Gasdermin D inhibition prevents multiple organ dysfunction during sepsis by blocking NET formation.

Multiple organ dysfunction is the most severe outcome of sepsis progression and is highly correlated with a worse prognosis. Excessive neutrophil extracellular traps (NETs) are critical players in the development of organ failure during sepsis. Therefore, interventions targeting NET release would likely effectively prevent NET-based organ injury associated with this disease. Herein, we demonstrate that the pore-forming protein gasdermin D (GSDMD) is active in neutrophils from septic humans and mice and plays a crucial role in NET release. Inhibition of GSDMD with disulfiram or genic deletion abrogated NET formation, reducing multiple organ dysfunction and sepsis lethality. Mechanistically, we demonstrate that during sepsis, activation of the caspase-11/GSDMD pathway controls NET release by neutrophils during sepsis. In summary, our findings uncover a novel therapeutic use for disulfiram and suggest that GSDMD is a therapeutic target to improve sepsis treatment.

FK506 impairs neutrophil migration that results in increased polymicrobial sepsis susceptibility.

OBJECTIVE: This study aimed to investigate the effects of FK506 on experimental sepsis immunopathology. It investigated the effect of FK506 on leukocyte recruitment to the site of infection, systemic cytokine production, and organ injury in mice with sepsis. METHODS: Using a murine cecal ligation and puncture (CLP) peritonitis model, the experiments were performed with wild-type (WT) mice and mice deficient in the gene Nfat1 (Nfat1-/-) in the C57BL/6 background. Animals were treated with 2.0 mg/kg of FK506, subcutaneously, 1 h before the sepsis model, twice a day (12 h/12 h). The number of bacteria colony forming units (CFU) was manually counted. The number of neutrophils in the lungs was estimated by the myeloperoxidase (MPO) assay. The expression of CXCR2 in neutrophils was determined using flow cytometry analysis. The expression of inflammatory cytokines in macrophage was determined using ELISA. The direct effect of FK506 on CXCR2 internalization was evaluated using HEK-293T cells after CXCL2 stimulation by the BRET method. RESULTS: FK506 treatment potentiated the failure of neutrophil migration into the peritoneal cavity, resulting in bacteremia and an exacerbated systemic inflammatory response, which led to higher organ damage and mortality rates. Failed neutrophil migration was associated with elevated CXCL2 chemokine plasma levels and lower expression of the CXCR2 receptor on circulating neutrophils compared with non-treated CLP-induced septic mice. FK506 did not directly affect CXCL2-induced CXCR2 internalization by transfected HEK-293 cells or mice neutrophils, despite increasing CXCL2 release by LPS-treated macrophages. Finally, the CLP-induced response of Nfat1-/- mice was similar to those observed in the Nfat1+/+ genotype, suggesting that the FK506 effect is not dependent on the NFAT1 pathway. CONCLUSION: Our data indicate that the increased susceptibility to infection of FK506-treated mice is associated with failed neutrophil migration due to the reduced membrane availability of CXCR2 receptors in response to exacerbated levels of circulating CXCL2.

Sepsis-Induced Immunosuppression Is Marked by an Expansion of a Highly Suppressive Repertoire of FOXP3+ T-Regulatory Cells Expressing TIGIT.

