Effect of Egyptian Spitting Cobra Naja nubiae Crude Venom on Immunogenic Activity of Rats.

Snakes show defensive activities, often counting visual or auditory displays against an aggressor. The study observed what happens to rats administered subcutaneously sub-lethal doses of crude venom Naja nubiae. The pro-inflammatory cytokines, such as tumor necrosis alpha (TNF-α) and interleukin-6 (IL-6), as well as the anti-inflammatory cytokines such as interleukin-10 (IL-10), and inflammatory mediator's prostaglandin E-2 (PG-E2), were evaluated. Vascular permeability (VP) was employed to assess how leaky or permeable blood vessels are in various tissues and organs, including the rat peritoneal cavity and lymphoid organs. Lymphoid organs' histological alterations brought on by Nubiae venom. The study found that the two venom doses-1/4 and 1/2 LD50-induced high levels of inflammatory activity as evidenced by the production of inflammatory cytokines. These findings demonstrated that venom enhanced innate immunity through specifically increased T helper cells, IL-6, TNF-α, IL-10, and PG-E2. The results reveal whether the venom has an immunomodulatory effect and promotes inflammation. The data have a substantial impact on the development of new drugs and treatments for inflammatory conditions.

SCUBE2 regulates adherens junction dynamics and vascular barrier function during inflammation.

AIMS: SCUBE2 (Signal peptide-CUB-epidermal growth factor-like domain-containing protein 2) is a secreted or membrane-bound protein originally identified from endothelial cells (ECs). Our previous work showed that SCUBE2 forms a complex with E-cadherin and stabilizes epithelial adherens junctions (AJs) to promote epithelial phenotypes. However, it remains unclear whether SCUBE2 also interacts with vascular endothelial (VE)-cadherin and modulates EC barrier function. In this study, we investigated whether and how SCUBE2 in ECs regulates vascular barrier maintenance. METHODS AND RESULTS: We showed that SCUBE2 colocalized and interacted with VE-cadherin and VE-protein tyrosine phosphatase (VE-PTP) within EC AJs. Furthermore, SCUBE2 knockdown disrupted EC AJs and increased EC permeability. Expression of EC SCUBE2 was suppressed at both mRNA and protein levels via the nuclear factor-κB (NF-κB) signaling pathway in response to pro-inflammatory cytokines or permeability-inducing agents. In line with these findings, EC-specific deletion of Scube2 (EC-KO) in mice impaired baseline barrier function and worsened vascular leakiness of peripheral capillaries after local injection of histamine or vascular endothelial growth factor. EC-KO mice were also sensitive to pulmonary vascular hyperpermeability and leukocyte infiltration in response to acute endotoxin- or influenza virus-induced systemic inflammation. Meanwhile, EC-specific SCUBE2-overexpressing mice were protected from these effects. Molecular studies suggested that SCUBE2 acts as a scaffold molecule enabling VE-PTP to dephosphorylate VE-cadherin, which prevents VE-cadherin internalization and stabilizes EC AJs. As such, loss of SCUBE2 resulted in hyperphosphorylation of VE-cadherin at tyrosine 685, which led to its endocytosis, thus destabilizing EC AJs and reducing barrier function. All of these effects were exacerbated by inflammatory insults. CONCLUSIONS: We found that SCUBE2 contributes to vascular integrity by recruiting VE-PTP to dephosphorylate VE-cadherin and stabilize AJs, thereby promoting EC barrier function. Moreover, our data suggest that genetic overexpression or pharmacological upregulation of SCUBE2 may help to prevent vascular leakage and edema in inflammatory diseases.

Microcirculation, endothelium and glycocalyx changes associated with the use of milrinone in children with septic shock.

