RESUMEN
Sterile inflammation after injury is important for tissue restoration. In injured human and mouse tissues, macrophages were recently found to accumulate perivascularly. This study investigates if macrophages adopt a mural cell phenotype important for restoration after ischemic injury. Single-cell RNA sequencing of fate-mapped macrophages from ischemic mouse muscles demonstrates a macrophage-toward-mural cell switch of a subpopulation of macrophages with downregulated myeloid cell genes and upregulated mural cell genes, including PDGFRß. This observation was further strengthened when including unspliced transcripts in the analysis. The macrophage switch was proven functionally relevant, as induction of macrophage-specific PDGFRß deficiency prevented their perivascular macrophage phenotype, impaired vessel maturation and increased vessel leakiness, which ultimately reduced limb function. In conclusion, macrophages in adult ischemic tissue were demonstrated to undergo a cellular program to morphologically, transcriptomically and functionally resemble mural cells while weakening their macrophage identity. The macrophage-to-mural cell-like phenotypic switch is crucial for restoring tissue function and warrants further exploration as a potential target for immunotherapies to enhance healing.
Asunto(s)
Modelos Animales de Enfermedad , Isquemia , Macrófagos , Animales , Macrófagos/metabolismo , Macrófagos/inmunología , Isquemia/metabolismo , Isquemia/patología , Isquemia/genética , Fenotipo , Músculo Esquelético/metabolismo , Músculo Esquelético/patología , Músculo Esquelético/lesiones , Cicatrización de Heridas/genética , Cicatrización de Heridas/fisiología , Ratones Endogámicos C57BL , Ratones , Masculino , Miembro Posterior/irrigación sanguínea , Neovascularización Fisiológica/genética , Regulación hacia Arriba , Transcriptoma , Análisis de la Célula Individual , Biomarcadores/metabolismo , Recuperación de la Función , Ratones NoqueadosRESUMEN
Treatments of colitis, inflammation of the intestine, rely on induction of immune suppression associated with systemic adverse events, including recurrent infections. This treatment strategy is specifically problematic in the increasing population of patients with cancer with immune checkpoint inhibitor (ICI)-induced colitis, as immune suppression also interferes with the ICI-treatment response. Thus, there is a need for local-acting treatments that reduce inflammation and enhance intestinal healing. Here, we investigated the effect and safety of bacterial delivery of short-lived immunomodulating chemokines to the inflamed intestine in mice with colitis. Colitis was induced by dextran sulfate sodium (DSS) alone or in combination with ICI (anti-PD1 and anti-CTLA-4), and Limosilactobacillus reuteri R2LC (L. reuteri R2LC) genetically modified to express the chemokine CXCL12-1α (R2LC_CXCL12, emilimogene sigulactibac) was given perorally. In addition, the pharmacology and safety of the formulated drug candidate, ILP100-Oral, were evaluated in rabbits. Peroral CXCL12-producing L. reuteri R2LC significantly improved colitis symptoms already after 2 days in mice with overt DSS and ICI-induced colitis, which in benchmarking experiments was demonstrated to be superior to treatments with anti-TNF-α, anti-α4ß7, and corticosteroids. The mechanism of action involved chemokine delivery to Peyer's patches (PPs), confirmed by local CXCR4 signaling, and increased numbers of colonic, regulatory immune cells expressing IL-10 and TGF-ß1. No systemic exposure or engraftment could be detected in mice, and product feasibility, pharmacology, and safety were confirmed in rabbits. In conclusion, peroral CXCL12-producing L. reuteri R2LC efficiently ameliorates colitis, enhances mucosal healing, and has a favorable safety profile.NEW & NOTEWORTHY Colitis symptoms are efficiently reduced by peroral administration of probiotic bacteria genetically modified to deliver CXCL12 locally to the inflamed intestine in several mouse models.