BACKGROUND: Although the literature shows that an increase in both the number and suppressive function of CD4+forkhead box P3 (FOXP3)+ T-regulatory cells (Tregs) during sepsis contributes to an immunosuppressed state, little is known about the identity of these cells. METHODS: Using the sepsis mouse model of cecal ligation and puncture (CLP), we analyzed the frequency and molecular signature of the T-cell immunoglobulin and ITIM domain (TIGIT)+ and TIGIT- Treg subsets, using flow cytometry and quantitative polymerase chain reaction. In addition, ST2-/- and signal transducer and activator of transcription 6 (STAT6)-/- mice were submitted to CLP or recombinant interleukin 33 (IL-33) treatment to investigate the mechanism whereby TIGIT+ Tregs differentiate during sepsis. RESULTS: Sepsis was marked by the sustained expansion of the highly suppressive TIGIT+ Treg subset, which expresses Helios, neuropilin 1, and high levels of Tnfrsf18 and Pdcd1 at 15 days after CLP. The increase in TIGIT+ Tregs was accompanied by higher susceptibility to nosocomial bacteria challenge, suggesting their association with post sepsis immunosuppression. Mechanistically, we found that the ST2 deletion abrogated the expansion of the TIGIT+ Treg subset during sepsis. Furthermore, treatment with recombinant IL-33 resulted in the expansion of TIGIT+ Tregs depending on the STAT6 and M2 macrophages. CONCLUSIONS: These findings demonstrated that only the TIGIT+ Tregs remain stably expanded at the late phase of sepsis. Moreover, the expansion of TIGIT+ Tregs is dependent on the IL-33/ST2/STAT6/M2 macrophage axis.

Neutrophil extracellular traps (NETs) exacerbate severity of infant sepsis.

BACKGROUND: Neutrophil extracellular traps (NETs) are innate defense mechanisms that are also implicated in the pathogenesis of organ dysfunction. However, the role of NETs in pediatric sepsis is unknown. METHODS: Infant (2 weeks old) and adult (6 weeks old) mice were submitted to sepsis by intraperitoneal (i.p.) injection of bacteria suspension or lipopolysaccharide (LPS). Neutrophil infiltration, bacteremia, organ injury, and concentrations of cytokine, NETs, and DNase in the plasma were measured. Production of reactive oxygen and nitrogen species and release of NETs by neutrophils were also evaluated. To investigate the functional role of NETs, mice undergoing sepsis were treated with antibiotic plus rhDNase and the survival, organ injury, and levels of inflammatory markers and NETs were determined. Blood samples from pediatric and adult sepsis patients were collected and the concentrations of NETs measured. RESULTS: Infant C57BL/6 mice subjected to sepsis or LPS-induced endotoxemia produced significantly higher levels of NETs than the adult mice. Moreover, compared to that of the adult mice, this outcome was accompanied by increased organ injury and production of inflammatory cytokines. The increased NETs were associated with elevated expression of Padi4 and histone H3 citrullination in the neutrophils. Furthermore, treatment of infant septic mice with rhDNase or a PAD-4 inhibitor markedly attenuated sepsis. Importantly, pediatric septic patients had high levels of NETs, and the severity of pediatric sepsis was positively correlated with the level of NETs. CONCLUSION: This study reveals a hitherto unrecognized mechanism of pediatric sepsis susceptibility and suggests that NETs represents a potential target to improve clinical outcomes of sepsis.

Hos1 deacetylates Smc3 to close the cohesin acetylation cycle.

Cohesion between sister chromatids is mediated by the chromosomal cohesin complex. In budding yeast, cohesin is loaded onto chromosomes during the G1 phase of the cell cycle. During S phase, the replication fork-associated acetyltransferase Eco1 acetylates the cohesin subunit Smc3 to promote the establishment of sister chromatid cohesion. At the time of anaphase, Smc3 loses its acetylation again, but the Smc3 deacetylase and the possible importance of Smc3 deacetylation are unknown. Here, we show that the class I histone deacetylase family member Hos1 is responsible for Smc3 deacetylation. Cohesin is protected from deacetylation while bound to chromosomes but is deacetylated as soon as it dissociates from chromosomes at anaphase onset. Nonacetylated Smc3 is required as a substrate for cohesion establishment in the following cell cycle. Our results complete the description of an Smc3 acetylation cycle and provide unexpected insight into the importance of de novo Smc3 acetylation for cohesion establishment.

Therapeutic potential and limitations of cholinergic anti-inflammatory pathway in sepsis.

