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M S Goligorsky

Publications and source records attributed to M S Goligorsky.

At least 19 recordsLinked to original sources
PubMed2006

Meeting report: ISN forefronts in nephrology on endothelial biology and renal disease: from bench to prevention.

This ISN-sponsored Forefront in Nephrology meeting, which has brought together 120 scientists from 21 countries, has been concerned with various aspects of endothelial function and dysfunction and their contribution to progression of chronic kidney disease and/or its cardiovascular complications. The following themes were discussed in great depth: (1) phenotypical changes in the vascular endothelium - permeability, senescence, and apoptosis; (2) regulation of endothelial nitric oxide (NO) synthase function - caveolar and shear stress mechanisms, epigenetic regulation, S-nitrosylation, and Rho-kinase regulation; (3) oxidative stress and hypoxia-induced changes; (4) organellar dysfunction - lysosomes, mitochondria, and endoplasmic reticulum; (5) NO-independent mechanisms of vasomotion - epoxides, heme oxygenase-1 and carbon monoxide, thromboxane, tumor necrosis factor-alpha, and uric acid; (6) endothelial crosstalk with podocytes, monocytes, smooth muscle cells, and platelets; (7) candidate clinical biomarkers of endothelial dysfunction - functional testing of macro- and micro-vascular functions, surrogate markers, circulating detached endothelial cells, and endothelial precursor cells; and culminated in Round Table discussion on the diagnosis of endothelial dysfunction and its treatment options. In conclusion, this meeting has focused on several key problems of endothelial cell pathobiology relevant to chronic kidney disease.

Animals↗
PubMed2006

Laser Doppler flowmetry detection of endothelial dysfunction in end-stage renal disease patients: correlation with cardiovascular risk.

Prediction of cardiovascular (CV) complications represents the Achilles' heel of end-stage renal disease. Surrogate markers of endothelial dysfunction have been advocated as predictors of CV risk in this cohort of patients. We have recently adapted a noninvasive laser Doppler flowmetry (LDF) functional testing of endothelium-dependent microvascular reactivity and demonstrated that end-stage renal disease patients are characterized by profound alterations in thermal hyperemic responsiveness. We hypothesized that such functional assessment of the cutaneous microcirculation may offer a valid, noninvasive test of the severity of endothelial dysfunction and CV risk. To test this hypothesis, we performed a cross-sectional study, in which we compared LDF measurements to conventional risk factors, and performed a pilot longitudinal study. LDF studies were performed in 70 patients and 33 controls. Framingham and Cardiorisk scores were near equivalent for low-risk patients, but more divergent as risk increased. C reactive protein (CRP) levels and LDF parameters (amplitude of thermal hyperemia (TH), area under the curve of TH) showed significant abnormality in high-risk vs low-risk patients calculated using either Framingham or Cardiorisk scores. Patients who had abnormal LDF parameters showed increased CV mortality, however, had similar risk assessments (Framingham, Cardiorisk, CRP, and homocysteine) to those with unimpaired LDF tracings. In conclusion, LDF parameters of microvascular reactivity offer a sensitive characterization of endothelial dysfunction, which may improve CV risk assessment through incorporation into the Framingham or Cardiorisk algorithm.

Adult↗
PubMed2004

Asymmetric dimethylarginine upregulates LOX-1 in activated macrophages: role in foam cell formation.

Intimal infiltration by monocytes and accumulation of lipids represent a critical step in the formation of fatty streaks during atherogenesis. Because elevated plasma levels of asymmetric dimethylarginine (ADMA), a potent nitric oxide (NO) synthase (NOS) inhibitor, are prevalent in diverse cardiovascular diseases, the goal of this study was to examine the contribution of NO deficiency to macrophage lipid accumulation. Inhibition of NO synthesis in PMA-primed human monocytic leukemia HL-60 cells resulted in a twofold increase in expression of the receptor for oxidized LDL (OxLDL), termed the lectin-like OxLDL receptor (LOX-1). Blockade of inducible NOS in activated macrophages resulted in 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI)-OxLDL accumulation and imparted macrophages with a foamy appearance as detected with oil-red O lipid staining. ADMA (15 microM) or N(G)-nitro-l-arginine methyl ester (l-NAME, 300 microM), both of which suppress inducible NOS activity, increased oil-red staining 1.9- and 2.8-fold, respectively. Macrophages treated with ADMA or l-NAME showed a 2.4-fold increase in accumulation of DiI-OxLDL. To examine the role of LOX-1 in this process, we used small interfering RNA (siRNA) duplex-mediated LOX-1 gene silencing. LOX-1 expression was suppressed twofold by siRNA as shown by Western blot analysis. This suppression was associated with a two- to fourfold decrease in DiI-OxLDL uptake as identified by fluorescence microscopy and decreased oil-red O staining by activated macrophages. In conclusion, accumulation of ADMA (a competitive inhibitor of NOS) in patients with chronic renal failure may be responsible for upregulation of LOX-1 receptor and increased OxLDL uptake, thus contributing to lipidosis and foam cell formation. The data illustrate an additional nonendothelial mode of antiatherogenic action of NO: prevention of LOX-1 induction and lipid accumulation by macrophages.