Background: The goal of fluid resuscitation and the use of inotropes in septic shock has traditionally focused on improving blood pressure and cardiac output, without considering the microcirculatory changes. Reaching macrocirculatory goals but with persistent microcirculatory abnormalities (hemodynamic incoherence) in septic shock has been associated with greater organ dysfunction and mortality. The objective of this study was to evaluate the microcirculation (flow and capillary density) and endothelial glycocalyx changes associated with the use of milrinone in children with septic shock, as well as their relationship with clinical variables and organ dysfunction. Methods: A prospective cohort study from February 2022 to January 2023 at a university hospital (Fundación Cardioinfantil-Instituto de Cardiología). Sublingual video microscopy was used to evaluate capillary density, microvascular flow rates and perfused boundary region (PBR-inverse parameter of glycocalyx thickness-abnormal if >2.0 microns). The primary outcome was the association between microcirculation and endothelial glycocalyx changes related to the use of milrinone. Results: A total of 140 children with a median age of two years [interquartile range (IQR) 0.58-12.1] were included. About 57.9% (81/140) of the patients received milrinone infusions. Twenty-four hours after receiving milrinone, the patients maintained functional capillary density (P<0.01) and capillary recruitment capacity (P=0.04) with no changes in capillary blood volume versus those who did not receive milrinone. Children under two years old who received milrinone had better 4-6-micron capillary density than older children [odds ratio (OR) 0.33; 95% confidence interval (95% CI): 0.12-0.89; P=0.02] and less endothelial glycocalyx degradation [adjusted OR (aOR) 0.34 95% CI: 0.11-0.99; P=0.04]. These changes persisted despite elevated ferritin (aOR 0.41; 95% CI: 0.18-0.93; P=0.03). Prolonged capillary refill and elevated lactate were correlated with microcirculation changes in both groups. The patients who died had the highest PBR levels (P=0.04). Conclusions: Children with septic shock who receive milrinone infusions have microcirculation changes compared with those who do not receive them. The group that received milrinone was found to maintain functional capillary density and capillary recruitment capacity and have less endothelial glycocalyx degradation 24 hours after administration. These changes were present despite the inflammatory response and were more significant in those under two years of age.

SphK1 deficiency ameliorates the development of atherosclerosis by inhibiting the S1P/S1PR3/Rhoa/ROCK pathway.

BACKGROUND AND AIMS: S1P is an important factor regulating the function of the vascular endothelial barrier. SphK1 is an important limiting enzyme for the synthesis of S1P. However, the role of the SphK1/S1P-mediated vascular endothelial barrier function in atherosclerosis has not been fully revealed. This study explored the roles and mechanisms of SphK1 on atherosclerosis in vivo and in vitro. METHODS: In vivo, ApoE-/- and SphK1-/-ApoE-/- mice were fed a high-fat diet to induce atherosclerosis. In vitro, ox-LDL induced HUVECs to establish a cell model. Aortic histological changes were measured by H&E staining, Oil Red O staining, EVG staining, Sirius scarlet staining, immunofluorescence, and Evans Blue Assay. Western blotting was performed to explore the specific mechanism. RESULTS: We validated that deficiency of SphK1 resulted in a marked amelioration of atherosclerosis, as indicated by the decreased lipid accumulation, inflammatory factors, oxidative stress, aortic plaque area, inflammatory factor infiltration, VCAM-1 expression, and vascular endothelial permeability. Moreover, deficiency of SphK1 downregulated the expression of aortic S1PR3, Rhoa, ROCK, and F-actin. The results of administration with the SphK1 inhibitor PF-543 and the S1PR3 inhibitor VPC23019 in vitro further confirmed the conclusion that deficiency of SphK1 reduced S1P level and S1PR3 protein expression, inhibited Rhoa/ROCK signaling pathway, regulated protein expression of F-actin, improved vascular endothelial dysfunction and permeability, and exerted anti-atherosclerotic effects. CONCLUSIONS: This study revealed that deficiency of SphK1 relieved vascular endothelial barrier function in atherosclerosis mice via SphK1/S1P/S1PR signaling pathway.

