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Protein-energy malnutrition (PEM) is a risk factor for developing visceral leishmaniasis (VL). However, the impact on adaptive immunity during infection is unknown. To study the effect of malnutrition on chronic VL, we used a polynutrient-deficient diet (deficient protein, energy, zinc, and iron), which mimics moderate human malnutrition, followed by Leishmania infantum infection. The polynutrient-deficient diet leads to growth stunting and reduced mass of visceral organs. Malnourished-infected mice harbored more parasites in the spleen and liver, had a reduced number of T lymphocytes, reduced production of IFN-γ by T cells, and exhibited enhanced IL-10 production. To test whether IL-10 blockade would lessen disease in the malnourished mice, we treated infected mice with monoclonal antibody α-IL-10R. α-IL-10R treatment reduced the parasite number of malnourished mice, restored the number of T cells producing IFN-γ, and enhanced hepatic granuloma formation. Our results indicate that malnutrition increases VL susceptibility due to a defective IFN-γ-mediated immunity attributable to increased IL-10 production., Author Summary: Malnutrition contributes to the development of VL. Despite the advances regarding this association, how malnutrition affects the adaptive immune mechanisms in VL is still unclear. We found that malnutrition disrupts the ability to control parasite replication in the spleen and liver in VL due to defective IFN-γ-mediated immunity, reduced hepatic granuloma formation, and enhanced IL-10 production. Blocking IL-10R signaling restored the protective mechanisms to control parasite replication in the malnourished mice without interfering with the undernutrition state. Thus, we demonstrate that malnutrition disrupts the adaptive immunity against VL due to an aberrant IL-10 production. Understanding the association between malnutrition and VL will provide insights into therapeutic approaches.

In many infectious diseases, the pathogen-induced inflammatory response could result in protective immunity that should be regulated to prevent tissue damage and death. In fact, in Trypanosoma cruzi infection, the innate immune and the inflammatory response should be perfectly controlled to avoid significant lesions and death. Here, we investigate the role of Blimp-1 expression in T cells in resistance to T. cruzi infection. Therefore, using mice with Blimp-1 deficiency in T cells (CKO) we determined its role in the controlling parasites growth and lesions during the acute phase of infection. Infection of mice with Blimp-1 ablation in T cells resulted failure the cytotoxic CD8 + T cells and in marked Th1-mediated inflammation, high IFN-γ and TNF production, and activation of inflammatory monocyte. Interestingly, despite high nitric-oxide synthase activation (NOS-2), parasitemia and mortality in CKO mice were increased compared with infected WT mice. Furthermore, infected-CKO mice exhibited hepatic lesions characteristic of steatosis, with significant AST and ALT activity. Mechanistically, Blimp-1 signaling in T cells induces cytotoxic CD8 + T cell activation and restricts parasite replication. In contrast, Blimp-1 represses the Th1 response, leading to a decreased monocyte activation, less NOS-2 activation, and, consequently preventing hepatic damage and dysfunction. These data demonstrate that T. cruzi -induced disease is multifactorial and that the increased IFN-γ, NO production, and dysfunction of CD8 + T cells contribute to host death. These findings have important implications for the design of potential vaccines against Chagas disease., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Benevides, Sacramento, Pioto, Barretto, Carregaro and Silva.)

