Biology subjects

Werts, C.

Publications and source records attributed to Werts, C..

11 recordsLinked to original sources

bioRxiv2026

Archaeal cell envelopes selectively activate innate immunity through TLR2 in contrasts with multi-receptor bacterial sensing

The cell envelope of bacteria and fungi is a major source of microbial-associated molecular patterns (MAMPs), which are recognised by multiple host pattern recognition receptors (PRRs) such as TLRs and NODs. The key role of this recognition in protecting against infection and regulating diverse physiological processes has been studied for decades. In contrast, although the archaea are growingly recognised as an important component of the animal and human microbiome, their interaction with the innate immune system has been largely understudied. Here, we asked whether the archaeal cell envelope acts as MAMP. We selected 15 archaeal species from different environments, including the human microbiome, and harbouring diverse envelope characteristics, and we tested their ability to activate innate immune responses in human and murine immune cells. We first show that archaeal cells activate the innate immune transcription factor NF-kB and induce IL-6 secretion. Using whole archaeal cells, supernatants, and purified envelope components, we then find that none of the main PRR receptors which recognise the bacterial envelope (TLR4, TLR5, NOD1 and NOD2) are involved in this activation. We only detected activation of TLR2, regardless of the environmental origins of the archaea tested, although with overall lower activity compared to bacteria. We further demonstrate that different TLR2 heterodimers are involved, depending on the species, although the archaeal agonist remains unknown. Altogether, our results highlight a surprising paucity of interaction of archaeal cell envelope components with host PPRs, in contrast to bacteria, and open new avenues of research toward further investigation of archaea/host interplay.

immunology↗

bioRxiv2026

β-lactam antibiotics trigger TLR2-dependent Jarisch-Herxheimer reaction in leptospirosis: bacteriostatic antibiotics offer a safer alternative

Leptospirosis is a neglected zoonotic disease causing ~1 million cases and 60,000 deaths annually. Antibiotic therapy can trigger a detrimental inflammatory Jarisch-Herxheimer reaction (JHR). We investigated antibiotic effects in leptospirosis, using a mouse model of severe disease and human whole blood from healthy donors infected ex vivo with bioluminescent Leptospira interrogans. We tested the effect of amoxicillin and ceftriaxone, two bactericidal beta-Lactams antibiotics and assessed bacterial survival, cytokines, pathophysiology and JHR mechanisms upon amoxicillin. Beta-lactams induced profound pro-inflammatory cytokine release. In mice, the inflammation was TLR2 and TLR4 dependent, accordingly to the TLR4 host-specificity of LPS recognition, and amoxicillin exacerbated disease severity within hours, notably worsening myocarditis. In human whole blood, this inflammation depended largely on TLR2 (lipoprotein receptor) and TLR5 (flagellin receptor). Interestingly, only stealthy virulent isolates of Leptospira interrogans, unlike some Leptospira species belonging to intermediate and non-virulent clades such as biflexa, induced in human whole blood a JHR-like effect, which was also observed with Borrelia burgdorferi. These findings demonstrate that host-specific TLR recognition of released spirochaetal components by antibiotic treatment drives JHR, evoking a cytokine storm and worsening outcomes.

immunology↗

bioRxiv2026

Primary human dermal fibroblasts selectively sense microbial ligands and initiate immune response through chemokines secretion

Fibroblasts are traditionally considered structural cells that maintain tissue homeostasis and facilitate repair. However, accumulating evidence suggests they also participate in innate immunity, although their pattern recognition capabilities remain incompletely characterized. Here, we systematically assessed the innate immune responses of commercially available primary human dermal fibroblasts from a male and a female donor. Fibroblasts were stimulated with a panel of microbe-associated molecular patterns (MAMPs) targeting various pattern recognition receptors (PRRs), including Toll-like receptors (TLRs), NOD-like receptors (NODs), Alpha kinase 1 (ALPK1) and STING. Innate immune activation was quantified by measuring the nuclear translocation of NF-{kappa}B via high content microscopy and cytokines and chemokines secretion by ELISA; baseline PRRs expression was determined by quantitative PCR. Only a restricted subset of agonists, specifically E. coli LPS (TLR4), Poly I:C (TLR3 / RIG-I) and unexpectedly ADP heptose (ALPK1) induced robust NF-{kappa}B activation and secretion of the chemokines IL-8 and MCP-1. Apart from IL-6 and RANTES, which were produced exclusively following Poly I:C stimulation, pro-inflammatory cytokines (IL-1{beta}, TNF, IFN-{beta}) and the anti-inflammatory cytokine IL-10 remained undetectable. Consistent with this limited reactivity, qPCR of PRRs revealed basal expression of TLR4 and ALPK1, whereas most other receptors were expressed at very low or undetectable levels. Notably, NOD1 was highly expressed although no cell activation was observed with several NOD1 agonists. Dose-response analysis revealed surprisingly high sensitivity to LPS. In conclusion, primary human dermal fibroblasts exhibit a highly selective but sensitive innate immune response, largely restricted to chemokine production upon PRR activation. This unexpected dissociation between chemokine and cytokine responses suggests that fibroblasts function as sentinel cells in early skin defense, capable of detecting key microbial patterns at low concentrations, to orchestrate local immune surveillance. Further investigation into interindividual variability and context-dependent activation is needed.