Importance of IgE-Induced Unique Plasma Leakage in the Skin for Urticaria-Like Symptoms in an Anaphylaxis-Dependent Spotted Distribution of Immune Complex in Skin (ASDIS) Mouse Model.

INTRODUCTION: Allergic diseases, such as anaphylaxis and urticaria, pose significant health concerns. The quest for improved prognostic outcomes in these diseases necessitates the exploration of novel therapeutic avenues. To address this need, we have developed a novel mouse model of anaphylaxis, denoted as anaphylaxis-dependent spotted distribution of immune complex in skin (ASDIS). ASDIS manifests as distinct dotted symptoms in the skin, detectable through in vivo imaging, resembling urticarial symptoms. In this study, we investigated the potential underlying mechanisms giving rise to these dotted symptoms, exploring the role of vascular permeability and characterizing the ASDIS model as a new urticaria model. METHODS: We employed haired and hairless HR mice (BALB/c background) and hairless HR-1 mice (a commercially available hairless strain with an unidentified genetic background). ASDIS was induced by the simultaneous intravenous injection of anti-ovalbumin IgE and fluorescein isothiocyanate (FITC)-ovalbumin, along with Evans blue - a recognized vascular permeability indicator. Anaphylaxis and scratching behavior were monitored through rectal temperature decrease and optical observation, respectively. Histamine, platelet-activating factor, and compound 48/80 were injected with or without FITC-ovalbumin for comparative analysis. The effects of an α1 adrenergic receptor agonist applied to the skin were also examined. RESULTS: In hairless mice, the simultaneous injection of histamine, compound 48/80, or IgE with FITC-ovalbumin induced comparable rectal temperature decreases and vascular permeability. However, only the combination of FITC-ovalbumin and IgE triggered ASDIS, specifically the dotted urticaria-like symptom. Evans blue visualization and optical observation of dotted swelling confirmed that the vascular permeability mediated the phenomenon. Hairless mice exhibited a more pronounced temperature decrease than their haired counterparts when exposed to histamine, platelet-activating factor, compound 48/80, and IgE with FITC-ovalbumin. The application of an α1 adrenergic receptor agonist to the skin attenuated the topical urticaria-like symptom. CONCLUSION: Our experiments revealed four findings. The first is that ASDIS mirrors urticaria-like symptoms resulting from increased vascular permeability, akin to human urticaria. The second finding is that the development of dotted symptoms involves an IgE-induced, yet unidentified, mechanism not triggered by histamine or compound 48/80 alone. The third finding highlights the heightened susceptibility of hairless mice to ASDIS induction. The fourth finding demonstrates that the inhibition of ASDIS by the topical application of an α1 adrenergic receptor agonist hints at a potential anti-urticarial application for this vasoconstrictor. Further elucidation of these unidentified IgE-dependent mechanisms and the specific generation of dotted symptoms by IgE-immune complexes could provide novel insights into allergic response processes and therapeutic interventions for these conditions.

Contact System Activation and Bradykinin Generation in Angioedema: Laboratory Assessment and Biomarker Utilization.

The role of contact system activation has been clearly established in the pathogenesis of hereditary angioedema due to C1 inhibitor deficiency (HAE-C1INH). C1 inhibitor (C1INH)-protease complexes, levels of functional C1INH, plasma kallikrein activation, and cleavage of high-molecular-weight kininogen have each been associated with disease activity. More recently, HAE with normal levels of C1INH (HAE-nl-C1INH) has been recognized. Six genetic mutations have been identified which are linked to HAE-nl-C1INH phenotypes. The majority of individuals with HAE-nl-C1INH fall into the unknown category. There is substantial evidence that bradykinin generation underlies the recurrent attacks of swelling in some of these cohorts.

Augmentation of the EPR effect by mild hyperthermia to improve nanoparticle delivery to the tumor.