Cutaneous leishmaniasis exhibits a spectrum of clinical presentations dependent upon the parasites' persistence and host immunopathologic responses. Although cytolytic CD8 T cells cannot control the parasites, they significantly contribute to pathologic responses. In a murine model of cutaneous leishmaniasis, we previously found that NKG2D plays a role in the ability of cytolytic CD8 T cells to promote disease in leishmanial lesions. Here, we investigated whether NKG2D plays a role in human disease. We found that NKG2D and its ligands were expressed within lesions from L. braziliensis-infected patients and that IL-15 and IL-1β were factors driving NKG2D and NKG2D ligand expression, respectively. Blocking NKG2D reduced degranulation by CD8 T cells in a subset of patients. Additionally, our transcriptional analysis of patients' lesions found that patients who failed the first round of treatment exhibited higher expression of KLRK1, the gene coding for NKG2D, than those who responded to treatment. These findings suggest that NKG2D may be a promising therapeutic target for ameliorating disease severity in cutaneous leishmaniasis caused by L. braziliensis infection., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Sacramento et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Objective: To evaluate the clinical efficacy of a mixture of probiotics (Lactobacillus and Bifidobacterium) in children and adolescents with atopic dermatitis (AD) and the effects on sensitization, inflammation, and immunological tolerance., Methods: In this double-blind, randomized, placebo-controlled clinical trial, we enrolled 60 patients aged between 6 months and 19 years with mild, moderate, or severe AD, according to the criteria proposed by Hanifin and Rajka. Patients were stratified to receive one gram per day of probiotics or placebo for 6 months. The primary outcome was a decrease in SCORing Atopic Dermatitis (SCORAD). Secondary outcomes were to assess the role of probiotics on the use of topical and oral medicines (standard treatment), serum IgE levels, skin prick test (SPT), and tolerogenic and inflammatory cytokines. Background therapy was maintained., Results: Forty patients completed the study (24 probiotics, 16 placebo). After treatment for six months, the clinical response was significantly better in the probiotics group; the SCORAD decreased [mean difference (MD) 27.69 percentage points; 95% confidence interval (CI), 2.44-52.94], even after adjustment for co-variables (MD 32.33 percentage points; 95%CI, 5.52-59.13), especially from the third month of treatment on. The reduction of the SCORAD in probiotic group persisted for three more months after the treatment had been discontinued, even after adjustment for co-variables (MD 14.24 percentage points; 95%CI, 0.78-27.70). Patients in the probiotics group required topical immunosuppressant less frequently at 6 and 9 months. No significant changes were found for IgE levels, SPT and cytokines., Conclusions: Children and adolescents with AD presented a significant clinical response after 6 months with a mixture of probiotics (Lactobacillus rhamnosus, Lactobacillus acidophilus, Lactobacillus paracasei, and Bifidobacterium lactis. However, this clinical benefit is related to treatment duration. Probiotics should be considered as an adjuvant treatment for AD., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Andrade, Maria e Silva, Carregaro, Sacramento, Roberti, Aragon, Carmona and Roxo-Junior.)

Innate lymphoid cells (ILCs) comprise a heterogeneous population of immune cells that maintain barrier function and can initiate a protective or pathological immune response upon infection. Here we show the involvement of IL-17A-producing ILCs in microbiota-driven immunopathology in cutaneous leishmaniasis. IL-17A-producing ILCs were RORγt+ and were enriched in Leishmania major infected skin, and topical colonization with Staphylococcus epidermidis before L. major infection exacerbated the skin inflammatory responses and IL-17A-producing RORγt+ ILC accumulation without impacting type 1 immune responses. IL-17A responses in ILCs were directed by Batf3 dependent CD103+ dendritic cells and IL-23. Moreover, experiments using Rag1-/- mice established that IL-17A+ ILCs were sufficient in driving the inflammatory responses as depletion of ILCs or neutralization of IL-17A diminished the microbiota mediated immunopathology. Taken together, this study indicates that the skin microbiota promotes RORγt+ IL-17A-producing ILCs, which augment the skin inflammation in cutaneous leishmaniasis., Competing Interests: The authors have declared that no competing interests exist.

Premise: Genera that are widespread, with geographically discontinuous distributions and represented by few species, are intriguing. Is their achieved disjunct distribution recent or ancient in origin? Why are they species-poor? The Rand Flora is a continental-scale pattern in which closely related species appear codistributed in isolated regions over the continental margins of Africa. Genus Camptoloma (Scrophulariaceae) is the most notable example, comprising three species isolated from each other on the northwest, eastern, and southwest Africa., Methods: We employed Sanger sequencing of nuclear and plastid markers, together with genomic target sequencing of 2190 low-copy nuclear genes, to infer interspecies relationships and the position of Camptoloma within Scrophulariaceae by using supermatrix and multispecies-coalescent approaches. Lineage divergence times and ancestral ranges were inferred with Bayesian Markov chain Monte Carlo (MCMC) approaches. The population history was estimated with phylogeographic coalescent methods., Results: Camptoloma rotundifolium, restricted to Southern Africa, was shown to be a sister species to the disjunct clade formed by C. canariense, endemic to the Canary Islands, and C. lyperiiflorum, distributed in the Horn of Africa-Southern Arabia. Camptoloma was inferred to be sister to the mostly South African tribes Teedieae and Buddlejeae. Stem divergence was dated in the Late Miocene, while the origin of the extant disjunction was inferred as Early Pliocene., Conclusions: The current disjunct distribution of Camptoloma across Africa was likely the result of fragmentation and extinction and/or population bottlenecking events associated with historical aridification cycles during the Neogene; the pattern of species divergence, from south to north, is consistent with the "climatic refugia" Rand Flora hypothesis., (© 2021 The Authors. American Journal of Botany published by Wiley Periodicals LLC on behalf of Botanical Society of America.)