immunology↗

bioRxiv2024

Myocarditis and neutrophil-mediated vascular leakage but not cytokine storm associated with fatal murine leptospirosis

Leptospirosis is a neglected re-emerging zoonosis caused by Leptospira spirochetes. Its pathophysiology remains mysterious, especially in the case of severe infection with L. interrogans. In the field of infectious diseases, the cause of death is rarely investigated in preclinical models. Here, for the first time, we identified unanticipated organ failures associated with death in a murine model of acute leptospirosis. Despite clinical similarities between bacterial sepsis and leptospirosis, striking differences were observed. Neither lung, liver, or kidney injury nor cytokine storm, or massive necroptosis could explain death. In contrast, severe leptospirosis was associated with high serum levels of the anti-inflammatory cytokine IL-10 and the chemokine RANTES, neutrophilia, pancreatitis and vascular damage. Unexpectedly, we demonstrated neutrophil-induced vascular permeability, making neutrophils a potential new therapeutic target. Strikingly, the main cause of death was myocarditis, an overlooked complication of human leptospirosis. These features are also found in patients, making this model a paradigm for better understanding human leptospirosis and designing novel therapeutic strategies.

immunology↗

bioRxiv2022

Inflammation drives age-induced loss of tissue resident macrophages

Low-grade chronic systemic inflammation, or inflammageing, is a hallmark of ageing and a risk factor for both morbidity and mortality in elderly people. Resident macrophages are tissue homeostasis sentinels that are embedded in their tissue of residence since embryonic development, thus been exposed to cumulative tissue insults throughout life. Therefore, resident macrophages, among other immune cells, emerge as potential key contributors to age-associated tissue dysfunction. Contrary to what is currently postulated, we demonstrate here that the pool of embryo-derived resident macrophages exhibits an age-dependent depletion in liver, and other solid organs and that they are not replaced by Hematopoietic Stem Cell (HSCs)-derived monocytes throughout life. Further, we demonstrate that gradual, cumulative inflammation during ageing induces this specific loss of tissue resident macrophages. Preserving a "youthful" density of resident macrophages attenuates classical hallmarks of liver age-associated dysfunction. SummaryThe pool of embryo-derived resident macrophages dwindles with age in most tissues, without compensation from Hematopoietic Stem Cell (HSC)-derived cells. This loss is not due to impaired self-renewal in old tissues but rather to increased cell death, which is driven by sustained inflammation. Attenuating inflammation sensing during ageing prevents age-induce macrophage loss and improves hallmarks of liver ageing.

immunology↗

bioRxiv2022

Leptospiral lipopolysaccharide dampens inflammation through upregulation of autophagy adaptor p62 and NRF2 signaling in macrophages

Leptospira interrogans are pathogenic bacteria responsible for leptospirosis, a worldwide zoonosis. All vertebrates can be infected, and some species like humans are susceptible to the disease whereas rodents such as mice are resistant and become asymptomatic renal carriers. Leptospires are stealth bacteria that are known to escape several immune recognition pathways and resist killing mechanisms. We recently published that leptospires may survive intracellularly and exit macrophages, in part by escaping xenophagy, a pathogen-targeting form of autophagy. Interestingly, autophagy is one of the antimicrobial mechanisms often highjacked by bacteria to evade the host immune response. In this study we therefore explored whether leptospires subvert the key molecular players of autophagy to facilitate the infection. We showed in macrophages that leptospires triggered a specific accumulation of autophagy-adaptor p62 in puncta-like structures, without major alteration of autophagy flux. Unlike active bacterial mechanisms described to date, we demonstrated that leptospires trigger p62 accumulation using a passive mechanism of LPS signaling via TLR4/TLR2. p62 is a central pleiotropic protein, not only involved in autophagy, but also mediating cell stress and death, via the translocation of transcription factors. We demonstrated that Leptospira-driven accumulation of p62 induced the translocation of transcription factor NRF2. However, NRF2 translocation upon Leptospira infection did not result as expected in antioxydant response, but dampened the production of inflammatory mediators such as iNOS/NO, TNF and IL6. Overall, these findings highlight a novel passive bacterial mechanism linked to p62/NRF2 signaling that decreases inflammation and contributes to the stealthiness of leptospires.

