Persistent p55TNFR expression impairs T cell responses during chronic tuberculosis and promotes reactivation

Acknowledgements We thank Lizette Fick for her contribution to histopathology. We thank Faried Abbass for technical support. We thank the support staff of the Division of Immunology and the Research Animal Facility at the University of Cape Town for their contribution to animal care and technical support. The study was supported by the University of Cape Town, National Research Foundation (South Africa), South African Medical Research Council (SAMRC) National Health Laboratory Service (South Africa), The European Union (contract number: 028190), FP6 NEST project N°028190 “TB REACT”. Research carried out within the scope of the Franco/South African Laboratory “TB Immunity” (Associated International Laboratory ‘AIL’).Peer reviewedPublisher PD

CD14 Works with Toll-Like Receptor 2 to Contribute to Recognition and Control of Listeria monocytogenes Infection

Toll-like receptor 2 (TLR2) signaling has been shown to contribute to resistance to Listeria monocytogenes infection, as TLR2-deficient mice have a heightened susceptibility to infection with this organism. Because CD14 may associate with TLR2, we investigated the role of CD14 in Listeria responses. In both CD14-deficient and TLR2- deficient macrophages, nuclear factor κB translocation; CD40 and CD86; and the production of interleukin (IL)- 12, IL-6, tumor necrosis factor, and nitric oxide are reduced. The absence of CD14 augmented susceptibility to Listeria infection, reduced survival, and diminished bacterial clearance, as observed in TLR2-deficient mice.Compared with C57BL/6 control mice, CD14-deficient mice were observed to have a greater number of hepatic microabscesses containing abundant neutrophils, these abscesses were larger in size, and there was reduced inducible nitric oxide synthase expression. Further, mice that are both CD14 deficient and TLR2 deficient display susceptibility to infection that is comparable to that of mice deficient in either CD14 or TLR2 alone. Therefore, the present data demonstrate the role of CD14 and TLR2 in the recognition and control of Listeria infection and host resistanc

Fatal Mycobacterium tuberculosis infection despite adaptive immune response in the absence of MyD88

Toll-like receptors (TLRs) such as TLR2 and TLR4 have been implicated in host response to mycobacterial infection. Here, mice deficient in the TLR adaptor molecule myeloid differentiation factor 88 (MyD88) were infected with Mycobacterium tuberculosis (MTB). While primary MyD88–/– macrophages and DCs are defective in TNF, IL-12, and NO production in response to mycobacterial stimulation, the upregulation of costimulatory molecules CD40 and CD86 is unaffected. Aerogenic infection of MyD88–/– mice with MTB is lethal within 4 weeks with 2 log10 higher CFU in the lung; high pulmonary levels of cytokines and chemokines; and acute, necrotic pneumonia, despite a normal T cell response with IFN-γ production to mycobacterial antigens upon ex vivo restimulation. Vaccination with Mycobacterium bovis bacillus Calmette-Guérin conferred a substantial protection in MyD88–/– mice from acute MTB infection. These data demonstrate that MyD88 signaling is dispensable to raise an acquired immune response to MTB. Nonetheless, this acquired immune response is not sufficient to compensate for the profound innate immune defect and the inability of MyD88–/– mice to control MTB infection

Natterin-Induced Neutrophilia Is Dependent on cGAS/STING Activation via Type I IFN Signaling Pathway