Arginine↗
PubMed2004

Upregulation of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) in endothelial cells by nitric oxide deficiency.

Endothelial cell dysfunction (ECD) is emerging as a common denominator for diverse cardiovascular abnormalities associated with inhibition of endothelial nitric oxide (NO) synthase (eNOS). Elevated levels of asymmetric dimethylarginine (ADMA), a potent eNOS inhibitor, are common in renal failure and may contribute to ECD. Through DNA microarray screening of genes modulated in human umbilical vein endothelial cells (HUVEC) by N(G)-nitro-l-arginine methyl ester (l-NAME), we found a 1.8-fold increase in low-density lipoprotein receptor-1 (LOX-1) expression. LOX-1 is a major endothelial receptor for oxidized low-density lipoproteins (OxLDL) and is assumed to play a role in the initiation and progression of atherosclerosis. Here, we confirmed the upregulation of LOX-1 mRNA and protein level by quantitative RT-PCR and Western blot analysis. Increased expression of LOX-1 was associated with the accumulation of DiI-labeled OxLDL (DiI-OxLDL) in ADMA- and l-NAME-pretreated HUVEC. To evaluate the contribution of LOX-1 in ADMA-induced accumulation of OxLDL by HUVEC, we used the competitive receptor inhibitor, soluble LOX-1. Treatment of HUVEC with soluble LOX-1 was associated with an approximately two- to threefold inhibition of DiI-OxLDL uptake in l-NAME- or ADMA-treated HUVEC. In conclusion, ADMA- or l-NAME-induced NO deficiency leads to the increased expression of LOX-1 mRNA and protein in HUVEC, which in turn results in the accumulation of OxLDL. Competition with LOX-1-soluble extracellular domain reduces OxLDL accumulation. In summary, elevated ADMA levels, i.e., in patients with renal failure, may be responsible for endothelial accumulation of OxLDL via upregulated LOX-1 receptor, thus contributing to endothelial lipidosis and dysfunction.

Arginine↗
PubMed2001

Nitric oxide attenuates signal transduction: possible role in dissociating caveolin-1 scaffold.

Caveolae harbor different serpentine receptors, intracellular components of signaling cascades, and certain enzymes, including endothelial nitric oxide synthase (eNOS). The regulation of eNOS activity by Ca(2+)/calmodulin and caveolin has been described. We have previously demonstrated that nitric oxide (NO) can modulate signaling initiated via receptors localized to caveolae. In the present study, we show that NO donors induced an increase in the monomeric form of this scaffolding protein in cultured endothelial cells, the effect mimicked by 8-bromo cGMP. Proximity imaging of endothelial cells transfected with the thermotolerant green fluorescent protein-caveolin-1 construct demonstrated that sodium nitroprusside resulted in the increased fluorescence ratio of 410:470 nm, consistent with the distancing of fluorescently tagged caveolin-1. Pulse labeling of endothelial cells with cholera toxin B subunit indicated that sodium nitroprusside reversibly decreased its binding. Signaling via G protein-coupled receptors resident to caveolae was inhibited by pretreatment with NO donor. The data demonstrate that NO modulation of cell signaling is accomplished in part by regulating the state of caveolin-1 oligomerization. NO-induced attenuation of signaling involves reversible dissociation of caveolin scaffold, thus providing both spatial and temporal modulation of signal transduction.

Animals↗
PubMed2001

Plasmin-dependent and -independent effects of plasminogen activators and inhibitor-1 on ex vivo angiogenesis.

