Author Correction: Heparanase-2 protects from LPS-mediated endothelial injury by inhibiting TLR4 signalling

The Acknowledgements section in the original version of this Article was incomplete. “We are grateful to Prof. Israel Vlodavsky (Technion, Haifa, Israel) for giving us 1c7 antibody to HPSE2. We are grateful to Prof. Fridrich Luft for critical editing the manuscript. Grants from German Federal Ministry of Education and Research (BMBF) Nr. 031A577A and 031A577B funded this research. This work was also supported by a grant for the German Research Council to H.H. Ha1388/17-1.” now reads: “We are grateful to Prof. Israel Vlodavsky (Technion, Haifa, Israel) for giving us 1c7 antibody to HPSE2. We are grateful to Prof. Fridrich Luft for critical editing the manuscript. Grants from German Federal Ministry of Education and Research (BMBF) Nr. 031A577A and 031A577B funded this research. This work was also supported by a grant for the German Research Council to H.H. Ha1388/17-1. This work was also supported by a grant from the Else Kröner-Fresenius-Stiftung (EKFS): Grant 2017_A96.” The original Article has been corrected

Heparanase-2 protects from LPS-mediated endothelial injury by inhibiting TLR4 signalling

The endothelial glycocalyx and its regulated shedding are important to vascular health. Endo-β-D-glucuronidase heparanase-1 (HPSE1) is the only enzyme that can shed heparan sulfate. However, the mechanisms are not well understood. We show that HPSE1 activity aggravated Toll-like receptor 4 (TLR4)-mediated response of endothelial cells to LPS. On the contrary, overexpression of its endogenous inhibitor, heparanase-2 (HPSE2) was protective. The microfluidic chip flow model confirmed that HPSE2 prevented heparan sulfate shedding by HPSE1. Furthermore, heparan sulfate did not interfere with cluster of differentiation-14 (CD14)-dependent LPS binding, but instead reduced the presentation of the LPS to TLR4. HPSE2 reduced LPS-mediated TLR4 activation, subsequent cell signalling, and cytokine expression. HPSE2-overexpressing endothelial cells remained protected against LPS-mediated loss of cell-cell contacts. In vivo, expression of HPSE2 in plasma and kidney medullary capillaries was decreased in mouse sepsis model. We next applied purified HPSE2 in mice and observed decreases in TNFα and IL-6 plasma concentrations after intravenous LPS injections. Our data demonstrate the important role of heparan sulfate and the glycocalyx in endothelial cell activation and suggest a protective role of HPSE2 in microvascular inflammation. HPSE2 offers new options for protection against HPSE1-mediated endothelial damage and preventing microvascular disease. © 2019, The Author(s)

Podocytic PKC-Alpha Is Regulated in Murine and Human Diabetes and Mediates Nephrin Endocytosis

Background: Microalbuminuria is an early lesion during the development of diabetic nephropathy. The loss of high molecular weight proteins in the urine is usually associated with decreased expression of slit diaphragm proteins. Nephrin, is the major component of the glomerular slit diaphragm and loss of nephrin has been well described in rodent models of experimental diabetes as well as in human diabetic nephropathy. Methodology/Principal Findings: In this manuscript we analyzed the role of PKC-alpha (PKCa) on endocytosis of nephrin in podocytes. We found that treatment of diabetic mice with a PKCa-inhibitor (GÖ6976) leads to preserved nephrin expression and reduced proteinuria. In vitro, we found that high glucose stimulation would induce PKCa protein expression in murine and human podocytes. We can demonstrate that PKCa mediates nephrin endocytosis in podocytes and that overexpression of PKCa leads to an augmented endocytosis response. After PKC-activation, we demonstrate an inducible association of PKCa, PICK1 and nephrin in podocytes. Moreover, we can demonstrate a strong induction of PKCa in podocytes of patients with diabetic nephropathy. Conclusions/Significance: We therefore conclude that activation of PKCa is a pathomechanistic key event during the development of diabetic nephropathy. PKCa is involved in reduction of nephrin surface expression and therefore PKC

The synthetic Tie2 agonist peptide vasculotide protects against vascular leakage and reduces mortality in murine abdominal sepsis