Sepsis is one of the main causes of mortality in hospitalized patients. Despite the recent technical advances and the development of novel generation of antibiotics, severe sepsis remains a major clinical and scientific challenge in modern medicine. Unsuccessful efforts have been dedicated to the search of therapeutic options to treat the deleterious inflammatory components of sepsis. Recent findings on neuronal networks controlling immunity raised expectations for novel therapeutic strategies to promote the regulation of sterile inflammation, such as autoimmune diseases. Interesting studies have dissected the anatomical constituents of the so-called "cholinergic anti-inflammatory pathway", suggesting that electrical vagus nerve stimulation and pharmacological activation of beta-2 adrenergic and alpha-7 nicotinic receptors could be alternative strategies for improving inflammatory conditions. However, the literature on infectious diseases, such as sepsis, is still controversial and, therefore, the real therapeutic potential of this neuroimmune pathway is not well defined. In this review, we will discuss the beneficial and detrimental effects of neural manipulation in sepsis, which depend on the multiple variables of the immune system and the nature of the infection. These observations suggest future critical studies to validate the clinical implications of vagal parasympathetic signaling in sepsis treatment.

Telomeres avoid end detection by severing the checkpoint signal transduction pathway.

Telomeres protect the normal ends of chromosomes from being recognized as deleterious DNA double-strand breaks. Recent studies have uncovered an apparent paradox: although DNA repair is prevented, several proteins involved in DNA damage processing and checkpoint responses are recruited to telomeres in every cell cycle and are required for end protection. It is currently not understood how telomeres prevent DNA damage responses from causing permanent cell cycle arrest. Here we show that fission yeast (Schizosaccharomyces pombe) cells lacking Taz1, an orthologue of human TRF1 and TRF2 (ref. 2), recruit DNA repair proteins (Rad22(RAD52) and Rhp51(RAD51), where the superscript indicates the human orthologue) and checkpoint sensors (RPA, Rad9, Rad26(ATRIP) and Cut5/Rad4(TOPBP1)) to telomeres. Despite this, telomeres fail to accumulate the checkpoint mediator Crb2(53BP1) and, consequently, do not activate Chk1-dependent cell cycle arrest. Artificially recruiting Crb2(53BP1) to taz1Δ telomeres results in a full checkpoint response and cell cycle arrest. Stable association of Crb2(53BP1) to DNA double-strand breaks requires two independent histone modifications: H4 dimethylation at lysine 20 (H4K20me2) and H2A carboxy-terminal phosphorylation (γH2A). Whereas γH2A can be readily detected, telomeres lack H4K20me2, in contrast to internal chromosome locations. Blocking checkpoint signal transduction at telomeres requires Pot1 and Ccq1, and loss of either Pot1 or Ccq1 from telomeres leads to Crb2(53BP1) foci formation, Chk1 activation and cell cycle arrest. Thus, telomeres constitute a chromatin-privileged region of the chromosomes that lack essential epigenetic markers for DNA damage response amplification and cell cycle arrest. Because the protein kinases ATM and ATR must associate with telomeres in each S phase to recruit telomerase, exclusion of Crb2(53BP1) has a critical role in preventing telomeres from triggering cell cycle arrest.

Histamine h2 receptor signaling in the pathogenesis of sepsis: studies in a murine diabetes model.