Plasminogen activator (PA) inhibitor-1 (PAI-1) has been recognized as a surrogate marker of endothelial dysfunction in diseases associated with impaired angiogenesis, including atherosclerosis, diabetic vasculopathy, and nephropathy. To establish the necessary and sufficient components of the PA system [PAI-1, urokinase-type PA (uPA), or tissue-type PA (tPA), and plasminogen (Plg)] for angiogenesis, we examined angiogenic competence of vascular explant cultures obtained from mice deficient in PAI-1, tPA, uPA, and Plg. To gain insight into the requirement for different matrix-degrading systems during endothelial cell migration across plasmin-degradable basement membranes compared with profibrotic areas containing plasmin-nondegradable collagen, we contrasted vascular sprouting in collagen with Matrigel lattices. PAI-1(-/-) vessels showed an increased capillary sprouting in both collagen and Matrigel. Deficiency of uPA significantly reduced the rate of sprouting, whereas tPA(-/-) vessels showed a profound inhibition of capillary sprouting. The Plg(-/-) vessels failed to sprout, a defect that was restored not only by exogenous Plg, but also by the addition of PAs; a nonproteolytic effect of tPA was observed in Matrigel. Zymography revealed no differences in the activity of metalloproteinase (MMP)-2 and -9 in wild-type and PAI-1(-/-) vessels, but demonstrated reduced MMP-9 activity in all angiogenesis-deficient vessels. In summary, 1) PAI-1 by itself is a modest inhibitor of endothelial sprouting, 2) tPA and Plg are indispensable for angiogenesis in this model, 3) Plg is not the only substrate for PAs, and 4) the activity of MMP-9 is undetectable in explant cultures from tPA and Plg knockout mice.

Animals↗
PubMed2001

Glucose scavenging of nitric oxide.

Endothelial dysfunction accompanies suboptimal glucose control in patients with diabetes mellitus. A hallmark of endothelial dysfunction is a deficiency in production or bioavailability of vascular nitric oxide (NO). Here we demonstrate that acute exposure of human endothelial cells to glucose, at levels found in plasma of diabetic patients, results in a significant blunting of NO responses to the endothelial nitric oxide synthase (eNOS) agonists bradykinin and A-23187. Monitoring of NO generation by purified recombinant bovine eNOS in vitro, using amperometric electrochemical detection and an NO-selective porphyrinic microelectrode, showed that glucose causes a progressive and concentration-dependent attenuation of detectable NO. Addition of glucose to pure NO solutions similarly elicited a sharp decrease in NO concentration, indicating that glucose promotes NO loss. Electrospray ionization-tandem mass spectrometry, using negative ion monitoring, directly demonstrated the occurrence of a covalent reaction involving unitary addition of NO (or a derived species) to glucose. Collectively, our findings reveal that hyperglycemia promotes the chemical inactivation of NO; this glucose-mediated NO loss may directly contribute to hypertension and endothelial dysfunction in diabetic patients.

Acids↗
PubMed2001

The concept of cellular "fight-or-flight" reaction to stress.

As animals respond to environmental stress with a set of default reactions described as the "fight-or-flight" response, so do epithelial and endothelial cells when they are confronting stressors in their microenvironment. This review will summarize a growing body of data suggesting the existence of a set of stereotypical cellular reactions to stress, provide some examples of diseases that are characterized by excessive flight reactions, describe the cellular mechanisms whereby the fight-or-flight reaction is accomplished, as well as cellular mechanisms triggering either fight or flight. It is proposed that cell-matrix adhesion is a sensitive indicator of the severity of stress. This indicator is interfaced with several default programs for cellular survival or death, thus dictating the fate of the cell. Some diagnostic and therapeutic applications of the concept, presently used and potentially useful, are outlined. The essential feature of this concept is its ability to categorize cellular events in terms of either type of default reaction, predict the details of each, and potentially exploit them clinically.

Animals↗
PubMed2001

Delayed branching of endothelial capillary-like cords in glycated collagen I is mediated by early induction of PAI-1.

