Mechanism Of Endothelial Cell Injury Research Articles

Lichen sclerosus: A C5B-9 mediated chronic microvascular injury syndrome potentially reflective of common adult comorbidities

Lichen sclerosus (LS) is a cutaneous disease of unknown etiology that often involves the vulva or foreskin but also can affect extragenital sites. Regardless of the anatomic site, the histomorphology and presumably pathogenesis are similar. Perhaps a clue to the pathophysiology of LS lies in its frequent association with morphea, specifically, when occurring in an extragenital context. In our experience a striking feature evident in established lichen sclerosis (LS) is one of superficial vascular drop out whereby residual vessels exhibited endothelial cell necrosis and microvascular basement membrane zone thickening, the latter reflective of antecedent episodes of microvascular injury. We sought to understand the pathophysiology that underlies the distinct vascular changes and in doing so, shed light on the pathogenesis of LS. We examined 44 cases of LS over a period of 2019 to 2021. We were able to obtain past medical histories in 34 of the 44 cases. Regarding pathological assessment, the predominant focus was on microvascular changes. We assessed the role of C5b-9 mediated vascular injury in the pathogenesis of the vasculopathy and enhanced type I interferon signaling in vessels given the morphologic semblance to the select interferonopathy syndromes, namely fibrosing dermatomyositis and Kohlmeier Degos disease. We examined the expression of CMV DNA and protein based on prior observations in an earlier study that isolated early protein expression in the microvasculature in the setting of LS and scleroderma. From a clinical perspective, the most striking association was an older age at the time of diagnosis (mean age of 62 years and median age of 61.5 years) and the presence of vascular comorbidities of diabetes, hypertension, and hyperlipidemia in almost 80% of cases. All cases showed significant microvascular changes in the superficial corium with the most frequent findings being those of significant basement membrane zone reduplication and vascular drop out. A number of cases showed prominent microvascular deposits of C5b-9 in the zone of hyalinizing fibrosis or subjacent to the discernible table of fibroplasia in the absence of enhanced type I interferon signaling. In no case were there viral cytopathic changes associated with CMV affecting the endothelium. The studies that encode CMV DNA or protein did not show a significant role for CMV reactivation in endothelium in the majority of the studied cases. It is concluded that the pathophysiology of LS includes a microvascular injury syndrome within the papillary dermis. The mechanism of endothelial cell injury is complement mediated at least in part and could reflect an adaptive immune response targeting endothelium indicative of classic complement pathway activation when coexisting with morphea or occurring in younger individuals. A non-immune based endothelial dysfunction and complement mediated injury unrelated to antibody driven classic complement pathway activation are more likely pathogenetically in the setting of certain diseases like diabetes mellitus and hypertension. Vascular drop out can be explained by the diminished endothelial progenitor pool needed to repopulate the damaged microvessels in certain settings like hypertension and diabetes.

STIM1 Regulates Endothelial Calcium Overload and Cytokine Upregulation During Sepsis