Type 1 diabetes enhances susceptibility to infection and favors the sepsis development. In addition, diabetic mice produced higher levels of histamine in several tissues and in the blood after LPS stimulation than nondiabetic mice. In this study, we aimed to explore the role of mast cells (MCs) and histamine in neutrophil migration and, consequently, infection control in diabetic mice with mild sepsis (MS) induced by cecum ligation and puncture. We used female BALB/c, MC-sufficient (WB/B6), MC-deficient (W/W(v)), and NOD mice. Diabetic mice given MS displayed 100% mortality within 24 h, whereas all nondiabetic mice survived for at least 5 d. The mortality rate of diabetic mice was reduced to 57% after the depletion of MC granules with compound 48/80. Moreover, this pretreatment increased neutrophil migration to the focus of infection, which reduced systemic inflammatory response and bacteremia. The downregulation of CXCR2 and upregulation of G protein-coupled receptor kinase 2 in neutrophils was prevented by pretreatment of diabetic mice given MS with compound 48/80. In addition, blocking the histamine H2 receptor restored neutrophil migration, enhanced CXCR2 expression, decreased bacteremia, and improved sepsis survival in alloxan-induced diabetic and spontaneous NOD mice. Finally, diabetic W/W(v) mice had neutrophil migration to the peritoneal cavity, increased CXCR2 expression, and reduced bacteremia compared with diabetic WB/B6 mice. These results demonstrate that histamine released by MCs reduces diabetic host resistance to septic peritonitis in mice.

An Eco1-independent sister chromatid cohesion establishment pathway in S. cerevisiae.

Cohesion between sister chromatids, mediated by the chromosomal cohesin complex, is a prerequisite for their alignment on the spindle apparatus and segregation in mitosis. Budding yeast cohesin first associates with chromosomes in G1. Then, during DNA replication in S-phase, the replication fork-associated acetyltransferase Eco1 acetylates the cohesin subunit Smc3 to make cohesin's DNA binding resistant to destabilization by the Wapl protein. Whether stabilization of cohesin molecules that happen to link sister chromatids is sufficient to build sister chromatid cohesion, or whether additional reactions are required to establish these links, is not known. In addition to Eco1, several other factors contribute to cohesion establishment, including Ctf4, Ctf18, Tof1, Csm3, Chl1 and Mrc1, but little is known about their roles. Here, we show that each of these factors facilitates cohesin acetylation. Moreover, the absence of Ctf4 and Chl1, but not of the other factors, causes a synthetic growth defect in cells lacking Eco1. Distinct from acetylation defects, sister chromatid cohesion in ctf4Δ and chl1Δ cells is not improved by removing Wapl. Unlike previously thought, we do not find evidence for a role of Ctf4 and Chl1 in Okazaki fragment processing, or of Okazaki fragment processing in sister chromatid cohesion. Thus, Ctf4 and Chl1 delineate an additional acetylation-independent pathway that might hold important clues as to the mechanism of sister chromatid cohesion establishment.

Paediatric sepsis survivors are resistant to sepsis-induced long-term immune dysfunction.

BACKGROUND AND PURPOSE: Sepsis-surviving adult individuals commonly develop immunosuppression and increased susceptibility to secondary infections, an outcome mediated by the axis IL-33/ILC2s/M2 macrophages/Tregs. Nonetheless, the long-term immune consequences of paediatric sepsis are indeterminate. We sought to investigate the role of age in the genesis of immunosuppression following sepsis. EXPERIMENTAL APPROACH: Here, we compared the frequency of Tregs, the activation of the IL-33/ILC2s axis in M2 macrophages and the DNA methylation of epithelial lung cells from post-septic infant and adult mice. Likewise, sepsis-surviving mice were inoculated intranasally with Pseudomonas aeruginosa or by subcutaneous inoculation of the B16 melanoma cell line. Finally, blood samples from sepsis-surviving patients were collected and the concentration of IL-33 and Tregs frequency were assessed. KEY RESULTS: In contrast to 6-week-old mice, 2-week-old mice were resistant to secondary infection and did not show impairment in tumour controls upon melanoma challenge. Mechanistically, increased IL-33 levels, Tregs expansion, and activation of ILC2s and M2-macrophages were observed in 6-week-old but not 2-week-old post-septic mice. Moreover, impaired IL-33 production in 2-week-old post-septic mice was associated with increased DNA methylation in lung epithelial cells. Notably, IL-33 treatment boosted the expansion of Tregs and induced immunosuppression in 2-week-old mice. Clinically, adults but not paediatric post-septic patients exhibited higher counts of Tregs and seral IL-33 levels. CONCLUSION AND IMPLICATIONS: These findings demonstrate a crucial and age-dependent role for IL-33 in post-sepsis immunosuppression. Thus, a better understanding of this process may lead to differential treatments for adult and paediatric sepsis.