Natterin is a potent pro-inflammatory fish molecule, inducing local and systemic IL-1β/IL-1R1-dependent neutrophilia mediated by non-canonical NLRP6 and NLRC4 inflammasome activation in mice, independent of NLRP3. In this work, we investigated whether Natterin activates mitochondrial damage, resulting in self-DNA leaks into the cytosol, and whether the DNA sensor cGAS and STING pathway participate in triggering the innate immune response. Employing a peritonitis mouse model, we found that the deficiency of the tlr2/tlr4, myd88 and trif results in decreased neutrophil influx to peritoneal cavities of mice, indicative that in addition to MyD88, TRIF contributes to neutrophilia triggered by TLR4 engagement by Natterin. Next, we demonstrated that gpcr91 deficiency in mice abolished the neutrophil recruitment after Natterin injection, but mice pre-treated with 2-deoxy-d-glucose that blocks glycolysis presented similar infiltration than WT Natterin-injected mice. In addition, we observed that, compared with the WT Natterin-injected mice, DPI and cyclosporin A treated mice had a lower number of neutrophils in the peritoneal exudate. The levels of dsDNA in the supernatant of the peritoneal exudate and processed IL-33 in the supernatant of the peritoneal exudate or cytoplasmic supernatant of the peritoneal cell lysate of WT Natterin-injected mice were several folds higher than those of the control mice. The recruitment of neutrophils to peritoneal cavity 2 h post-Natterin injection was intensely impaired in ifnar KO mice and partially in il-28r KO mice, but not in ifnγr KO mice. Finally, using cgas KO, sting KO, or irf3 KO mice we found that recruitment of neutrophils to peritoneal cavities was virtually abolished in response to Natterin. These findings reveal cytosolic DNA sensors as critical regulators for Natterin-induced neutrophilia

Pre-conception maternal helminth infection transfers via nursing long-lasting cellular immunity against helminths to offspring

Maternal immune transfer is the most significant source of protection from early-life infection, but whether maternal transfer of immunity by nursing permanently alters offspring immunity is poorly understood. Here, we identify maternal immune imprinting of offspring nursed by mothers who had a pre-conception helminth infection. Nursing of pups by helminth-exposed mothers transferred protective cellular immunity to these offspring against helminth infection. Enhanced control of infection was not dependent on maternal antibody. Protection associated with systemic development of protective type 2 immunity in T helper 2 (TH2) impaired IL-4R-/- offspring. This maternally acquired immunity was maintained into maturity and required transfer (via nursing) to the offspring of maternally derived TH2-competent CD4 T cells. Our data therefore reveal that maternal exposure to a globally prevalent source of infection before pregnancy provides long-term nursing-acquired immune benefits to offspring mediated by maternally derived pathogen-experienced lymphocytes. © 2019 by the Authors

GM-CSF Priming Drives Bone Marrow-Derived Macrophages to a Pro-Inflammatory Pattern and Downmodulates PGE(2) in Response to TLR2 Ligands

In response to pathogen recognition by Toll-like receptors (TLRs) on their cell surface, macrophages release lipid mediators and cytokines that are widely distributed throughout the body and play essential roles in host responses. Granulocyte macrophage colony-stimulating factor (GM-CSF) is important for the immune response during infections to improve the clearance of microorganisms. In this study, we examined the release of mediators in response to TLR2 ligands by bone marrow-derived macrophages (BMDMs) primed with GM-CSF. We demonstrated that when stimulated with TLR2 ligands, non-primed BMDMs preferentially produced PGE(2) in greater amounts than LTB4. However, GM-CSF priming shifted the release of lipid mediators by BMDMs, resulting in a significant decrease of PGE(2) production in response to the same stimuli. The decrease of PGE(2) production from primed BMDMs was accompanied by a decrease in PGE-synthase mRNA expression and an increase in TNF-alpha and nitric oxide (NO) production. Moreover, some GM-CSF effects were potentiated by the addition of IFN-gamma. Using a variety of TLR2 ligands, we established that PGE(2) release by GM-CSF-primed BMDMs was dependent on TLR2 co-receptors (TLR1, TLR6), CD14, MyD88 and the nuclear translocation of NF kappa B but was not dependent on peroxisome proliferator-activated receptor-gamma (PPAR-gamma) activation. Indeed, GM-CSF priming enhanced TLR2, TLR4 and MyD88 mRNA expression and phospho-I kappa B alpha formation. These findings demonstrate that GM-CSF drives BMDMs to present a profile relevant to the host during infections.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)FAPESP - Fundacao de Amparo a Pesquisa do Estado de Sao PauloConselho Nacional de Pesquisa (CNPq)CNPq - Conselho Nacional de Pesquis

Lung inflammation and interstitial fibrosis by targeted alveolar epithelial type I cell death

