Bacterial spillage into a sterile environment following intestinal hollow-organ perforation leads to peritonitis and fulminant sepsis. Outcome of sepsis critically depends on macrophage activation by ...extracellular ATP-release and associated autocrine signalling via purinergic receptors. ATP-release mechanisms, however, are poorly understood. Here, we show that TLR-2 and -4 agonists trigger ATP-release via Connexin-43 hemichannels in macrophages leading to poor sepsis survival. In humans, Connexin-43 was upregulated on macrophages isolated from the peritoneal cavity in patients with peritonitis but not in healthy controls. Using a murine peritonitis/sepsis model, we identified increased Connexin-43 expression in peritoneal and hepatic macrophages. Conditional
mice were developed to specifically assess Connexin-43 impact in macrophages. Both macrophage-specific Connexin-43 deletion and pharmacological Connexin-43 blockade were associated with reduced cytokine secretion by macrophages in response to LPS and CLP, ultimately resulting in increased survival. In conclusion, inhibition of autocrine Connexin-43-dependent ATP signalling on macrophages improves sepsis outcome.
Antimicrobial host defense is dependent on the rapid recruitment of inflammatory cells to the site of infection, the elimination of invading pathogens, and the efficient resolution of inflammation ...that minimizes damage to the host. The peritoneal cavity provides an accessible and physiologically relevant system where the delicate balance of these processes may be studied. Here, we describe murine models of peritoneal inflammation that enable studies of competent antimicrobial immunity and inflammation-associated tissue damage as a consequence of recurrent bacterial challenge. The inflammatory hallmarks of these models reflect the clinical and molecular features of peritonitis seen in renal failure patients on peritoneal dialysis. The development of these models relies on the preparation of a cell-free supernatant derived from an isolate of Staphylococcus epidermidis (termed SES). Intraperitoneal administration of SES induces a Toll-like receptor 2-driven acute inflammatory response that is characterized by an initial transient influx of neutrophils that are replaced by a more sustained recruitment of mononuclear cells and lymphocytes. Adaptation of this model using a repeated administration of SES allows investigations into the development of adaptive immunity and the hallmarks associated with tissue remodelling and fibrosis. These models are therefore clinically relevant and provide exciting opportunities to study innate and adaptive immunity and the response of the stromal tissue compartment to bacterial infection and the ensuing inflammatory reaction.
The local microenvironment shapes macrophage differentiation in each tissue. We hypothesized that in the peritoneum, local factors in addition to retinoic acid can support GATA6-driven ...differentiation and function of peritoneal large cavity macrophages (LCMs). We found that soluble proteins produced by mesothelial cells lining the peritoneal cavity maintained GATA6 expression in cultured LCMs. Analysis of global gene expression of isolated mesothelial cells highlighted mesothelin (Msln) and its binding partner mucin 16 (Muc16) as candidate secreted ligands that potentially regulate GATA6 expression in peritoneal LCMs. Mice deficient for either of these molecules showed diminished GATA6 expression in peritoneal and pleural LCMs that was most prominent in aged mice. The more robust phenotype in older mice suggested that monocyte-derived macrophages were the target of Msln and Muc16. Cell transfer and bone marrow chimera experiments supported this hypothesis. We found that lethally irradiated Msln-/- and Muc16-/- mice reconstituted with wild-type bone marrow had lower levels of GATA6 expression in peritoneal and pleural LCMs. Similarly, during the resolution of zymosan-induced inflammation, repopulated peritoneal LCMs lacking expression of Msln or Muc16 expressed diminished GATA6. These data support a role for mesothelial cell-produced Msln and Muc16 in local macrophage differentiation within large cavity spaces such as the peritoneum. The effect appears to be most prominent on monocyte-derived macrophages that enter into this location as the host ages and also in response to infection.
To determine whether synthetic phosphorylated hexa-acyl disaccharides provide antimicrobial protection in clinically relevant models of bacterial infection.
Laboratory study.
University laboratory.
...BALB/c, C57BL/10J, and C57BL/10ScNJ mice.
Mice were treated with lactated Ringer's (vehicle) solution, monophosphoryl lipid A, or phosphorylated hexa-acyl disaccharides at 48 and 24 hours prior to intraperitoneal Pseudomonas aeruginosa or IV Staphylococcus aureus infection. Leukocyte recruitment, cytokine production, and bacterial clearance were measured 6 hours after P. aeruginosa infection. In the systemic S. aureus infection model, one group of mice was monitored for 14-day survival and another for S. aureus tissue burden at 3 days postinfection. Duration of action for 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide was determined at 3, 10, and 14 days using a model of intraperitoneal P. aeruginosa infection. Effect of 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide on in vivo leukocyte phagocytosis and respiratory burst was examined. Leukocyte recruitment, cytokine production, and bacterial clearance were measured after P. aeruginosa infection in wild-type and toll-like receptor 4 knockout mice treated with 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide or vehicle to assess receptor specificity.
