Local microvascular leakage promotes trafficking of activated neutrophils to remote organs.
Ontology highlight
ABSTRACT:
SUBMITTER: Owen-Woods C
PROVIDER: S-EPMC7190919 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Alcohol intoxication can increase inflammation and worsen injury, yet the mechanisms involved are not clear. We investigated whether acute alcohol intoxication increases microvascular permeability and investigated potential signaling mechanisms in endothelial cells that may be involved.Conscious rats received a 2.5 g/kg alcohol bolus via gastric catheters to produce acute intoxication. Microvascular leakage of intravenously administered fluorescein isothiocyanate (FITC)-conjugated albumin (FITC-albumin) from the mesenteric microcirculation was assessed by intravital microscopy. Endothelial-specific mechanisms were studied using cultured endothelial cell monolayers. Transendothelial electrical resistance (TER) served as an index of barrier function, before and after treatment with alcohol or its metabolite acetaldehyde. Pharmacologic agents were used to test the roles of alcohol metabolism, oxidative stress, p38 mitogen-activated protein kinase (MAPK), myosin light-chain kinase (MLCK), rho kinase (ROCK), and exchange protein activated by cAMP (Epac). VE-cadherin localization was investigated to assess junctional integrity. Rac1 and RhoA activation was assessed by ELISA assays.Alcohol significantly increased FITC-albumin extravasation from the mesenteric microcirculation. Alcohol also significantly decreased TER and disrupted VE-cadherin organization at junctions. Acetaldehyde significantly decreased TER, but inhibition of alcohol dehydrogenase or application of a superoxide dismutase mimetic failed to prevent alcohol-induced decreases in TER. Inhibition of p38 MAPK, but not MLCK or ROCK, significantly attenuated the alcohol-induced barrier dysfunction. Alcohol rapidly decreased GTP-bound Rac1 but not RhoA during the drop in TER. Activation of Epac increased TER, but did not prevent alcohol from decreasing TER. However, activation of Epac after initiation of alcohol-induced barrier dysfunction quickly resolved TER to baseline levels.Our results suggest that alcohol intoxication increases microvascular permeability to plasma proteins. The data also suggest the endothelial-specific mechanism involves the p38 MAPK, Rac1, and reorganization of VE-cadherin at junctions. Last, activation of Epac can quickly resolve alcohol-induced endothelial barrier dysfunction.
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Project description:The complement C5a pathway has been shown to be an important mediator of inflammation and tissue injury. To further understand the role of C5a receptor (C5aR) pathway in ischemia/reperfusion (I/R) injury, and to evaluate the potential of antagonizing C5aR to protect from I/R injury, we tested the effect of eliminating C5aR using C5aR knockout (KO) mice and their wild-type (WT) littermates in a superior mesenteric artery occlusion (SMAO) intestinal I/R injury model. C5aR KO and WT mice were subjected to SMAO or sham for 45 min. After 3 h of reperfusion, the percentage of injured ileal villi was twice as high in WT mice subjected to SMAO as compared with the C5aR KO mice. In addition, the number of neutrophils was 34% higher in WT mice subjected to SMAO as compared with the C5aR KO mice. Moreover, ileum and lung myeloperoxidase activities after SMAO were significantly higher in WT than C5aR KO mice. Apoptotic cell death was induced after reperfusion in WT-SMAO and was reduced by more than 50% in C5aR KO mice. The plasma level of TNF-alpha was increased approximately 3.74-fold in WT subjected to SMAO compared with sham. In contrast, the level was increased only approximately 1.18-fold in the C5aR KO mice subjected to SMAO. In conclusion, this study demonstrates that elimination of the C5aR pathway protects the intestine from I/R injury and diminishes intestine-derived pulmonary neutrophil sequestration. Blocking C5aR may be considered as a potential therapeutic intervention for I/R injury.
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Project description:Reperfusion injury is a very common complication of various indicated therapies such as the re-opening of vessels in the myocardium or brain as well as reflow in hemodynamic shutdown (cardiac arrest, severe trauma, aortic cross-clamping). The treatment and prevention of reperfusion injury has therefore been a topic of immense interest in terms of mechanistic understanding, the exploration of interventions in animal models and in the clinical setting in major prospective studies. While a wealth of encouraging results has been obtained in the lab, the translation into clinical success has met with mixed outcomes at best. Considering the still very high medical need, progress continues to be urgently needed. Multi-target approaches rationally linking interference with pathophysiological pathways as well as a renewed focus on aspects of microvascular dysfunction, especially on the role of microvascular leakage, are likely to provide new insights.
