Chitin is the second most abundant polysaccharide in nature and linked to fungal infection and asthma. However, bona fide immune receptors directly binding chitin and signaling immune activation and ...inflammation have not been clearly identified because polymeric crude chitin with unknown purity and molecular composition has been used. By using defined chitin (N‐acetyl‐glucosamine) oligomers, we here identify six‐subunit‐long chitin chains as the smallest immunologically active motif and the innate immune receptor Toll‐like receptor (TLR2) as a primary fungal chitin sensor on human and murine immune cells. Chitin oligomers directly bind TLR2 with nanomolar affinity, and this fungal TLR2 ligand shows overlapping and distinct signaling outcomes compared to known mycobacterial TLR2 ligands. Unexpectedly, chitin oligomers composed of five or less subunits are inactive, hinting to a size‐dependent system of immuno‐modulation that appears conserved in plants and humans. Since blocking of the chitin‐TLR2 interaction effectively prevents chitin‐mediated inflammation in vitro and in vivo, our study highlights the chitin‐TLR2 interaction as a potential target for developing novel therapies in chitin‐related pathologies and fungal disease.
Synopsis
Chitin, a polysaccharide linked to fungal infection and allergic asthma, directly binds to the innate immune receptor TLR2 and triggers inflammation dependent on oligomer size. Blocking the chitin‐TLR2 interaction effectively prevents chitin‐mediated inflammation.
Oligomeric chains of fungal chitin directly bind to TLR2 and trigger inflammation.
Blocking of the chitin‐TLR2 interaction prevents chitin‐mediated inflammation in vitro and in vivo.
Size‐dependent chitin recognition based on oligomers is found in both plants and humans.
Chitin, a polysaccharide linked to fungal infection and allergic asthma, directly binds to the innate immune receptor TLR2 and triggers inflammation dependent on oligomer size. Blocking the chitin‐TLR2 interaction effectively prevents chitin‐mediated inflammation.
Abstract Skin repair is an important field of the tissue engineering, especially in the case of extended third-degree burns, where the current treatments are still insufficient in promoting ...satisfying skin regeneration. Bio-inspired bi-layered physical hydrogels only constituted of chitosan and water were processed and applied to the treatment of full-thickness burn injuries. The aim of the study was at assessing whether this material was totally accepted by the host organism and allowed in vivo skin reconstruction of limited area third-degree burns. A first layer constituted of a rigid protective gel ensured good mechanical properties and gas exchanges. A second soft and flexible layer allowed the material to follow the geometry of the wound and ensured a good superficial contact. To compare, highly viscous solutions of chitosan were also considered. Veterinary experiments were performed on pig's skins and biopsies at days 9, 17, 22, 100 and 293, were analysed by histology and immuno-histochemistry. Only one chitosan material was used for each time. All the results showed that chitosan materials were well tolerated and promoted a good tissue regeneration. They induced inflammatory cells migration and angiogenetic activity favouring a high vascularisation of the neo-tissue. At day 22, type I and IV collagens were synthesised under the granulation tissue and the formation of the dermal–epidermal junction was observed. After 100 days, the new tissue was quite similar to a native skin, especially by its aesthetic aspect and its great flexibility.
Sirtuin 2 (SIRT2) and SIRT3 are cytoplasmic and mitochondrial NAD-dependent deacetylases. SIRT2 and SIRT3 target proteins involved in metabolic, proliferation and inflammation pathways and have been ...implicated in the pathogenesis of neurodegenerative, metabolic and oncologic disorders. Both pro- and anti-inflammatory effects have been attributed to SIRT2 and SIRT3, and single deficiency in SIRT2 or SIRT3 had minor or no impact on antimicrobial innate immune responses. Here, we generated a SIRT2/3 double deficient mouse line to study the interactions between SIRT2 and SIRT3. SIRT2/3
mice developed normally and showed subtle alterations of immune cell populations in the bone marrow, thymus, spleen, blood and peritoneal cavity that contained notably more anti-inflammatory B-1a cells and less NK cells.
, SIRT2/3
macrophages favored fatty acid oxidation (FAO) over glycolysis and produced increased levels of both proinflammatory and anti-inflammatory cytokines. In line with metabolic adaptation and increased numbers of peritoneal B-1a cells, SIRT2/3
mice were robustly protected from endotoxemia. Yet, SIRT2/3 double deficiency did not modify endotoxin tolerance. Overall, these data suggest that sirtuins can act in concert or compensate each other for certain immune functions, a parameter to be considered for drug development. Moreover, inhibitors targeting multiple sirtuins developed for clinical purposes may be useful to treat inflammatory diseases.