During intraperitoneal P. aeruginosa infection, phosphorylated hexa-acyl disaccharides significantly attenuated infection-induced hypothermia, augmented leukocyte recruitment and bacterial clearance, and decreased cytokine production. At 3 days post S. aureus infection, bacterial burden in lungs, spleen, and kidneys was significantly decreased in mice treated with monophosphoryl lipid A or phosphorylated hexa-acyl disaccharides, which was associated with improved survival. Leukocyte phagocytosis and respiratory burst functions were enhanced after treatment with monophosphoryl lipid A or phosphorylated hexa-acyl disaccharides. A time course study showed that monophosphoryl lipid A- and 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide-mediated protection against P. aeruginosa lasts for up to 10 days. Partial loss of augmented innate antimicrobial responses was observed in toll-like receptor 4 knockout mice treated with 3-deacyl 6-Acyl phosphorylated hexa-acyl disaccharide.
Phosphorylated hexa-acyl disaccharides significantly augment resistance against clinically relevant Gram-negative and Gram-positive infections via enhanced leukocyte recruitment, phagocytosis, and respiratory burst functions of innate leukocytes. Improved antimicrobial protection persists for up to 10 days and is partially mediated through toll-like receptor 4.
Mouse models of ovarian cancer commonly transfer large numbers of tumor cells into the peritoneal cavity to establish experimental metastatic disease, which may not adequately model early metastatic ...spread from a primary tumor site. We hypothesized we could develop an ovarian cancer model that predictably represents micro-metastatic disease.
Murine ID8VEGF ovarian cancer cells were transduced to express enhanced luciferase (eLuc) to enable intravital detection of microscopic disease burden and injected beneath the ovarian bursa of C57Bl/6 mice. At 6 or 10 weeks after orthotopic injection, when mice had detectable metastases, hysterectomy and bilateral salpingo-oophorectomy was performed to remove all macroscopic disease, and survival monitored. Immunohistochemistry and gene expression profiling were performed on primary and metastatic tumors.
eLuc-transduced ID8VEGF cells were brighter than cells transduced with standard luciferase, enabling in vivo visualization of microscopic intra-abdominal metastases developing after orthotopic injection. Primary surgical cytoreduction removed the primary tumor mass but left minimal residual disease in all mice. Metastatic sites that developed following orthotopic injection were similar to metastatic human ovarian cancer sites. Gene expression and immune infiltration were similar between primary and metastatic mouse tumors. Surgical cytoreduction prolonged survival compared to no surgery, with earlier cytoreduction more beneficial than delayed, despite micro-metastatic disease in both settings.
Mice with primary ovarian tumors established through orthotopic injection develop progressively fatal metastatic ovarian cancer, and benefit from surgical cytoreduction to remove bulky disease. This model enables the analysis of therapeutic regimens designed to target and potentially eradicate established minimal residual disease.
•Orthotopic tumor inoculation establishes a progressive primary tumor that disseminates to form metastatic disease.•Metastases recapitulate the immune profile of the originating primary tumor and metastatic human disease.•Surgical cytoreduction of primary tumors prolongs survival.•Surgical cytoreduction retains micro-metastatic disease for evaluating therapeutic interventions.
In vivo imaging and actuation of a swarm of magnetic helical microswimmers by external magnetic fields (less than 10 mT) in deep tissue is demonstrated for the first time. This constitutes a major ...milestone in the field, yielding a generation of micrometer‐scale transporters with numerous applications in biomedicine including synthetic biology, assisted fertilization, and drug/gene delivery.
The biology of serous cavity macrophages Bain, Calum C.; Jenkins, Stephen J.
Cellular immunology,
August 2018, 2018-08-00, 20180801, Letnik:
330
Journal Article
Recenzirano
Odprti dostop
•The serous cavities contain phenotypically and functionally distinct macrophage subsets.•The developmental origin of these subsets is distinct and influenced by age and sex.•Peritoneal macrophages ...play vital homeostatic roles but can contribute to inflammatory disease.•The goal is to understand the role of discrete macrophage subsets to pathology of the serous cavities and the organs they cover.•Peritoneal macrophage subsets may be targets for novel therapeutics in conditions such as endometriosis.
For decades, it has been known that the serous cavities, which include the peritoneal, pleural and pericardial cavities, harbour large numbers of macrophages. In particular, due to the ease of isolating these cells, the peritoneal cavity has been used as a convenient source of macrophages to examine many facets of macrophage biology over the last 50–60 years. Despite this, it is only recently that the true heterogeneity of serous cavity mononuclear phagocyte compartment, which includes macrophages and dendritic cells, has been revealed. Advances in technologies such as multi-parameter flow cytometry and the ‘OMICs’ revolution have uncovered the presence of distinct populations of mononuclear phagocytes in the serous cavities. Given that peritoneal macrophages have been implicated in many pathologies, including peritonitis, pancreatitis, endometriosis and acute liver injury, it is imperative to understand the biology of these cells. Here, we review the recent advances in understanding the identity, origin and function of discrete serous cavity mononuclear phagocyte subsets in homeostasis and how these may change when homeostasis is perturbed, focusing on peritoneal and pleural cavities and highlighting differences in the mononuclear phagocytes found in each.