immunology↗

bioRxiv2022

L.interrogans prevents macrophage death and pyroptotic IL1β release through its lipopolysaccharide

Leptospira interrogans are bacteria that can infect all vertebrates and are responsible for leptospirosis, a neglected zoonosis. Some hosts are susceptible to leptospirosis whereas mice are resistant and get chronically colonized. Although leptospires escape recognition by some immune receptors, they activate NLRP3-inflammasome and trigger IL1{beta} secretion. Classically, IL1{beta} secretion is associated with lytic inflammatory cell death called pyroptosis, resulting from cytosolic LPS binding to inflammatory caspases. Interestingly, we showed that L. interrogans do not trigger cell death in either murine, human, hamster, or bovine macrophages, escaping both pyroptosis and apoptosis. Strikingly, we also revealed in murine cells, a potent antagonistic effect of leptospires and their atypical LPS on spontaneous and E. coli LPS-induced cell death. The leptospiral LPS efficiently prevents caspase 11 dimerization and subsequent gasdermin D cleavage. Finally, we showed that pyroptosis escape by leptospires prevents massive IL1 {beta} release, and we consistently found no major role of IL1-Receptor in controlling experimental leptospirosis in vivo. Overall, our findings described a novel mechanism by which leptospires dampen inflammation, thus potentially contributing to their stealthiness. Graphical abstract O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY

immunology↗

bioRxiv2020

Anti-Leptospira immunoglobulin profiling in mice reveals strain specific IgG and persistent IgM responses associated with virulence and renal colonization

Leptospira interrogans is a pathogenic spirochete responsible for leptospirosis, a neglected, zoonotic reemerging disease. Humans are sensitive hosts and may develop severe disease. Some animal species, such as rats and mice can become asymptomatic renal carriers. More than 350 leptospiral serovars have been identified, classified on the basis of the antibody response directed against the lipopolysaccharide (LPS). Similarly to whole inactivated bacteria used as human vaccines, this response is believed to confer only short-term, serogroup-specific protection. The immune response of hosts against leptospires has not been thoroughly studied and correlates of protection would be required to test vaccine candidates. In this work, we studied the immunoglobulin (Ig) profiles in mice infected with L. interrogans over time to determine whether this humoral response confers long-term protection after homologous challenge six months post-infection. Groups of mice were injected intraperitoneally with 2x107 leptospires of one of three pathogenic serovars (Manilae, Copenhageni or Icterohaemorrhagiae), attenuated mutants or heat-killed bacteria. Leptospira-specific immunoglobulin (IgA, IgM, IgG and 4 subclasses) produced in the first weeks up to 6 months post-infection were measured by ELISA. Strikingly, we found sustained high levels of IgM in mice infected with the pathogenic Manilae and Copenhageni strains, both colonizing the kidney. In contrast, the Icterohaemorrhagiae strain did not lead to kidney colonization, even at high dose, and triggered a classical IgM response that peaked at day 8 post-infection and disappeared. The virulent Manilae and Copenhageni serovars elicited high levels and similar profiles of IgG subclasses in contrast to Icterohaemorrhagiae strains that stimulated weaker antibody responses. Inactivated heat-killed Manilae strains elicited very low responses. However, all mice pre-injected with leptospires challenged with high doses of homologous bacteria did not develop acute leptospirosis, and all antibody responses were boosted after challenge. Furthermore, we showed that 2 months post challenge, mice pre-infected with the M895 Manilae LPS mutant or heat-killed bacterin were completely protected against renal colonization. In conclusion, we observed a sustained IgM response potentially associated with chronic leptospiral renal infection. We also demonstrated in mice different profiles of protective antibody response after L. interrogans infection, depending on the serovar and virulence of strains. Author summaryLeptospira interrogans is a pathogenic spirochete responsible for leptospirosis, a neglected zoonotic reemerging disease. The immune response of hosts against these bacteria has not been thoroughly studied. Here, we studied over 6 months the immunoglobulin (Ig) profiles in mice infected with L. interrogans and determined whether this humoral response confers long-term protection after homologous challenge six months after primary infection. Groups of mice were infected intraperitoneally with 2x107 bacteria of one of three different pathogenic serovars (Manilae, Copenhageni and Icterohaemorrhagiae) and some corresponding attenuated avirulent mutants. We measured by ELISA each type of Leptospira-specific Ig (IgA, IgM, IgG and 4 subclasses) produced in the first weeks up to 6 months post-infection. We showed different profiles of antibody response after L. interrogans challenge in mice, depending on the serovar and virulence of strains. However, all infected mice, including the ones harboring low antibody levels, like mice vaccinated with an inactivated, heat-killed strain, were protected against leptospirosis after challenge. Notably, we also showed an unusual sustained IgM response associated with chronic leptospiral colonization. Altogether, this long-term immune protection is different from what is known in humans and warrants further investigation.