The clinical translation of the nanoparticle (NP)-based anticancer therapies is still unsatisfactory due to the heterogeneity of the enhanced permeability and retention (EPR) effect. Despite the promising preclinical outcome of the pharmacological EPR enhancers, their systemic toxicity can limit their clinical application. Hyperthermia (HT) presents an efficient tool to augment the EPR by improving tumor blood flow (TBF) and vascular permeability, lowering interstitial fluid pressure (IFP), and disrupting the structure of the extracellular matrix (ECM). Furthermore, the HT-triggered intravascular release approach can overcome the EPR effect. In contrast to pharmacological approaches, HT is safe and can be focused to cancer tissues. Moreover, HT conveys direct anti-cancer effects, which improve the efficacy of the anti-cancer agents encapsulated in NPs. However, the clinical application of HT is challenging due to the heterogeneous distribution of temperature within the tumor, the length of the treatment and the complexity of monitoring.

Screening and construction of nanobodies against human CD93 using phage libraries and study of their antiangiogenic effects.

Background: Cluster of Differentiation 93 (CD93) plays an important role in angiogenesis and is considered an important target for inhibiting tumor angiogenesis, but there are currently no therapeutic antibodies against CD93 in the clinic. Thus, we describe the screening of novel nanobodies (Nbs) targeting human CD93 from a phage library of shark-derived Nbs. Methods: Screening and enrichment of phage libraries by enzyme-linked immunosorbent assay (ELISA). Anti-CD93 Nbs were purified by expression in E. coli. The binding affinity of anti-CD93 Nbs NC81/NC89 for CD93 was examined by flow cytometry (FC) and ELISA. The thermal stability of NC81/NC89 was examined by ELISA and CD spectroscopy. Afterward, the anti-angiogenic ability of NC81/NC89 was examined by MTT, wound healing assay, and tube formation assay. The expression level of VE-cadherin (VE-Ca) and CD93 was detected by Western Blot (WB). The binding sites and binding forms of NC81/NC89 to CD93 were analyzed by molecular docking. Results: The anti-CD93 Nbs were screened in a phage library, expressed in E. coli, and purified to >95% purity. The results of FC and ELISA showed that NC81/NC89 have binding ability to human umbilical vein endothelial cells (HUVECs). The results of ELISA and CD spectroscopy showed that NC81/NC89 retained the ability to bind CD93 at 80°C and that the secondary structure remained stable. In vitro, the results showed that NC81 and NC89 significantly inhibited the proliferation and migration of human umbilical vein endothelial cells (HUVECs) as well as tube formation on Matrigel. Western Blot showed that NC81 and NC89 also inhibited the expression of VE-Ca thereby increasing vascular permeability. It was found during molecular docking that the CDR regions of NC81 and NC89 could be attached to CD93 by strong hydrogen bonds and salt bridges, and the binding sites were different. Conclusion: We have successfully isolated NC81 and NC89, which bind CD93, and both Nbs significantly inhibit angiogenesis and increase vascular permeability. These results suggest that NC81 and NC89 have potential clinical applications in angiogenesis-related therapies.

Segmental Shielding: A Rare Case of Acute Respiratory Distress Syndrome with Middle Lobe Sparing.

Acute respiratory distress syndrome (ARDS) presents a formidable challenge in critical care, often resulting in high mortality rates, particularly in severe cases or those compounded by preexisting conditions. Despite substantial advancements in critical care, the heterogeneous nature of ARDS necessitates nuanced clinical approaches. ARDS is generally diagnosed through clinical evaluation, radiographic imaging, and laboratory tests, as well as acute onset, bilateral lung infiltrates on imaging, and a partial pressure of oxygen in arterial blood (PaO2)/fraction of inspiratory oxygen concentration (FiO2) ratio of less than 300 mmHg. Management involves measurements to improve oxygenation and provide mechanical ventilation to assist breathing. The typical manifestation of ARDS is diffuse lung involvement, which affects multiple lobes symmetrically. Here, we report an unusual case of ARDS in a 53-year-old female who was brought into the hospital in an unresponsive state, exhibiting hypoxic and hypotension requiring intubation. Subsequent imaging revealed a distinctive pattern: the preservation of the right middle lobe, diverging from the conventional diffuse pulmonary affliction. This case underscores the need for clinical vigilance and adaptability, as such atypical presentations can confound diagnosis and management, posing unique clinical challenges. This case highlights the importance of recognizing ARDS' diverse presentations. Moreover, understanding the mechanisms behind the lobar sparing could provide greater insight into the disease heterogeneity and guide tailored therapeutic approaches. The imperative for further research into these uncommon presentations is clear, as it may be vital to improving outcomes for a broader spectrum of ARDS patients.