Endometriosis is a disorder associated with a general inflammatory response in the peritoneal cavity. Oxidative stress is a potential factor involved in the pathophysiology of this disease, and ...reactive oxygen species (ROS) are implicated in this process. Indeed, in healthy individuals, ROS and antioxidants are in balance, but when balance is tipped toward an overabundance of ROS, oxidative stress occurs and can impact the entire reproductive lifespan of a woman. Reactive oxygen species are intermediaries produced by normal oxygen metabolism but are known to have deleterious effects. Excessive release of ROS induces cellular damage and alters cellular function by regulating protein activity and gene expression, leading to harmful effects. To protect themselves, cells have developed antioxidant systems to limit production of ROS, inactivate them, and repair cell damage. Understanding of the control of hemoglobin, heme, and iron-induced redox balance in endometriosis led us to propose a number of hypotheses to explain why oxidative stress is induced in case of pelvic endometriosis. Erythrocytes, apoptotic endometrial tissue, and cell debris transplanted into the peritoneal cavity by menstrual reflux and macrophages have all been cited as potential inducers of oxidative stress. Erythrocytes are likely to release pro-oxidant and proinflammatory factors, such as hemoglobin and its highly toxic by-products heme and iron, into the peritoneal environment. Iron and heme are essential to living cells, but unless appropriately chelated, free iron, and to a lesser extent heme, play a key role in the formation of deleterious ROS.
Tissue-resident macrophages require specific milieus for the maintenance of defining gene-expression programs. Expression of the transcription factor GATA6 is required for the homeostasis, function ...and localization of peritoneal cavity-resident macrophages. Gata6 expression is maintained in a non-cell autonomous manner and is elicited by the vitamin A metabolite, retinoic acid. Here, we found that the GATA6 transcriptional program is a common feature of macrophages residing in all visceral body cavities. Retinoic acid-dependent and -independent hallmark genes of GATA6+ macrophages were induced by mesothelial and fibroblastic stromal cells that express the transcription factor Wilms’ Tumor 1 (WT1), which drives the expression of two rate-limiting enzymes in retinol metabolism. Depletion of Wt1+ stromal cells reduced the frequency of GATA6+ macrophages in the peritoneal, pleural and pericardial cavities. Thus, Wt1+ mesothelial and fibroblastic stromal cells constitute essential niche components supporting the tissue-specifying transcriptional landscape and homeostasis of cavity-resident macrophages.
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•Large cavity macrophages (LCMs) reside in peritoneal, pleural, and pericardial spaces•LCMs express the retinoic-acid-responsive transcription factor GATA6•Wt1+ stromal cells generate retinoic acid and maintain LCM GATA6 expression•Genetic ablation of Wt1+ stroma elicits loss of LCM across cavity spaces
Buechler et al. show that large cavity macrophages (LCMs), which rely on the retinoic-acid-responsive transcription factor GATA6, are found in all visceral cavity spaces. Wt1+ mesothelial and fibroblastic stromal cells maintain the LCM transcriptome in part via the generation of retinoic acid and are required for LCM homeostasis in vivo.
Acinetobacter baumannii infection has become a major cause of healthcare-associated infection and a critical pathogen in the WHO antimicrobial resistance research and development priority list. ...Catheter-related septicemia is one of the major clinical manifestations of A. baumannii infection associated with high morbidity and mortality. In this study, we used a clinical A. baumannii strain (LAC-4) that is hypervirulent to immunocompetent C57BL/6 and BALB/c mice and established a mouse model of intraperitoneal (i.p.) A. baumannii infection. Our study showed that i.p. LAC-4 infection of C57BL/6 and BALB/c mice induces a lethal or sublethal infection with high bacterial burdens in peritoneal cavity, blood and tissues and the infected mice either succumbed to the infection within 24 hours or completely recovered from the infection. The infection induces acute peritoneal recruitment of neutrophils and other innate immune cells, and the local and systemic production of proinflammatory cytokines and chemokines (IL-1β, IL-5, IL-6, TNF-α, RANTES, MIP-1β, MCP-1, KC and IL-10). Mechanistic studies suggest an important role of macrophages in the host innate defense in this model in that in vitro stimulation of peritoneal macrophages with killed LAC-4 induced a similar pattern of cytokine/chemokine responses to those in the infected mice, and depletion of peritoneal macrophages rendered the mice significantly more susceptible to the infection. Thus, this mouse infection model will provide an alternative and useful tool for future pathogenesis studies of A. baumannii-associated septicemia and identification and characterization of important virulence factors, as well as serve as a surrogate model for rapid evaluation of novel therapeutics and vaccines for this emerging infectious agent.
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