immunology↗

bioRxiv2020

Humanized TLR4 immunocompetent mice are more susceptible to kidney disease after infection with Leptospira interrogans than wild-type mice

Toll-Like Receptor (TLR) 4, the LPS receptor, plays a central role in the control of leptospirosis and absence of TLR4 results in lethal infection in mice. Because human TLR4 does not sense the atypical leptospiral-LPS, we hypothesized that TLR4/MD-2 humanized transgenic mice (huTLR4) may be more susceptible to leptospirosis than wild-type mice, and thus may constitute a model of acute human leptospirosis. Therefore, we infected huTLR4 mice, which express human TLR4 but not murine TLR4, with a high but sublethal dose of L. interrogans serovar Copenhageni FioCruz (Leptospira) in comparison to C57BL/6J wildtype (WT) and, as a control, a congenic strain in which the tlr4 coding sequences are deleted (muTLR4Lps-del). We show that the huTLR4 gene is fully functional in the murine background. We found that dissemination of Leptospira in blood, shedding in urine, colonization of the kidney and overall kinetics of leptospirosis progression is equivalent between WT and huTLR4 C57BL/6J mice. Furthermore, inflammation of the kidney appeared to be subdued in huTLR4 compared to WT mice in that we observed less infiltrates of mononuclear lymphocytes, less innate immune markers and no relevant differences in fibrosis markers. Contrary to our hypothesis, huTLR4 mice showed less inflammation and kidney pathology, and are not more susceptible to leptospirosis than WT mice. This study is significant as it indicates that one intact TLR4 gene, be it mouse or human, is necessary to control acute leptospirosis. Contribution to the fieldDifferences of recognition exist between mouse and human TLR4, in that the anchor of LPS in the outer membrane of Leptospira (LipidA) activates murine, but not human TLR4. We hypothesized that if human TLR4 does not sense leptospiral-LPS, then transgenic mice in which murine TLR4 was replaced with human TLR4, should be more susceptible to Leptospira dissemination as compared to congenic wild-type mice, which could result in a more robust inflammatory response and pathology in the kidney. However, we found that impaired sensing of leptospiral-LPS in huTLR4 mice did not affect overall infection in comparison to wild-type mice and does not result in increased pathology of the kidney. Our study indicates that rather than leptospiral-LPS sensing, the presence of a fully functional TLR4 receptor is necessary to control acute leptospirosis.

microbiology↗

bioRxiv2020

Escape of TLR5 Recognition by Leptospira spp: A Rationale for Atypical Endoflagella