Neutrophils - an understudied bystander in dengue?

Dengue is a mosquito-borne viral disease which causes significant morbidity and mortality each year. Previous research has proposed several mechanisms of pathogenicity that mainly involve the dengue virus and host humoral immunity. However, innate immune cells, such as neutrophils, may also play an important role in dengue, albeit a much less defined role. In this review, we discuss the emerging roles of neutrophils in dengue and their involvement in pathologies associated with severe dengue. We also describe the potential use of several neutrophil proteins as biomarkers for severe dengue. These studies suggest that neutrophils are important players in dengue, and a better understanding of neutrophil-dengue biology is urgently needed.

Impaired high-density lipoprotein function and endothelial barrier stability in severe anaphylaxis.

BACKGROUND: Growing evidence demonstrates the importance of high- and low-density lipoprotein cholesterol in certain immune and allergy-mediated diseases. OBJECTIVE: This study aimed to evaluate levels of high- and low-density lipoprotein cholesterol and apolipoproteins A1 and B in sera from a cohort of patients presenting with hypersensitivity reactions. We further assessed the function of high-density lipoprotein particles as well as their involvement in the molecular mechanisms of anaphylaxis. METHODS: Lipid profile determination was performed in paired (acute and baseline) serum samples from 153 patients. Thirty-eight experienced a non-anaphylactic reaction and 115 had an anaphylactic reaction (88 moderate and 27 severe). Lecithin cholesterol acyl transferase activity was assessed in patient sera, and we also evaluated macrophage cholesterol efflux in response to the serum samples. Last, the effect of anaphylactic-derived high-density lipoprotein (HDL) particles on the endothelial barrier was studied. Detailed methods are provided in the Methods section in this article's Online Repository available at www.jacionline.org. RESULTS: Serum samples from severe anaphylactic reactions show statistically significant low levels of HDL cholesterol, low-density lipoprotein cholesterol, and apolipoproteins A1 and B, which points to their possible role as biomarkers. Specifically, HDL particles play a protective role in cardiovascular diseases. Using functional human serum cell assays, we observed impaired capacity of apolipoprotein B-depleted serum to induce macrophage cholesterol efflux in severe anaphylactic reactions. In addition, purified HDL particles from human anaphylactic sera failed to stabilize and maintain the endothelial barrier. CONCLUSION: These results encourage further research on HDL functions in severe anaphylaxis, which may lead to new diagnostic and therapeutic strategies.

Hereditary angioedema classification: Expanding knowledge by genotyping and endotyping.

Hereditary angioedema (HAE) encompasses a group of diseases characterized by recurrent, genetically mediated angioedema associated with increased vascular permeability primarily due to bradykinin. The disease poses diagnostic challenges, leading to underdiagnosis and delayed therapy. Severe manifestations include laryngeal and intestinal angioedema, contributing to significant morbidity and mortality. If left undiagnosed, the estimated mortality rate of the disease ranges from 25% to 40% due to asphyxiation caused by laryngeal angioedema. There is a pressing need to enhance awareness of hereditary angioedema and its warning signs. The acronym "H4AE" may facilitate the memorization of these signs. This study comprehensively reviews clinical, laboratory, and physiopathological features of documented HAE subtypes. The study advocates for an improved HAE classification based on endotypes, building on the knowledge of angioedema pathophysiology. The proposed endotype classification of HAE offers a clear and applicable framework, encouraging advancements in disease understanding and classification.