IntroductionThe pathogenesis of chronic lung diseases is multifaceted with a major role of recurrent micro-injuries of the epithelium. While several reports clearly indicated a prominent role for surfactant-producing alveolar epithelial type 2 (AT2) cells, the contribution of gas exchange-permissive alveolar epithelial type 1 (AT1) cells has not been addressed yet. Here, we investigated whether repeated injury of AT1 cells leads to inflammation and interstitial fibrosis.MethodsWe chose an inducible model of AT1 cell depletion following local diphtheria toxin (DT) administration using an iDTR flox/flox (idTRfl/fl) X Aquaporin 5CRE (Aqp5CRE) transgenic mouse strain.ResultsWe investigated repeated doses and intervals of DT to induce cell death of AT1 cells causing inflammation and interstitial fibrosis. We found that repeated DT administrations at 1ng in iDTRfl/fl X Aqp5CRE mice cause AT1 cell death leading to inflammation, increased tissue repair markers and interstitial pulmonary fibrosis.DiscussionTogether, we demonstrate that depletion of AT1 cells using repeated injury represents a novel approach to investigate chronic lung inflammatory diseases and to identify new therapeutic targets

Mycobacterial PIMs Inhibit Host Inflammatory Responses through CD14-Dependent and CD14-Independent Mechanisms

Mycobacteria develop strategies to evade the host immune system. Among them, mycobacterial LAM or PIMs inhibit the expression of pro-inflammatory cytokines by activated macrophages. Here, using synthetic PIM analogues, we analyzed the mode of action of PIM anti-inflammatory effects. Synthetic PIM1 isomer and PIM2 mimetic potently inhibit TNF and IL-12 p40 expression induced by TLR2 or TLR4 pathways, but not by TLR9, in murine macrophages. We show inhibition of LPS binding to TLR4/MD2/CD14 expressing HEK cells by PIM1 and PIM2 analogues. More specifically, the binding of LPS to CD14 was inhibited by PIM1 and PIM2 analogues. CD14 was dispensable for PIM1 and PIM2 analogues functional inhibition of TLR2 agonists induced TNF, as shown in CD14-deficient macrophages. The use of rough-LPS, that stimulates TLR4 pathway independently of CD14, allowed to discriminate between CD14-dependent and CD14-independent anti-inflammatory effects of PIMs on LPS-induced macrophage responses. PIM1 and PIM2 analogues inhibited LPS-induced TNF release by a CD14-dependent pathway, while IL-12 p40 inhibition was CD14-independent, suggesting that PIMs have multifold inhibitory effects on the TLR4 signalling pathway

All-Trans Retinoic Acid Promotes TGF-β-Induced Tregs via Histone Modification but Not DNA Demethylation on Foxp3 Gene Locus

It has been documented all-trans retinoic acid (atRA) promotes the development of TGF-β-induced CD4(+)Foxp3(+) regulatory T cells (iTreg) that play a vital role in the prevention of autoimmune responses, however, molecular mechanisms involved remain elusive. Our objective, therefore, was to determine how atRA promotes the differentiation of iTregs.Addition of atRA to naïve CD4(+)CD25(-) cells stimulated with anti-CD3/CD28 antibodies in the presence of TGF-β not only increased Foxp3(+) iTreg differentiation, but maintained Foxp3 expression through apoptosis inhibition. atRA/TGF-β-treated CD4(+) cells developed complete anergy and displayed increased suppressive activity. Infusion of atRA/TGF-β-treated CD4(+) cells resulted in the greater effects on suppressing symptoms and protecting the survival of chronic GVHD mice with typical lupus-like syndromes than did CD4(+) cells treated with TGF-β alone. atRA did not significantly affect the phosphorylation levels of Smad2/3 and still promoted iTreg differentiation in CD4(+) cells isolated from Smad3 KO and Smad2 conditional KO mice. Conversely, atRA markedly increased ERK1/2 activation, and blockade of ERK1/2 signaling completely abolished the enhanced effects of atRA on Foxp3 expression. Moreover, atRA significantly increased histone methylation and acetylation within the promoter and conserved non-coding DNA sequence (CNS) elements at the Foxp3 gene locus and the recruitment of phosphor-RNA polymerase II, while DNA methylation in the CNS3 was not significantly altered.We have identified the cellular and molecular mechanism(s) by which atRA promotes the development and maintenance of iTregs. These results will help to enhance the quantity and quality of development of iTregs and may provide novel insights into clinical cell therapy for patients with autoimmune diseases and those needing organ transplantation