tregs in the development of a trypanotolerant phenotype during African trypanosomiasis.

Experimental expansion of the regulatory T cell population increases resistance to African trypanosomiasis.

Inflammatory responses mounted to eliminate parasites can be lethal if not counterbalanced by regulatory responses protecting the host from collateral tissue damage. Here, we show that the maintained inflammation associated with tissue damage, anemia, and reduced survival of Trypanosoma brucei-infected mice correlates with the absence of the expansion of the regulatory T (T(reg)) cell population. Induction of T(reg) cell expansion via CD28 superagonist antibody treatment in these mice down-regulated interferon-gamma production by T cells and tumor necrosis factor-alpha and reactive oxygen species production by classically activated macrophages, triggered the development of alternatively activated macrophages, delayed the onset of liver injury, diminished the anemia burden, and prolonged the survival of infected animals. Thus, triggering the expansion of the T(reg) cell population coupled with the induction of alternatively activated macrophages can restore the balance between pro- and anti-inflammatory signals and thereby limit the pathogenicity of African trypanosomiasis.

A Trypanosoma brucei Kinesin Heavy Chain Promotes Parasite Growth by Triggering Host Arginase Activity

Background In order to promote infection, the blood-borne parasite Trypanosoma brucei releases factors that upregulate arginase expression and activity in myeloid cells. Methodology/Principal findings By screening a cDNA library of T. brucei with an antibody neutralizing the arginase-inducing activity of parasite released factors, we identified a Kinesin Heavy Chain isoform, termed TbKHC1, as responsible for this effect. Following interaction with mouse myeloid cells, natural or recombinant TbKHC1 triggered SIGN-R1 receptor-dependent induction of IL-10 production, resulting in arginase-1 activation concomitant with reduction of nitric oxide (NO) synthase activity. This TbKHC1 activity was IL-4Rα-independent and did not mirror M2 activation of myeloid cells. As compared to wild-type T. brucei, infection by TbKHC1 KO parasites was characterized by strongly reduced parasitaemia and prolonged host survival time. By treating infected mice with ornithine or with NO synthase inhibitor, we observed that during the first wave of parasitaemia the parasite growth-promoting effect of TbKHC1-mediated arginase activation resulted more from increased polyamine production than from reduction of NO synthesis. In late stage infection, TbKHC1-mediated reduction of NO synthesis appeared to contribute to liver damage linked to shortening of host survival time. Conclusion A kinesin heavy chain released by T. brucei induces IL-10 and arginase-1 through SIGN-R1 signaling in myeloid cells, which promotes early trypanosome growth and favors parasite settlement in the host. Moreover, in the late stage of infection, the inhibition of NO synthesis by TbKHC1 contributes to liver pathogenicity.

IL-10 limits production of pathogenic TNF by M1 myeloid cells through induction of nuclear NF-κB p50 member in Trypanosoma congolense infection-resistant C57BL/6 mice

A balance between parasite elimination and control of infection‐associated pathogenicity is crucial for resistance to African trypanosomiasis. By producing TNF and NO, CD11b+ myeloid cells with a classical activation status (M1) contribute to parasitemia control in experimental Trypanosoma congolense infection in resistant C57BL/6 mice. However, in these mice, IL‐10 is required to regulate M1‐associated inflammation, avoiding tissue/liver damage and ensuring prolonged survival. In an effort to dissect the mechanisms behind the anti‐inflammatory activity of IL‐10 in T. congolense‐infected C57BL/6 mice, we show, using an antibody blocking the IL‐10 receptor, that IL‐10 impairs the accumulation and M1 activation of TNF/iNOS‐producing CD11b+Ly6C+ cells in the liver. Using infected IL‐10flox/floxLysM‐Cre+/+ mice, we show that myeloid cell‐derived IL‐10 limits M1 activation of CD11b+Ly6C+ cells specifically at the level of TNF production. Moreover, higher production of TNF in infected IL‐10flox/floxLysM‐Cre+/+ mice is associated with reduced nuclear accumulation of the NF‐κB p50 subunit in CD11b+ M1 cells. Furthermore, in infected p50−/− mice, TNF production by CD11b+Ly6C+ cells and liver injury increases. These data suggest that preferential nuclear accumulation of p50 represents an IL‐10‐dependent anti‐inflammatory mechanism in M1‐type CD11b+ myeloid cells that regulates the production of pathogenic TNF during T. congolense infection in resistant C57BL/6 mice.