A striking feature of human visceral leishmaniasis (VL) is chronic inflammation in the spleen and liver, and VL patients present increased production levels of multiple inflammatory mediators, which contribute to tissue damage and disease severity. Here, we combined an experimental model with the transcriptional profile of human VL to demonstrate that the TLR4-IFN-β pathway regulates the chronic inflammatory process and is associated with the asymptomatic form of the disease. Tlr4-deficient mice harbored fewer parasites in their spleen and liver than wild-type mice. TLR4 deficiency enhanced the Th1 immune response against the parasite, which was correlated with an increased activation of dendritic cells (DCs). Gene expression analyses demonstrated that IRF1 and IFN-β were expressed downstream of TLR4 after infection. Accordingly, IRF1- and IFNAR-deficient mice harbored fewer parasites in the target organs than wild-type mice due to having an increased Th1 immune response. However, the absence of TLR4 or IFNAR increased the serum transaminase levels in infected mice, indicating the presence of liver damage in these animals. In addition, IFN-β limits IFN-γ production by acting directly on Th1 cells. Using RNA sequencing analysis of human samples, we demonstrated that the transcriptional signature for the TLR4 and type I IFN (IFN-I) pathways was positively modulated in asymptomatic subjects compared with VL patients and thus provide direct evidence demonstrating that the TLR4-IFN-I pathway is related to the nondevelopment of the disease. In conclusion, our results demonstrate that the TLR4-IRF1 pathway culminates in IFN-β production as a mechanism for dampening the chronic inflammatory process and preventing immunopathology development., Competing Interests: The authors have declared that no competing interests exist.

Monocytes play key roles in the maintenance of homeostasis and in the control of the infection. Monocytes are recruited from the bone marrow to inflammatory sites and are essential for antimicrobial activity to limit tissue damage and promote adaptive T cell responses. Here, we investigated the role of Nuclear Factor of Activated T cells 1 (NFAT1) in the regulation of Ly6C hi inflammatory monocyte recruitment to the CNS upon T. gondii infection. We show that NFAT-1-deficient monocytes are unable to migrate to the CNS of T. gondii -infected mice. Moreover, NFAT1 -/- mice are highly susceptible to chronic T. gondii infection due to a failure to control parasite replication in the CNS. The inhibition of Ly6C hi inflammatory monocyte recruitment to the CNS severely blocked CXCL10 production and consequently the migration of IFN-γ-producing CD4 + T cells. Moreover, the transfer of Ly6C hi monocytes to infected NFAT1 -/- mice favored CD4 + T cell migration to the CNS and resulted in the inhibition of parasite replication and host defense. Together, these results demonstrated for the first time the contribution of NFAT1 to the regulation of Ly6C hi monocyte recruitment to the CNS and to resistance during chronic T. gondii infection.