(Group B
, GBS) is a leading pathogen of neonatal sepsis. The host-pathogen interactions underlying the progression to life-threatening infection in newborns are incompletely understood. Macrophages ...are first line in host defenses against GBS, contributing to the initiation, amplification, and termination of immune responses. The goal of this study was to compare the response of newborn and adult monocyte-derived macrophages (MDMs) to GBS.
Monocytes from umbilical cord blood of healthy term newborns and from peripheral blood of healthy adult subjects were cultured with M-CSF to induce MDMs. M-CSF-MDMs, GM-CSF- and IFNγ-activated MDMs were exposed to GBS COH1, a reference strain for neonatal sepsis.
GBS induced a greater release of IL-1β, IL-6, IL-10, IL-12p70 and IL-23 in newborn compared to adult MDMs, while IL-18, IL-21, IL-22, TNF, RANTES/CCL5, MCP-1/CCL2 and IL-8/CXCL8 were released at similar levels. MDM responses to GBS were strongly influenced by conditions of activation and were distinct from those to synthetic bacterial lipopeptides and lipopolysaccharides. Under similar conditions of opsonization, newborn MDMs phagocytosed and killed GBS as efficiently as adult MDMs.
Altogether, the production of excessive levels of Th1- (IL-12p70), Th17-related (IL-1β, IL-6, IL-23) and anti-inflammatory (IL-10) cytokines is consistent with a dysregulated response to GBS in newborns. The high responsiveness of newborn MDMs may play a role in the progression of GBS infection in newborns, possibly contributing to the development of life-threatening organ dysfunction.
Patients admitted to the intensive care unit (ICU) often experience endotoxemia, nosocomial infections and sepsis. Polymorphonuclear and monocytic myeloid-derived suppressor cells (PMN-MDSCs and ...M-MDSCs) can have an important impact on the development of infectious diseases, but little is known about their potential predictive value in critically ill patients. Here, we used unsupervised flow cytometry analyses to quantify MDSC-like cells in healthy subjects challenged with endotoxin and in critically ill patients admitted to intensive care units and at risk of developing infections. Cells phenotypically similar to PMN-MDSCs and M-MDSCs increased after endotoxin challenge. Similar cells were elevated in patients at ICU admission and normalized at ICU discharge. A subpopulation of M-MDSC-like cells expressing intermediate levels of CD15 (CD15
M-MDSCs) was associated with overall mortality (
= 0.02). Interestingly, the high abundance of PMN-MDSCs and CD15
M-MDSCs was a good predictor of mortality (
= 0.0046 and 0.014), with area under the ROC curve for mortality of 0.70 (95% CI = 0.4-1.0) and 0.86 (0.62-1.0), respectively. Overall, our observations support the idea that MDSCs represent biomarkers for sepsis and that flow cytometry monitoring of MDSCs may be used to risk-stratify ICU patients for targeted therapy.
Viruses have developed strategies to counteract signalling through Toll-like receptors (TLRs) that are involved in the detection of viruses and induction of proinflammatory cytokines and IFNs. ...Vaccinia virus (VACV) encodes A46 protein which disrupts TLR signalling by interfering with TLR: adaptor interactions. Since the innate immune response to viruses is critical to induce protective immunity, we studied whether deletion of A46R gene in a NYVAC vector expressing HIV-1 Env, Gag, Pol and Nef antigens (NYVAC-C) improves immune responses against HIV-1 antigens. This question was examined in human macrophages and in mice infected with a single A46R deletion mutant of the vaccine candidate NYVAC-C (NYVAC-C-ΔA46R). The viral gene A46R is not required for virus replication in primary chicken embryo fibroblast (CEF) cells and its deletion in NYVAC-C markedly increases TNF, IL-6 and IL-8 secretion by human macrophages. Analysis of the immune responses elicited in BALB/c mice after DNA prime/NYVAC boost immunization shows that deletion of A46R improves the magnitude of the HIV-1-specific CD4 and CD8 T cell immune responses during adaptive and memory phases, maintains the functional profile observed with the parental NYVAC-C and enhances anti-gp120 humoral response during the memory phase. These findings establish the immunological role of VACV A46R on innate immune responses of macrophages in vitro and antigen-specific T and B cell immune responses in vivo and suggest that deletion of viral inhibitors of TLR signalling is a useful approach for the improvement of poxvirus-based vaccine candidates.
Trained immunity refers to the ability of the innate immune system exposed to a first challenge to provide an enhanced response to a secondary homologous or heterologous challenge. We reported that ...training induced with β-glucan one week before infection confers protection against a broad-spectrum of lethal bacterial infections. Whether this protection persists over time is unknown. To tackle this question, we analyzed the immune status and the response to
(
) of mice trained 9 weeks before analysis. The induction of trained immunity increased bone marrow myelopoiesis and blood counts of Ly6C
inflammatory monocytes and polymorphonuclear neutrophils (PMNs).
