key: cord-0006027-w1x1i0im authors: Volk, T.; Kox, W.J. title: Endothelium function in sepsis date: 2000 journal: Inflamm Res DOI: 10.1007/s000110050579 sha: 59dc7ad95d8b787374db69c83fff167e6b5bfd0d doc_id: 6027 cord_uid: w1x1i0im Endothelial cells can be the prime target for an infection and infected endothelial cells may serve as an initiating system for a systemic response as these cells are able to secrete many mediators known to be of paramount importance. Endothelial cell functions in turn are regulated by these circulating mediators. Cellular interactions with leukocytes revealed protective and destructive functions. Single cell and animal studies indicate that endothelial permeability is increased and apart from clinical obvious edema formation in septic patients, the endothelial component remains unknown. Endothelial coagulation activation has been shown in vitro, however human data supporting an endothelial procoagulatory state are lacking. Defects in endothelium dependent vasoregulation in animal models are well known and again human studies are largely missing.¶An imbalanced production of reactive oxygen species including nitric oxide has been found to be involved in all endothelial functions and may provide a common link which at present can be supported only in animal studies. Sepsis is considered the leading cause of death in noncoronary intensive care units. It has been defined as a systemic inflammatory reaction to an infection. Among many cellular disturbances endothelial cells play a major role in the pathogenesis of this disease as these cells are critically involved in maintaining a delicate balance between vasoconstriction and vasodilation, blood cell adherence and nonadherence, anticoagulation and procoagulation, permeability and thightness. All these functions are believed to be imbalanced and impairment is believed to precede clinically recognizable alterations (e.g. bleeding, edema, organ dysfunction and shock) but it is by no means clear whether these changes are the cause or consequence of endothelial dysfunction in sepsis. Endothelial functions have largely been studied in vitro from the first successful attempts of culturing isolated human endothelial cells dating back to the early 70ties. Endothelial cells from and within different organs or different species behave differently and tend to change properties the longer they are kept in culture. To overcome isolation variabilities several endothelial cell lines are currently available which have retained at least some features. However, experiments from single cell cultures are flawed in many ways and results from these may never be of any relevance to medical practice. It is inherent to any cell culturing that data obtained from these experiments often are restricted to cell type, time of culture and general working conditions. In vitro stressed endothelial cells almost uniquely tend to activate a functional program leading to a proinflammatory, procoagulatory and hyperpermeable phenotype. In this review we want to summarize investigations of disturbed endothelial functions including infection, mediator and cellular interactions, permeability, coagulation and vasoactivc properties from molecular findings to patient care. Staphylococcus aureus is the most prevalent bacterial pathogen isolated from patients with blood stream infection in north america [1] . S. aureus has been reported to directly infect human umbilical vein endothelial cells (HUVEC) thereby inducing secretion of cytokines and functional upregulation of adhesion molecules [2] . Internalized S. aureus may lead to apoptosis in HUVEC [3] or persistence as small colony variants [4] . Group A streptococci can enter HUVEC [5] , a process which may render these cells particularly sen-sitive to otherwise subtoxic concentrations of hydrogen peroxide [6] . Group B streptococci (GBS) are the most common cause of neonatal sepsis and pneumonia. GBS-induced endothelial cell injury can be confirmed by histological findings at autopsy, in animal studies and in vitro [7] . GBS invasion and subsequent damage of endothelial cells may be inhibited by cytochalasin D in HUVEC implicating that cytoskeletal interactions are important for toxicity. However, invasion of brain microvascular endothelial cells by GBS may be dose dependently cytotoxic due to beta-hemolysin production [8] . Streptococcus pneumoniae may enter activated endothelial cells using PAF-receptors [9] . This receptor engagement may also serve as a sorting signal for endothelial transcytosis [10] . Infection and activation of endothelial cells by Listeria monocytogenes is believed to be a critical component of the pathogenesis of this disease and includes ceramide generation, transcription factor activation and increases in adhesion molecule expression on HUVEC [11] . Listeria have been described to enter HUVEC either directly via internalin B or by a cell-to-cell spread from infected monocytes [12] . Gram negative bacteria have lipopolysaccarides (LPS) within their cell wall. Cellular binding of LPS usually is accomplished by CD14. Endothelial cells lack CD14 receptors and LPS effects on endothelial cells generally require the presence of CD14 in the serum. LPS effects from gram negative live bacteria (B. fragilis, E. cloacae, H. influenzae, K pneumoniae) on endothelial cells have been demonstrated by transcription factor activation and subsequent surface expression of E-Selectin and tissue factor which was not seen from viable or heat-killed gram-positive