Vascular complications with injection implants: treatment with hyperbaric chamber.

INTRODUCTION: The use of facial fillers has increased over the years because they are less invasive and present lower risks and faster results, along with shorter recovery time. OBJECTIVE: This study aimed to analyze the use of hyperbaric oxygen therapy as a possible treatment for vascular complications resulting from facial fillers. METHODOLOGY: This is a retrospective study of a series of patients treated at with hyperbaric oxygen therapy at the Brazilian Institute for Hyperbaric Oxygen Therapy (Instituto de Oxigenoterapia Hiperbárica do Brasil), after vascular complications from facial fillers. CONCLUSION: The association of oxygen therapy in a hyperbaric chamber with the conventional protocol for treating vascular occlusion from facial filler procedures was found to be effective.

Potential Fast COVID-19 Containment With Trehalose.

Countries worldwide have confirmed a staggering number of COVID-19 cases, and it is now clear that no country is immune to the SARS-CoV-2 infection. Resource-poor countries with weaker health systems are struggling with epidemics of their own and are now in a more uncertain situation with this rapidly spreading infection. Frontline healthcare workers are succumbing to the infection in their efforts to save lives. There is an urgency to develop treatments for COVID-19, yet there is limited clinical data on the efficacy of potential drug treatments. Countries worldwide implemented a stay-at-home order to "flatten the curve" and relieve the pressure on the health system, but it is uncertain how this will unfold after the economy reopens. Trehalose, a natural glucose disaccharide, is known to impair viral function through the autophagy system. Here, we propose trehalose as a potential preventative treatment for SARS-CoV-2 infection and transmission.

Analgesic and side effects of intravenous recombinant Phα1ß.

BACKGROUND: Intrathecal injection of voltage-sensitive calcium channel blocker peptide toxins exerts analgesic effect in several animal models of pain. Upon intrathecal administration, recombinant Phα1ß exerts the same analgesic effects as the those of the native toxin. However, from a clinical perspective, the intrathecal administration limits the use of anesthetic drugs in patients. Therefore, this study aimed to investigate the possible antinociceptive effect of intravenous recombinant Phα1ß in rat models of neuropathic pain, as well as its side effects on motor, cardiac (heart rate and blood pressure), and biochemical parameters. METHODS: Male Wistar rats and male Balb-C mice were used in this study. Giotto Biotech® synthesized the recombinant version of Phα1ß using Escherichia coli expression. In rats, neuropathic pain was induced by chronic constriction of the sciatic nerve and paclitaxel-induced acute and chronic pain. Mechanical sensitivity was evaluated using von Frey filaments. A radiotelemeter transmitter (TA11PA-C10; Data Sciences, St. Paul, MN, USA) was placed on the left carotid of mice for investigation of cardiovascular side effects. Locomotor activity data were evaluated using the open-field paradigm, and serum CKMB, TGO, TGP, LDH, lactate, creatinine, and urea levels were examined. RESULTS: Intravenous administration of recombinant Phα1ß toxin induced analgesia for up to 4 h, with ED50 of 0.02 (0.01-0.03) mg/kg, and reached the maximal effect (Emax = 100% antinociception) at a dose of 0.2 mg/kg. No significant changes were observed in any of the evaluated motor, cardiac or biochemical parameters. CONCLUSION: Our data suggest that intravenous administration of recombinant Phα1ß may be feasible for drug-induced analgesia, without causing any severe side effects.

Health Innovation Laboratories: towards strong Primary Health Care (PHC)in the Federal District of Brasilia. / Laboratórios de inovação em saúde: por uma Atenção Primária à Saúde forte no Distrito Federal, Brasil.