Asunto(s)
Quimiocina CXCL12 , Colitis , Sulfato de Dextran , Modelos Animales de Enfermedad , Limosilactobacillus reuteri , Animales , Colitis/inmunología , Colitis/inducido químicamente , Colitis/tratamiento farmacológico , Colitis/terapia , Colitis/metabolismo , Ratones , Quimiocina CXCL12/metabolismo , Quimiocina CXCL12/genética , Administración Oral , Conejos , Probióticos/administración & dosificación , Ratones Endogámicos C57BL , Femenino , Colon/metabolismo , Colon/microbiología , Colon/inmunología , MasculinoRESUMEN
Increased levels of the anti-inflammatory peptide Catestatin (CST), a cleavage product of the pro-hormone chromogranin A, correlate with less severe outcomes in hypertension, colitis, and diabetes. However, it is unknown how CST reduces the infiltration of monocytes and macrophages (MÏs) in inflamed tissues. Here, it is reported that CST blocks leukocyte migration toward inflammatory chemokines. By in vitro and in vivo migration assays, it is shown that although CST itself is chemotactic, it blocks migration of monocytes and neutrophils to inflammatory attracting factor CC-chemokine ligand 2 (CCL2) and C-X-C motif chemokine ligand 2 (CXCL2). Moreover, it directs CX3 CR1+ MÏs away from pancreatic islets. These findings suggest that the anti-inflammatory actions of CST are partly caused by its regulation of chemotaxis.
Asunto(s)
Quimiotaxis de Leucocito , Quimiotaxis , Antiinflamatorios/farmacología , Quimiocina CCL2/farmacología , Quimiocinas/farmacología , Cromogranina A/farmacología , Ligandos , Neutrófilos , Fragmentos de Péptidos , Péptidos/farmacologíaRESUMEN
The tissue microenvironment in the mouse pancreas has been shown to promote very different polarizations of resident macrophages with islet-resident macrophages displaying an inflammatory "M1" profile and macrophages in the exocrine tissue mostly displaying an alternatively activated "M2" profile. The impact of this polarization on tissue homeostasis and diabetes development is unclear. In this study, the ability of pancreas-resident macrophages to phagocyte bacterial and endogenous debris was investigated. Mouse endocrine and exocrine tissues were separated, and tissue-resident macrophages were isolated by magnetic immunolabeling. Isolated macrophages were subjected to flow cytometry for polarization markers and qPCR for phagocytosis-related genes. Functional in vitro investigations included phagocytosis and efferocytosis assays using pH-sensitive fluorescent bacterial particles and dead fluorescent neutrophils, respectively. Intravital confocal imaging of in situ phagocytosis and efferocytosis in the pancreas was used to confirm findings in vivo. Gene expression analysis revealed no significant overall difference in expression of most phagocytosis-related genes in islet-resident vs. exocrine-resident macrophages included in the analysis. In this study, pancreas-resident macrophages were shown to differ in their ability to phagocyte bacterial and endogenous debris depending on their microenvironment. This difference in abilities may be one of the factors polarizing islet-resident macrophages to an inflammatory state since phagocytosis has been found to imprint macrophage heterogeneity. It remains unclear if this difference has any implications in the development of islet dysfunction or autoimmunity.
Asunto(s)
Macrófagos/fisiología , Páncreas/citología , Fagocitosis/fisiología , Animales , Apoptosis/fisiología , Macrófagos/citología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Neutrófilos/citologíaRESUMEN
[Figure: see text].
Asunto(s)
Quimiocina CXCL12/metabolismo , Isquemia/terapia , Macrófagos/fisiología , Músculo Esquelético/irrigación sanguínea , Óxido Nítrico Sintasa de Tipo II/metabolismo , Flujo Sanguíneo Regional/fisiología , Animales , Movimiento Celular , Proliferación Celular , Quimiocina CCL2/metabolismo , Humanos , Isquemia/fisiopatología , Macrófagos/citología , Macrófagos/metabolismo , Ratones , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo II/deficiencia , Óxido Nítrico Sintasa de Tipo III/metabolismo , Plásmidos/metabolismo , Receptores CCR2/metabolismo , Receptores CXCR4/metabolismoRESUMEN
OBJECTIVE: The full spectrum of coronavirus disease 2019 (COVID-19) infection ranges from asymptomatic to acute respiratory distress syndrome, characterized by hyperinflammation and thrombotic microangiopathy. The pathogenic mechanisms are poorly understood, but emerging evidence suggest that excessive neutrophil extracellular trap (NET) formation plays a key role in COVID-19 disease progression. Here, we evaluate if circulating markers of NETs are associated with COVID-19 disease severity and clinical outcome, as well as to markers of inflammation and in vivo coagulation and fibrinolysis. Approach and Results: One hundred six patients with COVID-19 with moderate to severe disease were enrolled shortly after hospital admission and followed for 4 months. Acute and convalescent plasma samples as well as plasma samples from 30 healthy individuals were assessed for markers of NET formation: citrullinated histone H3, cell-free DNA, NE (neutrophil elastase). We found that all plasma levels of NET markers were elevated in patients with COVID-19 relative to healthy controls, that they were associated with respiratory support requirement and short-term mortality, and declined to those found in healthy individuals 4 months post-infection. The levels of the NET markers also correlated with white blood cells, neutrophils, inflammatory cytokines, and C-reactive protein, as well as to markers of in vivo coagulation, fibrinolysis, and endothelial damage. CONCLUSIONS: Our findings suggest a role of NETs in COVID-19 disease progression, implicating their contribution to an immunothrombotic state. Further, we observed an association between circulating markers of NET formation and clinical outcome, demonstrating a potential role of NET markers in clinical decision-making, as well as for NETs as targets for novel therapeutic interventions in COVID-19.