Development of micro- and macrovascular disease in diabetes mellitus (DM) warrants a thorough investigation into the repertoire of endothelial cell (EC) responses to diabetic environmental cues. Using human umbilical vein EC (HUVEC) cultured in three-dimensional (3-D) native collagen I (NC) or glycated collagen I (GC), we observed capillary cord formation that showed a significant reduction in branching when cells were cultured in GC. To gain insight into the molecular determinants of this phenomenon, HUVEC subjected to GC vs. NC were studied using a PCR-selected subtraction approach. Nine different genes were identified as up- or downregulated in response to GC; among those, plasminogen activator inhibitor-1 (PAI-1) mRNA was found to be upregulated by GC. Western blot analysis of HUVEC cultured on GC showed an increase in PAI-1 expression. The addition of a neutralizing anti-PAI-1 antibody to HUVEC cultured in GC restored the branching pattern of formed capillary cords. In contrast, supplementation of culture medium with the constitutively active PAI-1 reproduced defective branching patterns in HUVEC cultured in NC. Ex vivo capillary sprouting in GC was unaffected in PAI-1 knockout mice but was inhibited in wild-type mice. This difference persisted in diabetic mice. In conclusion, the PCR-selected subtraction technique identified PAI-1 as one of the genes characterizing an early response of HUVEC to the diabetic-like interstitial environment modeled by GC and responsible for the defective branching of endothelial cells. We propose that an upregulation of PAI-1 is causatively linked to the defective formation of capillary networks during wound healing and eventual vascular dropout characteristic of diabetic nephropathy.

Animals↗
PubMed2001

Oxidative and nitrosative stress in acute renal ischemia.

Generation of reactive oxygen species and nitric oxide in hypoxia-reperfusion injury may form a cytotoxic metabolite, peroxynitrite, which is capable of causing lipid peroxidation and DNA damage. This study was designed to examine the contribution of oxidative and nitrosative stress to the renal damage in ischemic acute renal failure (iARF). iARF was initiated in rats by 45-min renal artery clamping. This resulted in lipid peroxidation, DNA damage, and nitrotyrosine modification confirmed both by Western and immunohistochemical analyses. Three groups of animals were randomly treated with an inhibitor of inducible nitric oxide synthase (NOS), L-N(6)-(1-iminoethyl)lysine (L-Nil), cell-permeable lecithinized superoxide dismutase (SOD), or both. Each treatment resulted in amelioration of renal dysfunction, as well as reduced nitrotyrosine formation, lipid peroxidation, and DNA damage, thus suggesting that peroxynitrite rather than superoxide anion is responsible for lipid peroxidation and DNA damage. Therefore, in a separate series of experiments, a scavenger of peroxynitrite, ebselen, was administered before the reperfusion period. This treatment resulted in a comparable degree of amelioration of iARF. In conclusion, the present study provides the first attempt to elucidate the role of peroxynitrite in initiation of the cascade of lipid peroxidation and DNA damage to ischemic kidneys. The results demonstrate that L-Nil, lecithinized SOD, and ebselen treatments improve renal function due to their suppression of peroxynitrite production or its scavenging, consequently preventing lipid peroxidation and oxidative DNA damage.

Animals↗
PubMed2001

Workshop: endothelial cell dysfunction leading to diabetic nephropathy : focus on nitric oxide.

Clinical manifestations of diabetic nephropathy are an expression of diabetic microangiopathy. This review revisits the previously proposed Steno hypothesis and advances our hypothesis that development of endothelial cell dysfunction represents a common pathophysiological pathway of diabetic complications. Specifically, the ability of glucose to scavenge nitric oxide is proposed as the initiation phase of endothelial dysfunction. Gradual accumulation of advanced glycated end products and induction of plasminogen activator inhibitor-1, resulting in the decreased expression of endothelial nitric oxide synthase and reduced generation of nitric oxide, are proposed to be pathophysiologically critical for the maintenance phase of endothelial dysfunction. The proposed conceptual shift toward the role of endothelial dysfunction in diabetic complications may provide new strategies for their prevention.

Albuminuria↗
PubMed2001

The ins and outs of endothelial dysfunction: much ado about NO-thing.

This article presents data linking eNOS function and nitric oxide (NO) generation or availability to vascular remodeling and signaling, hyperlipidemias, advanced glycated end products and hyperglycemia. These data should be viewed within the broader framework of endothelial cell dysfunction (ECD). It is possible that vascular dysfunctions are capable of triggering early preclinical forms of generalized ECD. Accordingly, it is important to learn more about simple noninvasive ways to assess the functional state of eNOS and the bioavailability of NO. Here we discuss the growing body of evidence--which suggests that disturbances of NO production or availability are major determinants of ECD--and the need for therapeutic efforts toward correction of eNOS activity and NO levels in blood vessels.

Journal Article↗
PubMed2000

A pivotal role of nitric oxide in endothelial cell dysfunction.