Conditional deficiency of Rho-associated kinases disrupts endothelial cell junctions and impairs respiratory function in adult mice.

The Ras homology (Rho) family of GTPases serves various functions, including promotion of cell migration, adhesion, and transcription, through activation of effector molecule targets. One such pair of effectors, the Rho-associated coiled-coil kinases (ROCK1 and ROCK2), induce reorganization of actin cytoskeleton and focal adhesion through substrate phosphorylation. Studies on ROCK knockout mice have confirmed that ROCK proteins are essential for embryonic development, but their physiological functions in adult mice remain unknown. In this study, we aimed to examine the roles of ROCK1 and ROCK2 proteins in normal adult mice. Tamoxifen (TAM)-inducible ROCK1 and ROCK2 single and double knockout mice (ROCK1flox/flox and/or ROCK2flox/flox;Ubc-CreERT2) were generated and administered a 5-day course of TAM. No deaths occurred in either of the single knockout strains, whereas all of the ROCK1/ROCK2 double conditional knockout mice (DcKO) had died by Day 11 following the TAM course. DcKO mice exhibited increased lung tissue vascular permeability, thickening of alveolar walls, and a decrease in percutaneous oxygen saturation compared with noninducible ROCK1/ROCK2 double-floxed control mice. On Day 3 post-TAM, there was a decrease in phalloidin staining in the lungs in DcKO mice. On Day 5 post-TAM, immunohistochemical analysis also revealed reduced staining for vascular endothelial (VE)-cadherin, ß-catenin, and p120-catenin at cell-cell contact sites in vascular endothelial cells in DcKO mice. Additionally, VE-cadherin/ß-catenin complexes were decreased in DcKO mice, indicating that ROCK proteins play a crucial role in maintaining lung function by regulating cell-cell adhesion.

The Vitamin K-Dependent Anticoagulant Factor, Protein S, Regulates Vascular Permeability.

Protein S (PROS1) is a vitamin K-dependent anticoagulant factor, which also acts as an agonist for the TYRO3, AXL, and MERTK (TAM) tyrosine kinase receptors. PROS1 is produced by the endothelium which also expresses TAM receptors, but little is known about its effects on vascular function and permeability. Transwell permeability assays as well as Western blotting and immunostaining analysis were used to monitor the possible effects of PROS1 on both endothelial cell permeability and on the phosphorylation state of specific signaling proteins. We show that human PROS1, at its circulating concentrations, substantially increases both the basal and VEGFA-induced permeability of endothelial cell (EC) monolayers. PROS1 induces p38 MAPK (Mitogen Activated Protein Kinase), Rho/ROCK (Rho-associated protein kinase) pathway activation, and actin filament remodeling, as well as substantial changes in Vascular Endothelial Cadherin (VEC) distribution and its phosphorylation on Ser665 and Tyr685. It also mediates c-Src and PAK-1 (p21-activated kinase 1) phosphorylation on Tyr416 and Ser144, respectively. Exposure of EC to human PROS1 induces VEC internalization as well as its cleavage into a released fragment of 100 kDa and an intracellular fragment of 35 kDa. Using anti-TAM neutralizing antibodies, we demonstrate that PROS1-induced VEC and c-Src phosphorylation are mediated by both the MERTK and TYRO3 receptors but do not involve the AXL receptor. MERTK and TYRO3 receptors are also responsible for mediating PROS1-induced MLC (Myosin Light Chain) phosphorylation on a site targeted by the Rho/ROCK pathway. Our report provides evidence for the activation of the c-Src/VEC and Rho/ROCK/MLC pathways by PROS1 for the first time and points to a new role for PROS1 as an endogenous vascular permeabilizing factor.