Introduction: Angiopoietin-1 (Angpt1), the natural agonist ligand for the endothelial Tie2 receptor, is a non-redundant endothelial survival and vascular stabilization factor that reduces endothelial permeability and inhibits leukocyte-endothelium interactions. Here we evaluate the efficacy of a novel polyethylene glycol (PEG)-clustered Tie2 agonist peptide, vasculotide (VT), to protect against vascular leakage and mortality in a murine model of polymicrobial abdominal sepsis. Methods: Polymicrobial abdominal sepsis in C57BL6 mice was induced by cecal-ligation-and-puncture (CLP). Mice were treated with different dosages of VT or equal volume of phosphate-buffered saline (PBS). Sham-operated animals served as time-matched controls. Results: Systemic administration of VT induced long-lasting Tie2 activation in vivo. VT protected against sepsis-induced endothelial barrier dysfunction, as evidenced by attenuation of vascular leakage and leukocyte transmigration into the peritoneal cavity. Histological analysis revealed that VT treatment ameliorated leukocyte infiltration in kidneys of septic mice, probably due to reduced endothelial adhesion molecule expression. VT-driven effects were associated with significantly improved organ function and reduced circulating cytokine levels. The endothelial-specific action of VT was supported by additional in vitro studies showing no effect of VT on either cytokine release from isolated peritoneal macrophages, or migratory capacity of isolated neutrophils. Finally, administration of VT pre-CLP (hazard ratio 0.39 [95% confidence interval 0.19-0.81] P < 0.001) and post-CLP reduced mortality in septic mice (HR 0.22 [95% CI 0.06-0.83] P < 0.05). Conclusions: We provide proof of principle in support of the efficacious use of PEGylated VT, a drug-like Tie2 receptor agonist, to counteract microvascular endothelial barrier dysfunction and reduce mortality in a clinically relevant murine sepsis model. Further studies are needed to pave the road for clinical application of this therapeutic concept

Alpha1-antitrypsin improves survival in murine abdominal sepsis model by decreasing inflammation and sequestration of free heme

BackgroundExcessive inflammation, hemolysis, and accumulation of labile heme play an essential role in the pathophysiology of multi-organ dysfunction syndrome (MODS) in sepsis. Alpha1-antitrypsin (AAT), an acute phase protein with heme binding capacity, is one of the essential modulators of host responses to inflammation. In this study, we evaluate the putative protective effect of AAT against MODS and mortality in a mouse model of polymicrobial abdominal sepsis.MethodsPolymicrobial abdominal sepsis was induced in C57BL/6N mice by cecal ligation and puncture (CLP). Immediately after CLP surgery, mice were treated intraperitoneally with three different forms of human AAT—plasma-derived native (nAAT), oxidized nAAT (oxAAT), or recombinant AAT (recAAT)—or were injected with vehicle. Sham-operated mice served as controls. Mouse survival, bacterial load, kidney and liver function, immune cell profiles, cytokines/chemokines, and free (labile) heme levels were assessed. In parallel, in vitro experiments were carried out with resident peritoneal macrophages (MPMΦ) and mouse peritoneal mesothelial cells (MPMC).ResultsAll AAT preparations used reduced mortality in septic mice. Treatment with AAT significantly reduced plasma lactate dehydrogenase and s-creatinine levels, vascular leakage, and systemic inflammation. Specifically, AAT reduced intraperitoneal accumulation of free heme, production of cytokines/chemokines, and neutrophil infiltration into the peritoneal cavity compared to septic mice not treated with AAT. In vitro experiments performed using MPMC and primary MPMΦ confirmed that AAT not only significantly decreases lipopolysaccharide (LPS)-induced pro-inflammatory cell activation but also prevents the enhancement of cellular responses to LPS by free heme. In addition, AAT inhibits cell death caused by free heme in vitro.ConclusionData from the septic CLP mouse model suggest that intraperitoneal AAT treatment alone is sufficient to improve sepsis-associated organ dysfunctions, preserve endothelial barrier function, and reduce mortality, likely by preventing hyper-inflammatory responses and by neutralizing free heme