BackgroundStromal interaction molecule 1 (STIM1)-mediated store-operated Ca2+ entry (SOCE) is now recognized as the main mechanism of the majority of nonexcitable cell calcium influx. Calcium overload is a primary mechanism of endothelial cell injury during systemic inflammatory response and sepsis. Whether STIM1-mediated SOCE plays a role in calcium overload in vascular endothelial cell injury remains unclear. Materials and methodsTo explore the role of STIM1-gated SOCE in vascular endothelial cell calcium overload and inflammation, we established a human septic serum or lipopolysaccharide (LPS)-induced human umbilical vein endothelial cell (HUVEC) experimental system and derived ribonucleic acid interference (RNAi)-mediated STIM1, ORAI1 (orai gene [HGNC: 25896 Entrez Gene: 84876] coding protein, ORAI Calcium Release-Activated Calcium Modulator 1), and transient receptor potential channel 1 (TRPC1) (core components of store-operated Ca2+[SOC]) downregulated HUVECs, as well as STIM1 overinduced HUVECs. ResultsOur results show that sepsis serum or LPS stimulation increased STIM1 in HUVECs and increased all cytokines except for VEGF and the inflammatory mediators tumor necrosis factor, intercellular cell adhesion molecule-1, and endothelin-1 in a time-dependent manner. RNAi-mediated knockdown of STIM1 significantly inhibited serum or LPS-induced inflammatory cytokine expression, and STIM1 overexpression in HUVECs promoted LPS-mediated induction of these cytokines. Meanwhile, similar to the blocking effect of the specific SOC inhibitors Gd3+ and La3+ on LPS-induced calcium influx, RNAi-mediated depletion of STIM1 or the SOC proteins TRPC1 and ORAI1 could significantly inhibit serum or LPS-induced extracellular calcium influx, as well as the expression of the inflammatory cytokines tumor necrosis factor, intercellular cell adhesion molecule-1, and endothelin-1. Simultaneous downregulation of the SOCE core units TRPC1 and ORAI1 inhibited LPS-induced calcium influx and cytokine expression, which could not be restored by inducing STIM1. Forced expression of nuclear factor-κB (NF-κB) in HUVECs significantly induced STIM1 expression, whereas RNAi-mediated depletion of NF-κB significantly inhibited STIM1 mRNA levels and significantly reduced the thapsigargin-mediated SOCE calcium influx, which was similar to results with the NF-κB inhibitor wogonin. ConclusionsSeptic serum stimulates the expression of STIM1, cytokines, and inflammatory mediators in HUVECs. STIM1-mediated SOCE is required for Ca2+ influx induced by LPS or septic serum and contributes cytokines and inflammatory mediators in septic serum–stimulated HUVECs. In addition, STIM1-mediated SOCE on Ca2+ influx by septic serum or LPS involves NF-κB signaling.

LncRNA ZEB1-AS1 knockdown alleviates oxidative low-density lipoprotein-induced endothelial cell injury via the miR-590-5p/ HDAC9 axis

Oxidative low-density lipoprotein (ox-LDL) is thought to induce vascular endothelial cell injury, which contributes to the aetiopathogenesis of atherosclerosis (AS). Several previous reports have identified that lncRNA ZEB1-AS1 participates in the regulatory mechanisms of endothelial cell injury, but the potential interaction mechanism between ZEB1-AS1 and miR-590-5p in ox-LDL-induced endothelial cell damage is not clear. ZEB1-AS1 and miR-590-5p expression were tested by quantitative real-time polymerase chain reaction (qRT-PCR) in ox-LDL-treated endothelial cells. The proliferation and apoptosis were determined by MTT and Annexin V/PI double-staining assay, respectively. The protein expression of HDAC9, tumor necrosis factor α (TNF-α), cleaved caspase-3, and cleaved PARP were measured by western blot analysis. Dual-luciferase reporter and RIP assays affirmed the functional targets of ZEB1-AS1. ZEB1-AS1 expression was upregulated in ox-LDL-treated HUVECs, and miR-590-5p was lessened in a dose- or time-depended manner, respectively. Knockdown of ZEB1-AS1 facilitated ox-LDL-treated endothelial cell proliferation and inhibited cell apoptosis. Moreover, miR-590-5p was directly targeted via ZEB1-AS1 in ox-LDL-treated HUVECs. ZEB1-AS1 silencing attenuated ox-LDL-induced cell injury via regulation of miR-590-5p expression. Furthermore, HDAC9 reversed the influence of miR-590-5p on propagation and apoptosis of ox-LDL-induced endothelial cells. Knockdown of ZEB1-AS1 alleviates ox-LDL-induced endothelial cell injury by regulating the miR-590-5p/HDAC9 axis.

Ginsenoside Re Attenuates High Glucose-Induced RF/6A Injury via Regulating PI3K/AKT Inhibited HIF-1α/VEGF Signaling Pathway.