Asunto(s)
COVID-19/sangre , COVID-19/complicaciones , Trampas Extracelulares/metabolismo , Anciano , Biomarcadores/sangre , Síndrome de Liberación de Citoquinas/sangre , Progresión de la Enfermedad , Endotelio Vascular/metabolismo , Femenino , Fibrinólisis , Humanos , Inflamación/sangre , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Trombosis/virologíaRESUMEN
The aim of this study was to compare the prevalence of different virulence factor (VF) genes in extended-spectrum beta-lactamase (ESBL) producing Escherichia coli strains isolated from the Baltic Sea region. A total of 432 strains of phenotypically ESBL positive E. coli were collected from 20 institutions located in Estonia, Latvia, Lithuania, and the region of St. Petersburg in Russia from January to May 2012 and analyzed for phylogenetic group and prevalence of 23 VF genes. The strains were collected from clinical material (urine, blood, wound, and respiratory tract). Bacterial isolates were compared according to phylogenetic group, clinical material, and geographical origin. Most of the VF genes were concentrated within phylogenetic group B2 and/or D. When comparing strains isolated from different countries, it was found that strains originating from Estonia and Latvia belonged mainly to group B2 and strains from Lithuania and Russia mainly to groups B2 and D. The P-fimbrial adhesin gene papEF was more prevalent in Russian strains, colicin gene cvaC in Lithuanian strains, and capsular gene kpsMTII in Latvian strains; serum resistant gene traT was less prevalent in Estonian strains. The regional differences of VF genes remained statistically significant after taking into account the phylogenetic distribution in the countries.
Asunto(s)
Toxinas Bacterianas/genética , Escherichia coli/clasificación , Escherichia coli/fisiología , Factores de Virulencia/genética , Microbiología del Agua , beta-Lactamasas/metabolismo , Genes Bacterianos/genética , Océanos y Mares , Especificidad de la EspecieRESUMEN
The spread of carbapenemase-producing Enterobacteriaceae is a global problem; however, no exact data on the epidemiology of carbapenemase in the Baltic countries and St. Petersburg area is available. We aimed to evaluate the epidemiology of carbapenemase-producing Escherichia coli and Klebsiella pneumoniae in the Baltic States and St. Petersburg, Russia, and to compare the different methods for carbapenemase detection. From January to May 2012, all K. pneumoniae (n = 1983) and E. coli (n = 7774) clinical isolates from 20 institutions in Estonia, Latvia, Lithuania, and St. Petersburg, Russia were screened for carbapenem susceptibility. The IMP, VIM, GIM, NDM, KPC, and OXA-48 genes were detected using real-time PCR and the ability to hydrolyze ertapenem was determined using MALDI-TOF MS. Seventy-seven strains were found to be carbapenem nonsusceptible. From these, 15 K. pneumoniae strains hydrolyzed ertapenem and carried the bla NDM gene. All of these strains carried integron 1 and most carried integron 3 as well as genes of the CTX-M-1 group. No carbapenemase-producing E. coli or K. pneumoniae strains were found in Estonia, Latvia, or Lithuania; however, NDM-positive K. pneumoniae was present in the hospital in St. Petersburg, Russia. A MALDI-TOF MS-based assay is a suitable and cost-effective method for the initial confirmation of carbapenemase production.