bacteria (S. aureus, E. faecalis, S. pneumoniae) [13] . Neisseria meningitidis adherence on endothelial cells has been reported to be influenced by Pilus protein C expression and CD66 on endothelial surfaces [14, 15] and may cause tissue factor expression due to the presence of LPS within the cell wall. Haemophilus influenzae generally is not toxic to endothelial cells except three clones of biogroup aegyptius causing Brazilian purpuric fever [16] . Toxicity has been reported to be independent of endotoxin, phagocytosis and replication since irradiation, cycloheximide, cytochalasin D and methylamine have no effect on the ability of the bacterium to invade and cause a cytotoxic response. Piliated Pseudomonas aeruginosa adheres to and enters human endothelial cells leading to progressive damage [17] or may persist by lysis of endosomal membranes [18] . Escherichia coli may invade human brain microvascular endothelial cells involving specialized proteins [19, 20] . Endothelial infection by Chlamydia pneumoniae also activates endothelial cells to produce cytokines and adhesion molecules and become procoagulant [21] [22] [23] . Bartonella quintana, the cause of trench fever transmitted by the body louse, has recently been implicated in culture-negative endocarditis and bacteraemia amongst homeless people [24] . Infection and damage of endothelial cells by B. quintana has been demonstrated in vitro and in vivo [25] . Interaction of Bartonella henselae with endothelial cells may result in bacterial aggregation on the cell surface and the subsequent internalisation of the bacterial aggregate by a unique struc-ture, the invasome [26] . Rickettsia rickettsii, an obligate intracellular gram-negative bacterium which causes Rocky Mountain spotted fever, inhibits endothelial apoptosis allowing to remain inside the host endothelium [27] but is also known to cause a proinflammatory endothelial phenotype. Rickettsia conorii causes Mediterranean spotted fever which causes endothelial infection with secretion of cytokines, increases in adhesion molecules and induction of surface tissue factor [28, 29] . Attachment of Borrelia burgdorferi, the agent inducing Lyme disease, to endothelial cells may be accomplished by CD14, alpha(v)beta3 and alpha5beta1 integrins or different classes of proteoglycans [30] [31] [32] . B. burgdorferi activates the transcription of chemokine and adhesion in molecule gene expression in endothelial cells [33] . Plosmodium infected erythrocytes may bind to endothelial cells via P-Selectin, CD36, intercellular adhesion molecule 1 (ICAM-1) or platelet endothelial cell adhesion molecule 1 (PECAM-1) on the endothelial surface [34] [35] [36] . Phagocytosed live Candida albicans stimulates cytokine secretion and inducible cyclooxygenase expression in endothelial cells [37, 38] . Some viral diseases are known to primarily infect endothelial cells and alter their function. Dengue virus infection of HUVEC leads to production of chemoattractant proteins RANTES and IL-8 [39] , herpes and measles virus infection of brain microvascular endothelial cells increases lymphocyte adhesion by increasing ICAM-1 [40] and measles virus and cytomegalovirus increases tissue factor expression on HUVEC [41, 42] . Hemorrhagic fever caused by hantaviruses may be accomplished by integrin mediated endothelial infection [43] . On the other hand, ebola virus infects endothelial cells with a transmembrane glycoprotein and inhibits inflammatory responses [44] . Exotoxins are known to directly activate endothelial cells. Platelet activating factor (PAF), NO˙and PGI 2 secretion from HUVEC has been demonstrated by E. coli hemolysin via inositolphosphate/diacylglycerol formation and by S. aureus alpha-toxin via transmembrane Ca 2+ entry [45] . There is increasing evidence that hemolytic uremic syndrome results from the systemic action of Verocytotoxin producing E. coli on vascular endothelial cells [46] . Alpha toxin from Clostridium perfringens is a phospholipase C and has been reported to induce adhesion molecule expression and secretion of chemokines from endothelial cells [47] . Brain capillary endothelial cells have been reported to express MBEC1, a protein that may serve as the C. perfringens enterotoxin receptor [48, 49] . The activity of small GTPase Rho has been shown to be altered by C. difficile toxin B [50] and Pasteurella mulrocida toxin [51] , which leads to alterations of endothelial permeability. P. aeruginosa exotoxin A may directly injure endothelial cells by a motif shared by many toxins [52] . Listeriolysin and phosphatidylinositol-specific phospholipase C secreted from L. monocytogenes has been shown to induce phosphatidyinositol metabolism and diacylglycerol formation in the absence of bacterial uptake by the endothelial cells [53] . Some bacterial exotoxins have been used for years as pharmacological tools like Pertussis toxin for studying G-protein dependent endothelial cell functions. Lipoteichoic acid and peptidoglycan from cell wall components of gram positive bacteria have been shown to induce sepsis in animal models. While lipoteichoic acid has been reported to directly activate endothelial cells [54] , peptidoglycan seems monocyte dependent [55] . Most in vitro experiments, however, were performed with different sources of LPS as a surrogate activator modelling gram negative bacterial infection. Endothelial stimulation using LPS in relevant concentrations are usually performed in the presence of serum containing soluble CD14-receptor because