T1/ST2 is a surface marker selectively expressed on type 2 helper (T H 2) effector cells. As Leishmania infection in susceptible BALB/c mice have ascribed to a polarized T H 2 response, this study aim to investigate the T1/ST2 (the receptor for IL-33), as a typical T H 2 marker in the postulation that a shift towards a beneficial T H 1 response would occur in the absence of ST2. For this, ST2 knockout (ST2 -/- ) and WT BALB/c mice were experimentally infected in the retro-orbital sinus with L. infantum. We showed that ST2 -/- animals displayed better control of parasite burden in both spleen and liver tissues at different time points of chronic phases, and reduced spleenomegaly and hepatomegaly compared with the wild-type (WT) mice. This was associated with increased in the IFN-γ levels and expression by CD4 + and CD8 + lymphocytes. The inflammatory response encompasses transaminases (AST and ALT) releases and NO productions were remarkably lower in ST2 -/- mice compared with WT. These data suggest that, ST2 -/- ) exert protection against L. infantum infection and probably shift the immune response toward T H 1 induction., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Adenosine is an endogenously released purine nucleoside that signals through four widely expressed G protein-coupled receptors: A1, A2 A , A2 B , and A3. Of these, A 2A R is recognized as mediating major adenosine anti-inflammatory activity. During cutaneous leishmaniasis, adenosine induces immunosuppression, which promotes the establishment of infection. Herein, we demonstrated that A 2A R signaling is exploited by Leishmania infantum parasites, the etiologic agent that causes Visceral Leishmaniasis, to successfully colonize the vertebrate host. A 2A R gene-deleted mice exhibited a well-developed cellular reaction with a strong Th1 immune response in the parasitized organs. An intense infiltration of activated neutrophils into the disease-target organs was observed in A 2A R -/- mice. These cells were characterized by high expression of CXCR2 and CD69 on their cell surfaces and increased cxcl1 expression. Interestingly, this phenotype was mediated by IFN-γ on the basis that a neutralizing antibody specific to this cytokine prevented neutrophilic influx into parasitized organs. In evaluating the immunosuppressive effects, we identified a decreased number of CD4 + FOXP3 + T cells and reduced il10 expression in A 2A R -/- infected mice. During ex vivo cell culture, A 2A R -/- splenocytes produced smaller amounts of IL-10. In conclusion, we demonstrated that the A 2A R signaling pathway is detrimental to development of Th1-type adaptive immunity and that this pathway could be associated with the regulatory process. In particular, it promotes parasite surveillance.

Visceral leishmaniasis (VL) is a chronic and fatal disease caused by Leishmania infantum in Brazil. Leukocyte recruitment to infected tissue is a crucial event for the control of infections such as VL. Among inflammatory cells, neutrophils are recruited to the site of Leishmania infection, and these cells may control parasite replication through oxidative or non-oxidative mechanisms. The recruitment, activation and functions of the neutrophils are coordinated by pro-inflammatory cytokines and chemokines during recognition of the parasite by pattern recognition receptors (PRRs). Here, we demonstrated that the Toll-like receptor 2 (TLR2) signaling pathway contributes to the development of the innate immune response during L. infantum infection. The protective mechanism is related to the appropriate recruitment of neutrophils to the inflammatory site. Neutrophil migration is coordinated by DCs that produce CXCL1 and provide a prototypal Th1 and Th17 environment when activated via TLR2. Furthermore, infected TLR2 -/- mice failed to induce nitric oxide synthase (iNOS) expression in neutrophils but not in macrophages. In vitro , infected TLR2 -/- neutrophils presented deficient iNOS expression, nitric oxide (NO) and TNF-α production, decreased expression of CD11b and reduced L. infantum uptake capacity. The non-responsive state of neutrophils is associated with increased amounts of IL-10. Taken together, these data clarify new mechanisms by which TLR2 functions in promoting the development of the adaptive immune response and effector mechanisms of neutrophils during L. infantum infection.

In vertebrate hosts, Leishmania braziliensis parasites infect mainly mononuclear phagocytic system cells, which when activated by T helper cell type 1 cytokines produce nitric oxide and kill the pathogens. Chemokine (C-C motif) receptor 2 is a chemokine receptor that binds primarily chemokine (C-C motif) ligand 2 and has an important role in the recruitment of monocytic phagocytes. Although it has been reported that Leishmania braziliensis infection induces CCR2 expression in the lesions, the role of CCR2 during Leishmania braziliensis infection remains unknown. Here, we showed that CCR2 has a role in mediating protection against Leishmania braziliensis infection in mice. The absence of CCR2 resulted in increased susceptibility to infection and was associated with low amounts of Ly6C(+) inflammatory dendritic cells in the lesions, which we found to be the major sources of tumor necrosis factor production and induced nitric oxide synthase expression in C57BL/6 mice lesions. Consequently, CCR2(-/-) mice showed decreased tumor necrosis factor production and induced nitric oxide synthase expression, resulting in impaired parasite elimination. We also demonstrated that CCR2 has a role in directly mediating the differentiation of monocytes into inflammatory dendritic cells at the infection sites, contributing to the accumulation of inflammatory dendritic cells in Leishmania braziliensis lesions and subsequent control of parasite replication. Therefore, these data provide new information on the role of chemokines during the immune response to infections and identify a potential target for therapeutic interventions in cutaneous leishmaniasis., (© Society for Leukocyte Biology.)