Hyperglycaemia-induced retinal microvascular endothelial cell apoptosis is a critical and principle event in diabetic retinopathy (DR), which involves a series of complex processes such as mitochondrial dysfunction and oxidative stress. Ginsenoside Re (Re), a key ingredients of ginseng, is considered to have various pharmacologic functions, such as antioxidative, inhibition of inflammation and anti-apoptotic properties. However, the effects of Re in DR and the related mechanisms of endothelial cell injury induced by high glucose (HG) exposure remain unclear. The present study was designed to investigate and evaluate the ability of Re to ameliorate HG-induced retinal endothelial RF/6A cell injury and the potential mechanisms involved in the hypoxia-inducible factor-1-alpha (HIF-1α)/vascular endothelial growth factor (VEGF) signaling regulated by phosphoinositide 3-kinase (PI3K)/AKT pathway. Our results showed that preincubation with Re exerted cytoprotective effects by reversing the HG-induced decrease in RF/6A cell viability, downregulation of apoptosis rate and inhibition of oxidative-related enzymes, thereby reducing the excess intracellular reactive oxygen species (ROS) and HG-triggered RF/6A cell injury. In addition, Western blot analysis results showed ginsenoside Re significantly increased HIF-1α expression in the cytoplasm but decreased its expression in the nucleus, suggesting that it reduced the translocation of HIF-1α from the cytoplasm to the nucleus, and downregulated VEGF level. Moreover, this effect is involved in the activation of the PI3K/Akt pathway. LY294002, a PI3K inhibitor, was used to block the Akt pathway. Afterwards, the effects of Re on the regulation of apoptotic related proteins, VEGF and HIF-1α nuclear transcription was partially reversed. These findings suggested the exerting protective effects of ginsenoside Re were associated with regulating of PI3K/AKT and HIF-1α/VEGF signaling pathway, which indicates that ginsenoside Re may ameliorates HG-induced retinal angiogenesis and suggests the potential for the development of Re as a therapeutic for DR.

Endothelial Cell Dysfunction and Injury in Subarachnoid Hemorrhage.

In the brain, vascular endothelial cells conserve blood viscosity, control blood flow, and form the interface between central nervous system and circulating blood. Clinical outcome after aneurysmal subarachnoid hemorrhage is linked to early brain injury, cerebral vasospasm, and other causes of delayed cerebral ischemia. The cerebral vasculature remains a unique target for therapies since it becomes rapidly disrupted after subarachnoid hemorrhage, and damage to the blood vessels continues into the delayed injury phase. The current failure of therapies to improve clinical outcome warrants a re-evaluation of current therapeutic approaches. The mechanisms of endothelial cell injury and blood-brain barrier breakdown are critical to the pathway of cerebral injury, and an improved understanding of these mechanisms may lead to novel therapeutic targets. This review provides an update on the current understanding of endothelial cell injury following aneurysmal subarachnoid hemorrhage, including blood-brain barrier dysfunction.

Endothelial factors in the pathogenesis and treatment of chronic kidney disease Part II: Role in disease conditions: a joint consensus statement from the European Society of Hypertension Working Group on Endothelin and Endothelial Factors and the Japanese Society of Hypertension.

: After examining in Part I the general mechanisms of endothelial cell injury in the kidney, the Working Group on Endothelin and Endothelial Factors of the European Society of Hypertension and the Japanese Society of Hypertension will herein review current knowledge on the role of endothelial dysfunction in multiple disease conditions that affect the kidney, including diabetes mellitus, preeclampsia, solid organ transplantation, hyperhomocysteinemia and antiangiogenic therapy in cancer. The few available randomized controlled clinical trials specifically designed to evaluate strategies for correcting endothelial dysfunction in patients with hypertension and/or chronic kidney disease are also discussed alongside their cardiovascular and renal outcomes.

TRB3 mediates homocysteine‐induced inhibition of endothelial cell proliferation

Hyperhomocysteinemia (HHcy) has been shown to induce endothelial dysfunction, an early event in the progression of atherosclerosis. However, the underlying mechanism of endothelial cell injury in HHcy has not been clearly elucidated. In this study, we examined the effect of homocysteine on tribbles-related protein 3 (TRB3)-mediated cell-cycle arrest in human umbilical vein endothelial cells (HUVECs). Treatment of HUVECs with homocysteine (0-250 µmol/L) resulted in inhibition of cell proliferation assessed by [(3)H]-thymidine incorporation into DNA. Homocysteine induced cell-cycle arrest in the G1 phase by up-regulating the protein levels of p27(kip1). Under these conditions, homocysteine did not induce endoplasmic reticulum stress. However, homocysteine up-regulated the expression of TRB3, thus leading to the dephosphorylation of Akt (Thr308). Knock-down of endogenous TRB3 using siRNA significantly suppressed the inhibitory effect of homocysteine on the proliferation of HUVECs. Homocysteine-induced TRB3 expression was mediated by the cAMP/cAMP response element-binding protein (CREB) pathway. These results demonstrate that TRB3 is a critical molecule in the homocysteine-mediated cell-cycle arrest in endothelial cells.