Obesity-induced blood-brain barrier dysfunction: phenotypes and mechanisms.

Obesity, a burgeoning global health issue, is increasingly recognized for its detrimental effects on the central nervous system, particularly concerning the integrity of the blood-brain barrier (BBB). This manuscript delves into the intricate relationship between obesity and BBB dysfunction, elucidating the underlying phenotypes and molecular mechanisms. We commence with an overview of the BBB's critical role in maintaining cerebral homeostasis and the pathological alterations induced by obesity. By employing a comprehensive literature review, we examine the structural and functional modifications of the BBB in the context of obesity, including increased permeability, altered transport mechanisms, and inflammatory responses. The manuscript highlights how obesity-induced systemic inflammation and metabolic dysregulation contribute to BBB disruption, thereby predisposing individuals to various neurological disorders. We further explore the potential pathways, such as oxidative stress and endothelial cell dysfunction, that mediate these changes. Our discussion culminates in the summary of current findings and the identification of knowledge gaps, paving the way for future research directions. This review underscores the significance of understanding BBB dysfunction in obesity, not only for its implications in neurodegenerative diseases but also for developing targeted therapeutic strategies to mitigate these effects.

Potential Therapeutic Strategies and Drugs That Target Vascular Permeability in Severe Infectious Diseases.

Severe infection pathogenicity is induced by processes such as pathogen exposure, immune cell activation, inflammatory cytokine production, and vascular hyperpermeability. Highly effective drugs, such as antipathogenic agents, steroids, and antibodies that suppress cytokine function, have been developed to treat the first three processes. However, these drugs cannot completely suppress severe infectious diseases, such as coronavirus disease 2019 (COVID-19). Therefore, developing novel drugs that inhibit vascular hyperpermeability is crucial. This review summarizes the mechanisms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-induced vascular hyperpermeability and identifies inhibitors that increase endothelial cell (EC) junction-related proteins and determines their efficacy in COVID-19 and endotoxemia models. Analyzing the effects of SARS-CoV-2 on vascular permeability revealed that SARS-CoV-2 suppresses Claudin-5 (CLDN5) expression, which is responsible for adhesion between ECs, thereby increasing vascular permeability. Inhibiting CLDN5 function in mice induced vascular hyperpermeability and pulmonary edema. In contrast, Enhancing CLDN5 expression suppressed SARS-CoV-2-induced endothelial hyperpermeability, suggesting that SARS-CoV-2-induced vascular hyperpermeability contributes to pathological progression, which can be suppressed by upregulating EC junction proteins. Based on these results, we focused on Roundabout4 (Robo4), another EC-specific protein that stabilizes EC junctions. EC-specific Robo4 overexpression suppressed vascular hyperpermeability and mortality in lipopolysaccharide-treated mice. An ALK1 inhibitor (a molecule that increases Robo4 expression), suppressed vascular hyperpermeability and mortality in lipopolysaccharide- and SARS-CoV-2-treated mice. These results indicate that Robo4 expression-increasing drugs suppress vascular permeability and pathological phenotype in COVID-19 and endotoxemia models.

Inhibition of leukocyte migration after ischemic stroke by VE-cadherin mutation in a mouse model leads to reduced infarct volumes and improved motor skills.