Leptospira interrogans are invasive bacteria responsible for leptospirosis, a worldwide zoonosis. They possess two periplasmic endoflagella that allow their motility. L. interrogans are stealth pathogens that escape the innate immune responses of the NOD-like receptors NOD1/2, and the human Toll-like receptor (TLR)4, sensing peptidoglycan and lipopolysaccharide (LPS), respectively. TLR5 is another receptor of bacterial cell wall components, recognizing flagellin subunits. To study the contribution of TLR5 in the host defense against leptospires, we infected WT and TLR5 deficient mice with pathogenic L. interrogans and tracked the infection by in vivo live imaging of bioluminescent bacteria or by q-PCR. We did not identify any protective or inflammatory role of murine TLR5 to control pathogenic Leptospira. Likewise, subsequent in vitro experiments showed that infections with different live strains of L. interrogans and L. biflexa did not trigger TLR5. However, unexpectedly, heat-killed bacteria stimulated human and bovine TLR5, although barely mouse TLR5. Abolition of TLR5 recognition required extensive boiling time of the bacteria or proteinase K treatment, showing an unusual high stability of the leptospiral flagellins. Interestingly, using antimicrobial peptides to destabilize live leptospires, we detected some TLR5 activity, suggesting that TLR5 could participate in the fight against leptospires in humans or cattle. Using different Leptospira strains with mutations in flagellin proteins, we further showed that neither FlaAs nor Fcps participated in the recognition by TLR5, suggesting a role for the FlaBs. These have structural homology to Salmonella FliC, and conserved residues important for TLR5 activation, as shown by in silico analyses. Accordingly, we found that leptospires regulate the expression of FlaB mRNA according to the growth phase in vitro, and that infection with L. interrogans in hamsters and in mice downregulated the expression of the FlaBs but not the FlaAs subunits. Altogether, in contrast to different bacteria that modify their flagellin sequences to escape TLR5 recognition, our study suggests that the peculiar central localization and stability of the FlaB monomers in the periplasmic endoflagella, associated with the downregulation of FlaB subunits in hosts, constitute an efficient strategy of leptospires to escape TLR5 recognition and the immune response.

immunology↗

bioRxiv2020

Leptospiral LPS escapes mouse TLR4 internalization and TRIF?associated antimicrobial responses through O antigen and associated lipoproteins

Leptospirosis is a worldwide re-emerging zoonosis caused by pathogenic Leptospira spp. All vertebrate species can be infected; humans are sensitive hosts whereas other species, such as rodents, may become long-term renal carrier reservoirs. Upon infection, innate immune responses are initiated by recognition of Microbial Associated Molecular Patterns (MAMPs) by Pattern Recognition Receptors (PRRs). Among MAMPs, the lipopolysaccharide (LPS) is recognized by the Toll-Like-Receptor 4 (TLR4) and activates both the MyD88-dependent pathway at the plasma membrane and the TRIF-dependent pathway after TLR4 internalization. We previously showed that leptospiral LPS is not recognized by the human TLR4, whereas it signals through murine TLR4, which mediates mouse resistance to acute leptospirosis. However, leptospiral LPS has low endotoxicity in mouse cells and is an agonist of TLR2, the sensor for bacterial lipoproteins. Here, using confocal microscopy and flow cytometry, we showed that the LPS of L. interrogans did not induce internalization of TLR4 in mouse macrophages, unlike the LPS of Escherichia coli. Consequently, the LPS failed to induce the production of the TRIF-dependent nitric oxide and RANTES, both important antimicrobial responses. Using shorter O antigen LPS and repurified leptospiral LPS with reporter HEK cells, we further found this TLR4-TRIF escape to be dependent on both the co-purifying lipoproteins and the full-length O antigen. Furthermore, our data suggest that the O antigen could alter the binding of the leptospiral LPS to the co-receptor CD14 that is essential for TLR4-TRIF activation. Overall, we describe here a novel immune escape mechanism linked to leptospiral LPS. We hypothesize that the LPS, already known as a virulence factor, plays a major role in the innate immune evasion of the leptospires, thereby contributing to their stealthiness and chronicity in mice. Author summaryLeptospira interrogans is a bacterial pathogen, responsible for leptospirosis, a worldwide neglected reemerging disease. L. interrogans may cause an acute severe disease in humans, whereas rodents and other animals asymptomatically carry the leptospires in their kidneys. They can therefore excrete live bacteria in urine and contaminate the environment. Leptospires are stealth pathogens known to escape the innate immune defenses of their hosts. They are covered in lipopolysaccharide (LPS), a bacterial motif recognized in mammals through the Toll-like receptor 4 (TLR4), which triggers two different signaling pathways. We showed previously that pathogenic leptospires escape TLR4 recognition in humans. Here we show in mice that the leptospiral LPS triggers only one arm of the TLR4 pathway and escapes the other, hence avoiding production of antimicrobial compounds. Removing the lipoproteins that always co-purify with the leptospiral LPS, or using shorter LPS, restores the stimulation of both pathways. This suggests a novel escape mechanism linked to the LPS and involving lipoproteins that could be instrumental for leptospires to escape the mouse defense and allows for their chronic renal colonization.

immunology↗