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Project description:It is widely understood that CRISPR-Cas9 technology is revolutionary, with well-recognized issues including the potential for off-target edits and the attendant need for spatiotemporal control of editing. Here, we describe a far-red light (FRL)-activated split-Cas9 (FAST) system that can robustly induce gene editing in both mammalian cells and mice. Through light-emitting diode-based FRL illumination, the FAST system can efficiently edit genes, including nonhomologous end joining and homology-directed repair, for multiple loci in human cells. Further, we show that FAST readily achieves FRL-induced editing of internal organs in tdTomato reporter mice. Finally, FAST was demonstrated to achieve FRL-triggered editing of the PLK1 oncogene in a mouse xenograft tumor model. Beyond extending the spectrum of light energies in optogenetic toolbox for CRISPR-Cas9 technologies, this study demonstrates how FAST system can be deployed for programmable deep tissue gene editing in both biological and biomedical contexts toward high precision and spatial specificity.
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Project description:Background and Purpose- Cerebral amyloid angiopathy (CAA) is a common small vessel disease that independently effects cognition in older individuals. The pathophysiology of CAA and CAA-related bleeding remains poorly understood. In this postmortem study, we explored whether blood-brain barrier leakage is associated with CAA and microvascular lesions. Methods- Eleven CAA cases (median [IQR] age=69 years [65-79 years], 8 males) and 7 cases without neurological disease or brain lesions (median [IQR] age=77 years [68-92 years], 4 males) were analyzed. Cortical sections were sampled from each lobe, and IgG and fibrin extravasation (markers of blood-brain barrier leakage) were assessed with immunohistochemistry. We hypothesized that IgG and fibrin extravasation would be increased in CAA cases compared with controls, that this would be more pronounced in parietooccipital brain regions compared with frontotemporal brain regions in parallel with the posterior predilection of CAA, and would be associated with CAA severity and number of cerebral microbleeds and cerebral microinfarcts counted on ex vivo magnetic resonance imaging of the intact brain hemisphere. Results- Our results demonstrated increased IgG positivity in the frontotemporal ( P=0.044) and parietooccipital ( P=0.001) cortex in CAA cases compared with controls. Within CAA cases, both fibrin and IgG positivity were increased in parietooccipital brain regions compared with frontotemporal brain regions ( P=0.005 and P=0.006, respectively). The percentage of positive vessels for fibrin and IgG was associated with the percentage of amyloid-β-positive vessels (Spearman ρ=0.71, P=0.015 and Spearman ρ=0.73, P=0.011, respectively). Moreover, the percentage of fibrin and IgG-positive vessels, but not amyloid-β-positive vessels, was associated with the number of cerebral microbleeds on magnetic resonance imaging (Spearman ρ=0.77, P=0.005 and Spearman ρ=0.70, P=0.017, respectively). Finally, we observed fibrin deposition in walls of vessels involved in cerebral microbleeds. Conclusions- Our results raise the possibility that blood-brain barrier leakage may be a contributory mechanism for CAA-related brain injury.
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Project description:ObjectiveTo investigate the replication of influenza A virus A/Puerto Rico/8/34 (H1N1) in pulmonary microvascular endothelial cells and its effect on endothelial barrier function.MethodsHuman pulmonary microvascular endothelial cells were infected with influenza A/Puerto Rico/8/34 (H1N1) virus. Plaque reduction assay, real-time quantitative PCR, immunofluorescence staining, and western blot were used to elucidate the replication process of virus-infected endothelial cells. In addition, real-time quantitative PCR was used to detect the relative expression levels of mRNA of some inflammatory factors. The endothelial resistance assay was used to determine the permeability of the endothelial monolayer. Excavation and analysis of data from open databases, such as the GeneCards database, DAVID Bioinformatics Resources, STRING search tool, and DGIdb database determined the genes, proteins, and signal pathways related to microvascular leakage caused by the H1N1 virus, and predicted the drugs that could be effective for treatment.ResultsIn vitro experiments showed that the influenza virus can infect endothelial cells, leading to a significant increase in the permeability of pulmonary microvascular endothelial cells and the release of pro-inflammatory cytokines, but does not efficiently replicate in endothelial cells. A total of 107 disease-related target genes were obtained from the Gene-cards database. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis showed that these genes mainly affected the pathways related to "Inflammatory bowel disease" (IBD), "Chagas disease" (American trypanosomiasis), "Influenza A", and also played a key role in anti-inflammation and regulation of immunity. After enrichment analysis, 46 hub genes were screened. A total of 42 FDA-approved drugs corresponding to the hub genes were screened from the DGIdb database, and these could be formulated for topical application. In addition, these drugs can be used to treat other diseases, including cancer, inflammatory diseases, immune system disorders, and cardiovascular diseases.ConclusionH1N1 influenza virus affects the barrier function of endothelial cells indirectly. Combined with bioinformatics tools, we can better understand the possible mechanism of action of influenza A (H1N1) virus causing pulmonary microvascular leakage and provide new clues for the treatment of pulmonary microvascular leakage.
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