AIMS: To distinguish between the genuine cellular impact of the ischemic cascade by leukocytes and unspecific effects of edema and humoral components, two knock-in mouse lines were utilized. Mouse lines Y731F and Y685F possess point mutations in VE-cadherin, which lead to a selective inhibition of transendothelial leukocyte migration or impaired vascular permeability. METHODS: Ischemic stroke was induced by a model of middle cerebral artery occlusion. Analysis contained structural outcomes (infarct volume and extent of brain edema), functional outcomes (survival analysis, rotarod test, and neuroscore), and the extent and spatial distribution of leukocyte migration (heatmaps and fluorescence-activated cell sorting (FACS) analysis). RESULTS: Inhibition of transendothelial leukocyte migration as in Y731F mice leads to smaller infarct volumes (52.33 ± 4719 vs. 70.43 ± 6483 mm3 , p = .0252) and improved motor skills (rotarod test: 85.52 ± 13.24 s vs. 43.06 ± 15.32 s, p = .0285). An impaired vascular permeability as in Y685F mice showed no effect on structural or functional outcomes. Both VE-cadherin mutations did not influence the total immune cell count or spatial distribution in ischemic brain parenchyma. CONCLUSION: Selective inhibition of transendothelial leukocyte migration by VE-cadherin mutation after ischemic stroke in a mouse model leads to smaller infarct volumes and improved motor skills.

Roles of Macrophages and Endothelial Cells and Their Crosstalk in Acute Lung Injury.

Acute lung injury (ALI) and its severe form, acute respiratory distress syndrome (ARDS), present life-threatening conditions characterized by inflammation and endothelial injury, leading to increased vascular permeability and lung edema. Key players in the pathogenesis and resolution of ALI are macrophages (Mφs) and endothelial cells (ECs). The crosstalk between these two cell types has emerged as a significant focus for potential therapeutic interventions in ALI. This review provides a brief overview of the roles of Mφs and ECs and their interplay in ALI/ARDS. Moreover, it highlights the significance of investigating perivascular macrophages (PVMs) and immunomodulatory endothelial cells (IMECs) as crucial participants in the Mφ-EC crosstalk. This sheds light on the pathogenesis of ALI and paves the way for innovative treatment approaches.

Dapagliflozin ameliorated retinal vascular permeability in diabetic retinopathy rats by suppressing inflammatory factors.

BACKGROUND: Diabetic retinopathy is a common microvascular complication of diabetes and one of the major causes of blindness in the working-age population. Emerging evidence has elucidated that inflammation drives the key mechanism of diabetes-mediated retinal disturbance. As a new therapeutic drug targeting diabetes, whether dapagliflozin could improve vascular permeability from the perspective of anti-inflammatory effect need to be further explored. METHODS: Type 2 diabetic retinopathy rat model was established and confirmed by fundus fluorescein angiography (FFA). ELISA detected level of plasma inflammatory factors and C-peptide. HE staining, immunohistochemistry and western blot detected histopathology changes of retina, expression of retinal inflammatory factors and tight junction proteins. RESULTS: Dapagliflozin exhibited hypoglycemic effect comparable to insulin, but did not affect body weight. By inhibiting expression of inflammatory factors (NLRP3, Caspase-1, IL-18, NF-κB) in diabetic retina and plasma, dapagliflozin reduced damage of retinal tight junction proteins and improved retinal vascular permeability. The anti-inflammatory effect of dapagliflozin was superior to insulin. CONCLUSIONS: Dapagliflozin improved retinal vascular permeability by reducing diabetic retinal and plasma inflammatory factors. The anti-inflammatory mechanism of dapagliflozin is independent of hypoglycemic effect and superior to insulin.

Akt3 activation by R-Ras in an endothelial cell enforces quiescence and barrier stability of neighboring endothelial cells via Jagged1.

Communication between adjacent endothelial cells is important for the homeostasis of blood vessels. We show that quiescent endothelial cells use Jagged1 to instruct neighboring endothelial cells to assume a quiescent phenotype and secure the endothelial barrier. This phenotype enforcement by neighboring cells is operated by R-Ras through activation of Akt3, which results in upregulation of a Notch ligand Jagged1 and consequential upregulation of Notch target genes, such as UNC5B, and VE-cadherin accumulation in the neighboring cells. These signaling events lead to the stable interaction between neighboring endothelial cells to continue to fortify juxtacrine signaling via Jagged1-Notch. This mode of intercellular signaling provides a positive feedback regulation of endothelial cell-cell interactions and cellular quiescence required for the stabilization of the endothelium.