The functional role of the vascular endothelium is a subject of growing interest and appreciation. Some of the key functions of the endothelium are modulated by the activity and expression of endothelial nitric oxide synthase (eNOS), suggesting a role for this enzyme in endothelial dysfunction. Several well-known angiogenic stimulators exert their effect only in the presence of the functional eNOS. In this setting NO production is responsible for the scalar podokinetic cell motility, which is a prerequisite for the acquisition of vectorial movement when guidance cues are applied. The mode of this NO action appears to lie in the accelerated turnover of focal adhesions through the process of activation/inactivation of protein tyrosine phosphatases. Localization of eNOS to the caveolar domains, in the proximity of clustered beta1 integrins, provides an additional level of regulatory complexity through the modulation of caveolar dynamics and the state of caveolin oligomerization. Therefore, eNOS serves various important functions in the endothelium and is a putative target for therapeutic interventions.

Animals↗
PubMed2000

Effects of homocysteine on endothelial nitric oxide production.

Hyperhomocysteinemia (HHCy) is an independent and graded cardiovascular risk factor. HHCy is prevalent in patients with chronic renal failure, contributing to the increased mortality rate. Controversy exists as to the effects of HHCy on nitric oxide (NO) production: it has been shown that HHCy both increases and suppresses it. We addressed this problem by using amperometric electrochemical NO detection with a porphyrinic microelectrode to study responses of endothelial cells incubated with homocysteine (Hcy) to the stimulation with bradykinin, calcium ionophore, or L-arginine. Twenty-four-hour preincubation with Hcy (10, 20, and 50 microM) resulted in a gradual decline in responsiveness of endothelial cells to the above stimuli. Hcy did not affect the expression of endothelial nitric oxide synthase (eNOS), but it stimulated formation of superoxide anions, as judged by fluorescence of dichlorofluorescein, and peroxynitrite, as detected by using immunoprecipitation and immunoblotting of proteins modified by tyrosine nitration. Hcy did not directly affect the ability of recombinant eNOS to generate NO, but oxidation of sulfhydryl groups in eNOS reduced its NO-generating activity. Addition of 5-methyltetrahydrofolate restored NO responses to all agonists tested but affected neither the expression of the enzyme nor formation of nitrotyrosine-modified proteins. In addition, a scavenger of peroxynitrite or a cell-permeant superoxide dismutase mimetic reversed the Hcy-induced suppression of NO production by endothelial cells. In conclusion, electrochemical detection of NO release from cultured endothelial cells demonstrated that concentrations of Hcy >20 microM produce a significant indirect suppression of eNOS activity without any discernible effects on its expression. Folates, superoxide ions, and peroxynitrite scavengers restore the NO-generating activity to eNOS, collectively suggesting that cellular redox state plays an important role in HCy-suppressed NO-generating function of this enzyme.

Animals↗
PubMed1999

RGD-recognizing integrins mediate interactions of human prostate carcinoma cells with endothelial cells in vitro.

BACKGROUND: Interactions of cancer cells with endothelium are a crucial step in metastatic invasion. RGD-recognizing integrins play a definitive role in these interactions. METHODS: Fluorescence-activated cell sorting (FACS) analysis of RGD-sensitive integrins in prostate epithelial cells was performed. Attachment inhibition assay was used to characterize functionality of particular integrins. Potential partners for RGD-binding integrins in human umbilical vein endothelial cells (HUVEC) were identified by Western blotting and attachment inhibition assay. To determine the RGD-flanking amino acids optimal for interactions with prostate cell integrins, these cells were biopanned with a phage library. RESULTS: Different expressions of RGD-recognizing integrins and distinctions in RGD-dependent adhesion of nonmalignant and cancer cells were observed. Cancer but not control cells were detached from culture plastic by incubation with RGD peptide. Adhesion of carcinoma cells to HUVEC was RGD-sensitive, in contrast to nonmalignant cells. Antibodies against alpha3, alpha5, beta1, and alpha(v)beta3 inhibited interactions of carcinoma cells with HUVEC. Potential ligands for alpha5beta1, alpha3beta1, and alphaVbeta3 integrins, fibronectin, and vitronectin, were detected on the HUVEC surface. Several phages which preferentially bound to the surface of particular prostate cells were selected. CONCLUSIONS: Interactions of prostate carcinoma with endothelium are mediated in part via alpha5beta1, alpha3beta1, and alpha(v)beta3 integrins. Because these interactions are RGD-sensitive, synthetic RGD peptides with optimized flanking amino acids can potentially be used as antimetastatic agents.

Amino Acid Sequence↗