This article emphasizes PHC as a fundamental strategy for the sustainability of the SUS, based on its impact on health indicators. The attributes and requirements for Robust PHC are based on statements from researchers, policy makers and institutions, including the PAHO/WHO Representation in Brazil. The model proposed is the result of discussions with workers, researchers and health managers in Brazil, endorsing the commitments outlined in the Alma Ata Declaration. The article details the methodology of Health Innovation Laboratories used by PAHO/WHO in Brazil to systematize knowledge generated by innovative health experiences, including the Healthy Brasilia Project, an ongoing activity run by the Health Department of the Federal District (DF) of Brasilia, which has made important changes in the healthcare model, with emphasis on the expansion of Family Health Strategy coverage. This article analyzes the results of the Innovation Laboratories in Robust PHC in the FD that will be consolidated in a Case Study. The initiative aims to raise awareness amongst managers and health workers about innovation in health processes and policies that are essential for the sustainability of theSUS, focusing on the exchange of knowledge between peers about relevant initiatives in PHCin Brazil.

O artigo defende a APS como estratégia fundamental para a sustentabilidade do SUS, subsidiado em resultados sobre o impacto da APS nos indicadores de saúde e outras áreas. São apresentados os atributos e requisitos para uma APS Forte, defendidos por pesquisadores, formuladores de políticas e instituições, entre elas a Representação da OPAS/OMS no Brasil. O formato advogado é fruto de discussão realizada com trabalhadores, pesquisadores e gestores de saúde do país, reafirmando os compromissos da Declaração de Alma Ata. O artigo detalha a metodologia de Laboratórios de Inovação em Saúde, utilizada pela OPAS/OMS no Brasil para identificar e sistematizar experiências inovadoras em saúde, incluindo o acompanhamento do Projeto Brasília Saudável, da Secretaria de Saúde do DF, que vem apresentando importantes transformações no modelo de atenção à saúde, com ênfase na ampliação da cobertura da Estratégia Saúde da Família. O artigo resgata e analisa os trabalhos desenvolvidos pelo Laboratório de Inovação em APS Forte no DF até o momento e que serão sistematizados em Estudo de Caso. A iniciativa visa sensibilizar gestores e trabalhadores em saúde para a inovação em processos e políticas de saúde, sendo essencial para a sustentabilidade do SUS, privilegiando troca de conhecimentos entre pares sobre iniciativas adotadas na APS do Brasil.

Perivascular adipose tissue contributes to lethal sepsis-induced vasoplegia in rats.

It is well established that sepsis induces vascular hyporesponsiveness to vasoconstrictors. Perivascular adipose tissue (PVAT) displays anti-contractile action in various blood vessels. We hypothesized that sepsis would increase the anti-contractile effect of PVAT aggravating sepsis-induced vasoplegia. Male Wistar Hannover rats were subjected to lethal sepsis by cecal ligation and puncture (CLP) method. Aorta or PVAT were collected for functional or biochemical assays 6 h after CLP surgery. Functional experiments showed that sepsis increased the anti-contractile action of PVAT in both endothelium-intact and endothelium-denuded aortas. Carboxy-PTIO, L-NAME and ODQ reversed the hypocontractility mediated by PVAT in aortas from septic rats. Inhibition of nNOS and iNOS with 7-nitroindazole and 1400 W attenuated PVAT-mediated hypocontractility during sepsis. Similar results were found in the presence of indomethacin and Ro1138452, a selective prostacyclin IP receptor antagonist. However, neither tiron nor catalase affected phenylephrine-induced contraction in aortas from septic rats. Increased levels of superoxide anion (O2•-) and 6-keto-prostaglandin F1α (stable product of prostacyclin) were detected in PVAT from septic rats. In situ quantification of reactive oxygen species and nitric oxide (NO) using fluorescent dyes revealed increased levels of both in PVAT from septic rats. The novelty of our study is that PVAT contributes to sepsis-induced vasoplegia by releasing NO and prostacyclin. These findings suggested that signaling pathways in PVAT may be considered as potential novel pharmacological therapeutic targets during sepsis-induced vasoplegia.