, whole blood, PMNs and monocytes from trained mice produced increased levels of cytokines in response to microbial products and limited the growth of
.
, following challenge with
, peripheral blood leukocytes were massively depleted in control mice but largely preserved in trained mice. PMNs were reduced also in the spleen from control mice, and increased in the spleen of trained mice. In transwell experiments, PMNs from trained mice showed increased spontaneous migration and CXCL2/MIP2α-induced chemotaxis, suggesting that training promotes the migration of PMNs in peripheral organs targeted by
. Trained PMNs and monocytes had higher glycolytic activity and mitochondrial respiration than control cells when exposed to
. Bacterial burden and dissemination in blood, spleen and liver as well as systemic cytokines and inflammation (multiplex bead assay and bioluminescence imaging) were reduced in trained mice. In full agreement with these results, mice trained 9 weeks before infection were powerfully protected from lethal listeriosis. Altogether, these data suggest that training increases the generation and the antimicrobial activity of PMNs and monocytes, which may confer prolonged protection from lethal bacterial infection.
The pro‐inflammatory cytokine macrophage migration inhibitory factor (MIF) acts as a physiological counter‐regulator of the immuno‐suppressive effects of glucocorticoids. However, the mechanisms ...whereby MIF exerts its counter‐balancing effect remain largely unknown. Here we report that MAPK phosphatase 1 (MKP‐1), an archetypal member of dual specificity phosphatase that inactivates MAPK activity in response to pro‐inflammatory stimuli, is a critical target of MIF‐glucocorticoid crosstalk. Recombinant MIF counter‐regulated in a dose‐dependent fashion dexamethasone inhibition of TNF and IL‐8 production by RAW 264.7 macrophages and U‐937 promonocytes stimulated with lipoplysaccharides (LPS) or with LPS plus phorbol 12‐myristate 13‐acetate. Stimulation of RAW 264.7 macrophages with dexamethasone or dexamethasone plus LPS led to a robust up‐regulation of MKP‐1 mRNA and protein expressions that were counter‐regulated by addition of recombinant MIF. Antisense MIF macrophages expressing reduced levels of endogenous MIF produced higher amount of MKP‐1 and lower amount of TNF after exposure to dexamethasone and dexamethasone plus LPS, indicating that endogenous MIF acts in an autocrine fashion to override glucocorticoid‐induced MKP‐1 expression and inhibition of cytokine production. Taken together, these data identify MKP‐1 as a molecular target of MIF‐glucocorticoid crosstalk and provide a molecular basis for the control of macrophage responses by a pair of physiological regulators of innate immunity.
See accompanying commentary: http://dx.doi.org/10.1002/eji.200535556
Abstract
Background
Clarithromycin may act as immune-regulating treatment in sepsis and acute respiratory dysfunction syndrome. However, clinical evidence remains inconclusive. We aimed to evaluate ...whether clarithromycin improves 28-day mortality among patients with sepsis, respiratory and multiple organ dysfunction syndrome.
Methods
We conducted a multicenter, randomized, clinical trial in patients with sepsis. Participants with ratio of partial oxygen pressure to fraction of inspired oxygen less than 200 and more than 3 SOFA points from systems other than the respiratory function were enrolled between December 2017 and September 2019. Patients were randomized to receive 1 gr of clarithromycin or placebo intravenously once daily for 4 consecutive days. The primary endpoint was 28-day all-cause mortality. Secondary outcomes were 90-day mortality; sepsis response (defined as at least 25% decrease in SOFA score by day 7); sepsis recurrence; and differences in peripheral blood cell populations and leukocyte transcriptomics.
Results
Fifty-five patients were allocated to each arm. By day 28, 27 (49.1%) patients in the clarithromycin and 25 (45.5%) in the placebo group died (risk difference 3.6% 95% confidence interval (CI) − 15.7 to 22.7;
P
= 0.703, adjusted OR 1.03 95%CI 0.35–3.06;
P
= 0.959). There were no statistical differences in 90-day mortality and sepsis response. Clarithromycin was associated with lower incidence of sepsis recurrence (OR 0.21 95%CI 0.06–0.68;
P
= 0.012); significant increase in monocyte HLA-DR expression; expansion of non-classical monocytes; and upregulation of genes involved in cholesterol homeostasis. Serious and non-serious adverse events were equally distributed.
Conclusions
Clarithromycin did not reduce mortality among patients with sepsis with respiratory and multiple organ dysfunction. Clarithromycin was associated with lower sepsis recurrence, possibly through a mechanism of immune restoration.
Clinical trial registration
clinicaltrials.gov identifier
NCT03345992
registered 17 November 2017; EudraCT 2017-001056-55.
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