Differential cytotoxic actions of Shiga toxin 1 and Shiga toxin 2 on microvascular and macrovascular endothelial cells

Shiga toxin (Stx)-mediated injury to vascular endothelial cells in the kidneys, brain and other organs underlies the pathogenesis of haemolytic uraemic syndrome (HUS) caused by enterohaemorrhagic Escherichia coli (EHEC). We present a direct and comprehensive comparison of cellular injury induced by the two major Stx types, Stx1 and Stx2, in human brain microvascular endothelial cells (HBMECs) and EA.hy 926 macrovascular endothelial cells. Scanning electron microscopy of microcarrier-based cell cultures, digital holographic microscopy of living single cells, and quantitative apoptosis/necrosis assays demonstrate that Stx1 causes both necrosis and apoptosis, whereas Stx2 induces almost exclusively apoptosis in both cell lines. Moreover, microvascular and macrovascular endothelial cells have different susceptibilities to the toxins: EA.hy 926 cells are slightly, but significantly (∼ 10 times) more susceptible to Stx1, whereas HBMECs are strikingly (≥ 1,000 times) more susceptible to Stx2. These findings have implications in the pathogenesis of HUS, and suggest the existence of yet to be delineated Stx type-specific mechanisms of endothelial cell injury beyond inhibition of protein biosynthesis.

A Fundamental Study on Increasing of Endothelial Permeability by Photochemical Reaction

As photochemical reaction (PR) is used in photodynamic therapy (PDT), high reactive oxygen speacies (ROS) concentration often induces cell and tissue injury. In this study, we focused on the mechanism of endothelial cell injury caused by PR. Human umbilical vein endothelial cells (HUVEC) were incubated with delta-aminolevulinic acid and were irradiated by LED light in 630 nm wavelength for 10 min. In vitro cell viability by MTT ssay did not change after PR, while in 3D-cultures of HUVEC in collagen that induces tube formation, PDT reduced the capillary-like structures and increase in endothelial permeability. ROS by PR leaded to shrinkage of endothelial cells, suppressed increase in subendothelial area after PR. Furthermore, F-actin was formed into stress fibers. These results suggest that oxidative stress by PDT strongly affects on endothelial permeability and morphological change of endothelial cells in which generation of stress fibers.

Endothelial cell fatty acid unsaturation mediates cold‐induced oxidative stress

Ultraprofound hypothermia (< 5 degrees C) induces changes to cell membranes such as liquid-to-gel lipid transitions and oxidative stress that have a negative effect on membrane function and cell survival. We hypothesized that fatty acid substitution of endothelial cell lipids and alterations in their unsaturation would modify cell survival at 0 degrees C, a temperature commonly used during storage and transportation of isolated cells or tissues and organs used in transplantation. Confluent bovine aortic endothelial cells were treated with 18-carbon fatty acids (C18:0, C18:1n-9, C18:2n-6, or C18:3n-3), C20:5n-3 or C22:6n-3 (DHA), and then stored at 0 degrees C without fatty acid supplements. Storage of control cells caused the release of lactate dehydrogenase (LDH) and a threefold increase in lipid peroxidation (LPO) when compared to control cells not exposed to cold. Pre-treating cells with C18:0 decreased the unsaturation of cell lipids and reduced LDH release at 0 degrees C by 50%, but all mono- or poly-unsaturated fatty acids increased injury in a concentration-dependent manner and as the extent of fatty acid unsaturation increased. DHA-treatment increased cell fatty acid unsaturation and caused maximal injury at 0 degrees C, which was prevented by lipophilic antioxidants BHT or vitamin E, the iron chelator deferoxamine, and to a lesser extent by vitamin C. Furthermore, the cold-induced increase in LPO was reduced by C18:0, vitamin E, or DFO but enhanced by DHA. In conclusion, the findings implicate iron catalyzed free radicals and LPO as a predominant mechanism of endothelial cell injury at 0 degrees C, which may be reduced by increasing lipid saturation or treating cells with antioxidants.