The relationship established between Leishmania infantum and the vertebrate host can lead to a self-healing infection or to the manifestation of visceral leishmaniasis, a chronic systemic infection associated with high rates of mortality. We hypothesized that regulatory cytokines, such as interleukin-27 (IL-27), play a role in susceptibility to L. infantum infection. IL-27 is a heterodimeric cytokine composed of IL-27p28 and EBi3 subunits which, when combined, bind to IL-27R, leading to STAT-1 and -3 activation, playing a role in the regulation of the immune response. We observed in this work that IL-27 regulates the Th1/Th17 profiles in a mouse model of visceral leishmaniasis (VL) caused by L. infantum We showed here that the pathogen recognition by endosomal Toll-like receptors triggers a type I interferon (IFN) response, which acts through the type I IFN receptor and interferon regulatory factor 1 to induce IL-27 production by macrophages. Furthermore, IL-27 plays a major regulatory role in vivo, because Ebi3(-/-) mice can efficiently control parasite replication despite reduced levels of IFN-γ compared to wild-type mice. On the other hand, the absence of Ebi3 leads to exacerbated IL-17A production in the infected organs as well as in a coculture system, suggesting a direct regulatory action of IL-27 during L. infantum infection. As a consequence of exacerbated IL-17A in Ebi3(-/-) mice, a greater neutrophil influx was observed in the target organs, playing a role in parasite control. Thus, this work unveiled the molecular steps of IL-27 production after L. infantum infection and demonstrated its regulatory role in the IL-17A-neutrophil axis., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)

A novel T cell-secreted cytokine, termed secreted osteoclastogenic factor of activated T cells (SOFAT) that induces osteoclastic bone resorption in a RANKL-independent manner, has been described. Our group have previously reported that SOFAT is highly expressed in gingival tissues of patients with chronic periodontitis suggesting a putative role in the bone loss associated with periodontal disease. The aim of the present study was to identify other potential cellular sources of SOFAT in the bone resorptive lesions of patients with periodontal disease. Gingival tissues were biopsied from systemically healthy subjects without periodontal disease (n=5) and patients with chronic periodontitis (n=5), and the presence of SOFAT was analyzed by immunohistochemistry and immunofluorescence staining. The present data demonstrated marked SOFAT staining in diseased periodontal tissues that was predominantly associated with the lymphocytic infiltration of gingival tissues. Notably, in addition to CD3+ T cells, B‑lineage cells including plasma cells also exhibited strong staining for SOFAT. As SOFAT has not previously been reported in B‑lineage cells, splenic T cells and B cells were further purified from BALB/c mice and activated using CD3/CD28 and lipopolysaccharide, respectively. SOFAT was quantified by reverse transcription‑quantitative polymerase chain reaction and was shown to be significantly expressed (P<0.05) in both activated T cells and B cells compared with unstimulated cells. These data support a putative role of SOFAT in the bone loss associated with chronic periodontal disease. In addition, to the best of our knowledge, this study demonstrates for the first time that in addition to T cells, B-lineage cells may also be a significant source of SOFAT in inflammatory states.