Narrativas de participantes do Curso Introdução à Saúde dos Povos Indígenas: protagonismo e diversidade para o trabalho nos contextos indígenas / Narratives from participants in the Introduction to Indigenous Peoples' Health Course: protagonism and diversity for work in indigenous contexts

Há uma demanda de formação de profissionais para atuação nos contextos de saúde indígena e cresce o protagonismo indígena nos diversos campos de saber para tratar de suas próprias questões sociopolíticas. O presente artigo traz uma análise documental com abordagem qualitativa e análise temática de conteúdo. Os documentos analisados foram narrativas redigidas por participantes do curso Introdução à Saúde dos Povos Indígenas, ofertado por uma universidade, com participação de indígenas e não indígenas, lideranças, trabalhadores e estudantes da saúde de diversas regiões do Brasil. Analisaram-se três categorias de experiências: presença e protagonismo indígena na experiência do curso; aprendizagens no encontro de diversidades com metodologias ativas de ensino-aprendizagem; formação para o trabalho na saúde indígena. Percebeu-se que a utilização de metodologias ativas possibilitou um curso participativo, com valorização da presença e do protagonismo indígenas. Apresentou-se de forma inovadora ao ofertar espaços de construção de conhecimentos e de formação profissional de forma acessível, no modelo remoto, com valorização dos diferentes conhecimentos e trajetórias dos participantes.

There is a demand for training professionals to work in indigenous health contexts and indigenous protagonism is growing in different fields of knowledge to address their own sociopolitical issues. This article presents a documentary analysis with a qualitative approach and thematic content analysis. The documents analyzed were narratives written by participants in the Introduction to the Health of Indigenous Peoples course, offered by a university, with the participation of indigenous and non indigenous people, leaders, health workers and students from different regions of Brazil. Three categories of experiences were analyzed: indigenous presence and protagonism in the course experience; learning in the encounter of diversities with active teaching-learning methodologies; training for work in indigenous health. It was noticed that the use of active methodologies enabled a participatory course, valuing indigenous presence and protagonism. It presented an innovative way by offering spaces for knowledge construction and professional training in an accessible way, in a remote model, valuing the different knowledge and trajectories of the participants.

CCR5-Positive Inflammatory Monocytes are Crucial for Control of Sepsis.

Neutrophils and inflammatory monocytes control sepsis by migration to the site of infection via their chemokine receptors. CCR5 is a chemokine receptor that is not expressed on neutrophils and inflammatory monocytes under homeostatic conditions. However, it has been demonstrated that CCR5 can become expressed on these cells during different models of inflammation. In the present study, we investigated if CCR5 is also expressed on neutrophil and inflammatory monocytes during sepsis, exerting an important role in the migration of these cells to the infectious focus. Using cecal ligation and puncture model to induce polymicrobial sepsis, we demonstrated that the expression of CCR5 is induced on CD11bLy6GLy6C inflammatory monocytes, but not on neutrophils (CD11bLy6GLy6C). Furthermore, CCR5 plays an important role for the migration of the inflammatory monocytes to infection focus during sepsis. CCR5-expressing inflammatory monocytes migrate from the bone marrow to the circulation and then into the site of infection, where they phagocytize and kill the bacteria. Consequently, CCR5 mice showed increased systemic inflammatory response and mortality compared to wild-type mice. These data therefore demonstrate a hitherto unrecognized protective role of CCR5 in sepsis.

Thimet Oligopeptidase (EC 3.4.24.15) Key Functions Suggested by Knockout Mice Phenotype Characterization.