Autoimmunity in CD73/Ecto-5′-Nucleotidase Deficient Mice Induces Renal Injury

Extracellular adenosine formed by 5′-ectonucleotidase (CD73) is involved in tubulo-glomerular feedback in the kidney but is also known to be an important immune modulator. Since CD73−/−mutant mice exhibit a vascular proinflammatory phenotype, we asked whether long term lack of CD73 causes inflammation related kidney pathologies. CD73−/−mice (13 weeks old) showed significantly increased low molecule proteinuria compared to C57BL6 wild type controls (4.8≥0.52 vs. 2.9±0.54 mg/24 h, p<0.03). Total proteinuria increased to 5.97±0.78 vs. 2.55±0.35 mg/24 h at 30 weeks (p<0.01) whereas creatinine clearance decreased (0.161±0.02 vs. 0.224±0.02 ml/min). We observed autoimmune inflammation in CD73−/−mice with glomerulitis and peritubular capillaritis, showing glomerular deposition of IgG and C3 and enhanced presence of CD11b, CD8, CD25 as well as GR-1-positive cells in the interstitium. Vascular inflammation was associated with enhanced serum levels of the cytokines IL-18 and TNF-α as well as VEGF and the chemokine MIP-2 (CXCL-2) in CD73−/−mice, whereas chemokines and cytokines in the kidney tissue were unaltered or reduced. In CD73−/−mice glomeruli, we found a reduced number of podocytes and endothelial fenestrations, increased capillaries per glomeruli, endotheliosis and enhanced tubular fibrosis. Our results show that adult CD73−/−mice exhibit spontaneous proteinuria and renal functional deterioration even without exogenous stress factors. We have identified an autoimmune inflammatory phenotype comprising the glomerular endothelium, leading to glomeruli inflammation and injury and to a cellular infiltrate of the renal interstitium. Thus, long term lack of CD73 reduced renal function and is associated with autoimmune inflammation

KEY COMPONENTS OF MICROSOMAL OXIDATION SYSTEM IN RETINA AND CEREBRAL CORTEX IN NORM AND AT EXTREME STATES OF RETINA

The work covers the retina and myocardium tissues of bulls, line Vistar rat and Campball line rat with hereditary degeneration of the retina. The aim of the work is to identify the unknown component in the microsomal oxidation system of retina and to study the influence of the extreme retina state on this system. The protein containing the unheme ferrous - key component in the microsomal oxidation system of this tissue has been extracted firstly from microsomes of the retina. The activation of this system at extreme retina states has been shown firstly. It has been determined that the development of the hereditary retina degeneration among rats is accompanied with the activation of microsomal oxidation system also in the cerebral cortex and strong breach of ferric homeostasis on the early stages of the disease development. The supposition about possible role of the ferrum ions in the pathogenesis of the hereditary retina degeneration has been brought upAvailable from VNTIC / VNTIC - Scientific & Technical Information Centre of RussiaSIGLERURussian Federatio

Correction: Calcium dobesilate reduces VEGF signaling by interfering with heparan sulfate binding site and protects from vascular complications in diabetic mice.

[This corrects the article DOI: 10.1371/journal.pone.0218494.]

Calcium dobesilate reduces VEGF signaling by interfering with heparan sulfate binding site and protects from vascular complications in diabetic mice.

Inhibiting vascular endothelial growth factor (VEGF) is a therapeutic option in diabetic microangiopathy. However, VEGF is needed at physiological concentrations to maintain glomerular integrity; complete VEGF blockade has deleterious effects on glomerular structure and function. Anti-VEGF therapy in diabetes raises the challenge of reducing VEGF-induced pathology without accelerating endothelial cell injury. Heparan sulfate (HS) act as a co-receptor for VEGF. Calcium dobesilate (CaD) is a small molecule with vasoprotective properties that has been used for the treatment of diabetic microangiopathy. Preliminary evidence suggests that CaD interferes with HS binding sites of fibroblast growth factor. We therefore tested the hypotheses that (1) CaD inhibits VEGF signaling in endothelial cells, (2) that this effect is mediated via interference between CaD and HS, and (3) that CaD ameliorates diabetic nephropathy in a streptozotocin-induced diabetic mouse model by VEGF inhibition. We found that CaD significantly inhibited VEGF165-induced endothelial cell migration, proliferation, and permeability. CaD significantly inhibited VEGF165-induced phosphorylation of VEGFR-2 and suppressed the activity of VEGFR-2 mediated signaling cascades. The effects of CaD in vitro were abrogated by heparin, suggesting the involvement of heparin-like domain in the interaction with CaD. In addition, VEGF121, an isoform which does not bind to heparin, was not inhibited by CaD. Using the proximity ligation approach, we detected inhibition of interaction in situ between HS and VEGF and between VEGF and VEGFR-2. Moreover, CaD reduced VEGF signaling in mice diabetic kidneys and ameliorated diabetic nephropathy and neuropathy, suggesting CaD as a VEGF inhibitor without the negative effects of complete VEGF blockade and therefore could be useful as a strategy in treating diabetic nephropathy