The prostaglandin, 15-deoxy Δ 12,14 -prostaglandin J 2 (15d-PGJ 2 ), is a lipid mediator that plays an important role in the control of chronic inflammatory disease. However, the role of prostanoid in rheumatoid arthritis (RA) is not well determined. We demonstrated the therapeutic effect of 15d-PGJ 2 in an experimental model of arthritis. Daily administration of 15d-PGJ 2 attenuated the severity of CIA, reducing the clinical score, pain, and edema. 15d-PGJ 2 treatment was associated with a marked reduction in joint levels of proinflammatory cytokines. Although the mRNA expression of ROR- γ t was profoundly reduced, FOXP3 was enhanced in draining lymph node cells from 15d-PGJ 2 -treated arthritic mice. The specific and polyclonal CD4 + Th17 cell responses were limited during the addition of prostaglandin to cell culture. Moreover, in vitro 15d-PGJ 2 increased the expression of FOXP3, GITR, and CTLA-4 in the CD4 + CD25 - population, suggesting the induction of Tregs on conventional T cells. Prostanoid addition to CD4 + CD25 - cells selectively suppressed Th17 differentiation and promoted the enhancement of FOXP3 under polarization conditions. Thus, 15d-PGJ 2 ameliorated symptoms of collagen-induced arthritis by regulating Th17 differentiation, concomitant with the induction of Tregs, and, consequently, protected mice from diseases aggravation. Altogether, these results indicate that 15d-PGJ 2 may represent a potential therapeutic strategy in RA.

Background: Sand fly saliva plays a crucial role in establishing Leishmania infection. We identified adenosine (ADO) and adenosine monophosphate (AMP) as active pharmacologic compounds present in Phlebotomus papatasi saliva that inhibit dendritic cell (DC) functions through a PGE2/IL 10-dependent mechanism., Methodology/principal Findings: Herein, we prepared a mixture of ADO and AMP in equimolar amounts similar to those present in the salivary-gland extract (SGE) form one pair of salivary glands of P. papatasi and co-injected it with Leishmania amazonensis or L. major into mouse ears. ADO+AMP mimicked exacerbative effects of P. papatasi saliva in leishmaniasis, increasing parasite burden and cutaneous lesions. Enzymatic catabolism of salivary nucleosides reversed the SGE-induced immunosuppressive effect associated with IL-10 enhancement. Immunosuppressive factors COX2 and IL-10 were upregulated and failed to enhance ear lesion and parasite burden in IL 10-/- infected mice. Furthermore, nucleosides increased regulatory T cell (Treg) marker expression on CD4+CD25- cells, suggesting induction of Tregs on effector T cells (T eff). Treg induction (iTreg) was associated with nucleoside-induced tolerogenic dendritic cells (tDCs) expressing higher levels of COX2 and IL-10. In vitro generation of Tregs was more efficient in DCs treated with nucleosides. Suppressive effects of nucleosides during cutaneous leishmaniasis were mediated through an A2AR-dependent mechanism. Using BALB/c mice deficient in A2A ADO receptor (A2AR-/-), we showed that co-inoculated mice controlled infection, displaying lower parasite numbers at infection sites and reduced iTreg generation., Conclusion/significance: We have demonstrated that ADO and AMP in P. papatasi saliva mediate exacerbative effects of Leishmania infection by acting preferentially on DCs promoting a tolerogenic profile in DCs and by generating iTregs in inflammatory foci through an A2AR mechanism.

Visceral leishmaniasis (VL) is a chronic and fatal disease caused by Leishmania infantum in Brazil. Leukocyte recruitment to infected tissue is a crucial event for the control of infections such as VL. Leucotriens are lipid mediators synthesized by 5-lipoxygenase (5-LO) and they display a protective role against protozoan parasites by inducing several functions in leucocytes. We determined the role of 5-LO activity in parasite control, focusing on the inflammatory immune response against Leishmania infantum infection. LTB4 is released during in vitro infection. The genetic ablation of 5-LO promoted susceptibility in highly resistant mice strains, harboring more parasites into target organs. The susceptibility was related to the failure of neutrophil migration to the infectious foci. Investigating the neutrophil failure, there was a reduction of proinflammatory cytokines involved in the related Th17 axis released into the organs. Genetic ablation of 5-LO reduced the CD4(+)T cells producing IL-17, without interfering in Th1 subset. L. infantum failed to activate DC from 5-LO(-/-), showing reduced surface costimulatory molecule expression and proinflammatory cytokines involved in Th17 differentiation. BLT1 blockage with selective antagonist interferes with DC maturation and proinflammatory cytokines release. Thus, 5-LO activation coordinates the inflammatory immune response involved in the control of VL.