Thimet oligopeptidase (THOP1) is thought to be involved in neuropeptide metabolism, antigen presentation, neurodegeneration, and cancer. Herein, the generation of THOP1 C57BL/6 knockout mice (THOP1-/-) is described showing that they are viable, have estrus cycle, fertility, and a number of puppies per litter similar to C57BL/6 wild type mice (WT). In specific brain regions, THOP1-/- exhibit altered mRNA expression of proteasome beta5, serotonin 5HT2a receptor and dopamine D2 receptor, but not of neurolysin (NLN). Peptidomic analysis identifies differences in intracellular peptide ratios between THOP1-/- and WT mice, which may affect normal cellular functioning. In an experimental model of multiple sclerosis THOP1-/- mice present worse clinical behavior scores compared to WT mice, corroborating its possible involvement in neurodegenerative diseases. THOP1-/- mice also exhibit better survival and improved behavior in a sepsis model, but also a greater peripheral pain sensitivity measured in the hot plate test after bradykinin administration in the paw. THOP1-/- mice show depressive-like behavior, as well as attention and memory retention deficits. Altogether, these results reveal a role of THOP1 on specific behaviors, immune-stimulated neurodegeneration, and infection-induced inflammation.

Galectin-3 aggravates experimental polymicrobial sepsis by impairing neutrophil recruitment to the infectious focus.

Sepsis is an overwhelming systemic inflammation resulting from an uncontrolled infection that causes extensive tissue damage, organ dysfunction and eventually death. A growing body of evidence indicates that impaired neutrophil migration to the site of infection is associated with poor outcome in sepsis. Here we show that galectin-3 (Gal-3), an endogenous glycan-binding protein, plays a critical role in sepsis outcome. We found that serum Gal-3 concentration increased in patients with septic shock and mice undergoing sepsis induced by cecal ligation and puncture (CLP). Mice deficient in Gal-3 (Gal-3 KO) are more resistant to sepsis induced by CLP, showing lower levels of biochemical markers and neutrophil infiltration for organ injury/dysfunction than those observed in wild-type mice (WT). Furthermore, Gal-3 KO mice show an increased number of neutrophils in the primary focus of infection and reduced bacterial loads in the peritoneal cavity, blood, and lungs. Mechanistically, blood neutrophils from septic mice show higher levels of surface-bound Gal-3 than neutrophils from naive mice. The deficiency of Gal-3 was associated with increased rolling and adhesion of these cells in mesenteric venules. Our results indicate that Gal-3, secreted during sepsis, inhibits neutrophil migration into the infectious focus, which promotes the bacterial spread and worsens the outcome of sepsis.

The Atypical Chemokine Receptor ACKR2 is Protective Against Sepsis.

Sepsis is a systemic inflammatory response as a result of uncontrolled infections. Neutrophils are the first cells to reach the primary sites of infection, and chemokines play a key role in recruiting neutrophils. However, in sepsis chemokines could also contribute to neutrophil infiltration to vital organs leading to multiple organ failure. ACKR2 is an atypical chemokine receptor, which can remove and degrade inflammatory CC chemokines. The role of ACK2 in sepsis is unknown. Using a model of cecal ligation and puncture (CLP), we demonstrate here that ACKR2 deficient () mice exhibited a significant reduction in the survival rate compared with similarly treated wild-type (WT) mice. However, neutrophil migration to the peritoneal cavity and bacterial load were similar between WT and ACKR2 mice during CLP. In contrast, ACKR2 mice showed increased neutrophil infiltration and elevated CC chemokine levels in the lung, kidney, and heart compared with the WT mice. In addition, ACKR2 mice also showed more severe lesions in the lung and kidney than those in the WT mice. Consistent with these results, WT mice under nonsevere sepsis (90% survival) had higher expression of ACKR2 in these organs than mice under severe sepsis (no survival). Finally, the lungs from septic patients showed increased number of ACKR2 cells compared with those of nonseptic patients. Our data indicate that ACKR2 may have a protective role during sepsis, and the absence of ACKR2 leads to exacerbated chemokine accumulation, neutrophil infiltration, and damage to vital organs.