EDITORIAL

This issue marks the third of our review articles written by presidents and immediate past presidents of the four leading international respiratory societies, namely the American Thoracic Society, the American Academy of Chest Physicians, the Asian Pacific Society of Respirology and the European Respiratory Society. Professor Sharon Rounds is the immediate past president of the American Thoracic Society. Professor Rounds is chief of Pulmonary/Critical Care at the Providence VA Medical Centre and is also Associate Dean of Medicine for the Medical Faculty and Professor of Medicine and of Pathology and Laboratory Medicine at Brown Medical School. Her Research group, in the Pulmonary Vascular Research Laboratory, studies mechanisms of pulmonary vascular injury with a focus on mechanisms of endothelial cell injury. Her co-authors are Dr James R. Klinger and Dr Kevin B. Martin. Dr Klinger is Associate Professor of Medicine at Brown Medical School. His research interests include the mechanisms of pulmonary vascular response to hypoxia and clinical studies of pulmonary arterial hypertension. Dr Martin is a pulmonary/critical care postdoctoral fellow at Brown Medical School. Professor Rounds and her co-authors have an extensive publication record and together they have provided us with new insights and new hopes into the mechanisms and management of ‘pulmonary arterial hypertension’. As is the tradition in Respirology, this issue features a special Editorial from one of our newly appointed Associate Editors Dr Y.C. Gary Lee. Dr Lee joined our team in April 2005 and was also a co-editor of the very successful pleural disease review series published in Respirology last year. He is a respiratory physician at University College London working in Geoff Laurent's laboratory as well as carrying out clinical studies in Oxford. He holds a Wellcome Trust Senior Research Fellowship. Dr Lee has made a major contribution to our understanding of pleural adhesions and pleurodesis. He is the co-editor of one of the definitive textbooks on pleural disease. His editorial is entitled ‘Pleural disease: a forgotten frontier in respiratory research’. We are delighted with the excellent review of all clinical papers published in 2005 by Dr Gary Lee and Professor Richard Beasley. Submission to Respirology can be made online as of 1 November 2005, and from 1 January 2006 it will be the only system available to authors. Online submissions will be made through the very user-friendly ScholarOne. We expect that this will allow our turnaround time to be vastly improved and the whole review process will be more transparent as authors will be able to check the status of their manuscript live. To submit a new manuscript or check the status of a submitted manuscript log on to manuscript central at: http://mc.manuscriptcentral.com/res If you need assistance, have lost your login details or have any problem with the new system, you can contact the Editorial Office at: respirol@cyllene.uwa.edu.au We hope you enjoy using this new system and that you agree that this is another important improvement in our Journal.

Generation of Reactive Oxygen Species by Neutrophils and Endothelial Cell Injury in Normal and Preeclamptic Pregnancies

The aim of this study was to investigate the role of neutrophil-derived reactive oxygen species on endothelial cell dysfunction in preeclampsia. We first assessed the correlation between nitrite and superoxide anion production in normal nonpregnant (n=10), normal pregnant (n=15), and preeclamptic women (n=12). We then examined neutrophil-mediated oxygen radical damage to human umbilical vein endothelial cells in vitro. Neutrophil superoxide release was measured by cytochrome C reduction; nitrite release was measured by the modified Griess reaction, and endothelial cell injury was measured by 51Cr release. N-formyl-methionyl-leucyl-phenylalanine-stimulated superoxide release by neutrophils was significantly increased in women with preeclampsia compared with the other 2 groups. Nitrite release by neutrophils was significantly decreased in preeclampsia compared with normal pregnancy. When neutrophils were pretreated with superoxide dismutase, nitrite release by neutrophils did not differ between normal pregnancy and preeclampsia, suggesting that excess superoxide anion in preeclampsia could reduce bioavailability of nitric oxide through neutrophil autocrine function. Neutrophil-mediated endothelial cell injury was significantly greater in women with preeclampsia than in the other 2 groups. Hydrogen peroxide was important in neutrophil-mediated endothelial cell injury in preeclampsia as catalase inhibited endothelial cell injury. When neutrophils were pretreated with NG-nitro-L-arginine methyl ester, neutrophil-mediated endothelial cell injury in preeclampsia was decreased, indicating a role for peroxynitrite formation as a mechanism of endothelial cell injury. In conclusion, the modulation of neutrophils causing superoxide production to dominate over nitrite release provides a reasonable explanation for endothelial cell dysfunction in preeclampsia.

Homocysteine enhances endothelial apoptosis via upregulation of Fas-mediated pathways.

Hyperhomocysteinemia is an independent risk factor for the development of atherosclerosis. However, the underlying mechanism of endothelial cell injury in hyperhomocysteinemia has not been elucidated. In this study, we examined the effect of homocysteine (Hcy) on Fas-mediated apoptosis in endothelial cells. Hcy-induced upregulation of Fas in endothelial cells (ECs) in a dose-dependent manner. At the same time, Hcy increased intracellular peroxide in ECs. Hcy-induced Fas expression was inhibited by the treatment with catalase. Hcy increased NF-kappaB DNA binding activity, and adenovirus-mediated transfection of a Ikappa-B mutant (Ikappa-B mt) gene inhibited Hcy-induced Fas expression. ECs were sensitive to Fas-mediated apoptosis when exposed to Hcy. Under these condition, Ikappa-B mt protected ECs from Fas-mediated apoptosis. In addition, Hcy inhibited expression of the caspase-8 inhibitor FLICE-inhibitory protein (FLIP). Adenovirus-mediated transfection of constitutively active Akt gene abolished the Hcy-mediated downregulation of FLIP. These data suggest that upregulation of Fas expression and downregulation of FLIP is a mechanism through which Hcy induces EC apoptosis.

Mechanisms of endothelial cell injury in vasculitis.

The aetiology of the primary systemic vasculitides remains obscure. Recent years have seen significant advances in our understanding of inflammation and in particular the role of and interaction between the vascular endothelium, mediators and immune effector cells. This has helped to further elucidate those specific processes relevant to vasculitis which result in endothelial cell damage. In Wegener's granulomatosis and microscopic polyarteritis the evidence favours an autoimmune inflammatory response characterised by specific mediators in which the endothelium is both target and active participant. Current treatment of these disorders with combinations of corticosteroids and cytotoxics is highly effective in inducing remission. However, long-term use of this therapy is potentially toxic and there remains also a significant risk of relapse. It is hoped that increased understanding of the pathogenesis of systemic vasculitis will enable more specific, less toxic and more effective therapies to be defined. Jayne et al. have suggested a beneficial effect of intravenous pooled normal human immunoglobulin (IVIG) in patients with ANCA-positive vasculitis. In vitro studies have shown that IVIG contains antiidiotypic antibodies to ANCA and AECA, capable of inhibiting the binding of these autoantibodies to their autoantigens. In vivo, IVIG may also provide the immunoregulatory elements needed for the idiotype network and control of the autoimmune repertoire. Mathieson et al. successfully used monoclonal antibodies to T cells (Campath-H directed against CDw52) in a patient with ANCA-negative dermal lymphocytic vasculitis. Monoclonal antibodies to CAMs have been used in human renal transplant rejection and reduced the inflammation and proteinuria in animal models of anti-glomerular basement membrane disease. In vasculitis, the therapeutic use of specific anti-CAM antibodies may result from further definition of the role of CAMs. Increased understanding of the pathogenesis of systemic vasculitis is likely to provide the basis for the use of more specific immunotherapies in the future.

Mechanisms of Endothelial Cell Injury Induced by Oxidatively Modified LDL

Mechanisms of Endothelial Cell Injury Induced by Oxidatively Modified LDL

Mechanism of Endothelial Cell Injury Induced by Activated Leukocytes

Mechanism of Endothelial Cell Injury Induced by Activated Leukocytes

Overview of the Process of Cellular Injury in Interstitial Lung Disease

Since the process of cellular injury occurring in lung inflammatory diseases is highly complex and not well understood, this presentation will be directed at emerging evidence regarding mechanisms of endothelial cell injury. A variety of in vivo and in vitro techniques have been utilized to derive information related to the process of cell injury. Summarized below are studies involving a group of collaborators (Drs Hans E Friedl, Kent J. Johnson, Sem H. Phan, Una S. Ryan, Gerd O. Till, James Varani, and Jeffrey S. Warren) on the use of endothelial cells obtained from the pulmonary vasculature of rat lungs. These cells have been used in order to be able to extrapolate between in vitro and in vivo studies.