endothelial cells generally lack CD14. Alternatively, proinflammatory cytokines including tumor necrosis factor alpha (TNF-a), interleukins (IL-1, IL-4) and interferon gamma (IFNg) acting on endothelial cells have extensively been investigated and shown to alter endothelial in vitro functions [56, 57] . The inflammatory response in endothelial cells has been linked to an alteration in reactive oxygen production including superoxide (O 2 -˙) , hydrogen peroxide (H 2 O 2 ), nitric oxide (NO˙), hydroxyl radicals (OH˙) and secondary reaction products thereof. O 2 -˙i s believed to be present in unstressed conditions in less than nanomolar quantities within cells. Mitochondrial and cytoplasmatic superoxide dismutase readily reacts with O 2 -˙f orming H 2 O 2 which has been measured in micromolar concentrations in human blood. Sources of endothelial O 2 -˙p roduction apart from mitochondrial leakage may include metabolism of CytP450 or other metabolic byproducts and production by a NADH oxidase system or by nitric oxide synthase in the absence of L-arginine. Some bacterial pathogens are known to be able to produce H 2 O 2 by themselves, however interaction with the endothelium may greatly enhance cellular alterations. Streptococcal hemolysin, streptolysin S, is capable of interacting with H 2 O 2 to injure vascular endothelial cells [6] . Iron bound to the P. aeruginosa siderophore, pyochelin, augments oxidantmediated endothelial cell injury by modification of transferrin to form iron complexes capable of catalyzing the formation of OH˙from O 2 -˙a nd H 2 O 2 [58] . R. rickettsii infection of the endothelial cell line EA.hy 926 and HUVEC has been demonstrated to cause glutathione depletion, a major intracellular antioxidant, and reduced glutathione peroxidase activity leading to increased amounts of intracellular peroxide [59] . An increase in reactive oxygen species production has been shown in endothelial cells after incubation with LPS, IL-1, TNF-a and IFN-g [60] [61] [62] [63] [64] [65] . TNF-a has many times been reported to increase, e.g., adhesion molecule expression inhibitable by various antioxidants [62, [66] [67] [68] . Nitric oxide, like O 2 -˙, is chemically not very reactive at all. Endothelial production has been established by either constitutive NO-synthase (NOS) III in a Ca 2+ and phosphorylation dependent manner or by inducible NOS II. Direct biochemical actions of NO˙include metalcomplex containing proteins, other radical species, oxygen and oxygen derivatives leading to oxidation, nitrosation and nitration. Relevant concentrations in vivo have been reported to range from nM to 100 mM. NO˙directly reacts with oxyhemoglobin (Fe 2 -O 2 ) leading to methemoglobin (Fe 3+ ) and nitrate (NO 3 -). NO˙reacts with O 2 forming nitrite (NO 2 -) via intermediate NO 2 and N 2 O 3 . Mainly N 2 O 3 is participating in N-or S-nitrosylations leading to nitrosamines or nitrosothioles, the latter may serve as a circulating source or as a transporter across cell membranes. Reduced glutathione has a high affinity to N 2 O 3 and may also be important in toxicity related to NO˙autoxidation. Toxicity related cellular targets of NO˙may include inhibition of cytochrome C oxidase, inhibition of catalase or DNA damage. On the other hand, NO˙has been reported to inhibit iron catalysed Fenton reaction and to inhibit lipid peroxidation. Iron nitrosyl formation in heme containing proteins are the best characterized reactions for NO˙in biology. This type of interaction includes very sensitive stimulation of guanylate cyclase and inhibition of cytochrome c oxidase. E. coli hemolysin and S. aureus alpha toxin induce NO˙formation in cultured porcine pulmonary endothelial cells [69] . Transformed mouse endothelial cells stimulated by the combination of IFN-g and TNF-a killed intracellular R.conorii by a mechanism that required the synthesis of NO˙ [70] . IFN-g, TNF-a and IL-1 stimulated murine endothelial cells have been shown to kill Schistosoma mansoni through the production of nitric oxide [71] . Both NO˙and O 2 -˙p roduction may have a limited influence on endothelial viability under resting conditions. Cultured bovine and porcine aortic endothelial cells showed decreased NO˙production after 1 h of LPS incubation which was related to decreases in capacitative Ca 2+ signals [72] . Posttranscriptional destabilization of NOS III mRNA may have accounted for this early decrease in NO˙production [73] . NOS II, which is believed to produce larger amounts of NO˙is also known to be regulated by many proinflammatory stimuli with significant cell type variablilities. At sites of endothelial involvement in an inflammatory process both vascular non-endothelial and non-resident cells are known to produce NO˙. As the amount and physiological consequences of NO˙produced from NOSs at the microcirculatory level is not known, it may well serve also to reduce inflammatory processes as recently implicated from a coculture experiment [74] . That an increase in reactive oxygen species is present in human sepsis has been documented by almost any study trying to quantify secondary reaction products by several methods, however the cellular sources remain speculative ( Table 2 ). In 1990 Beckman et al. showed that the presence of both NO˙and O 2 -˙p roduced peroxynitrite (ONOO -) which may decompose to produce HO˙like molecules and thereby kill endothelial cells [75] . At pH 7 its lifetime is in the order of a second. Half of the ONOO formed is rapidly equilibrated to peroxynitrous acid (ONOOH) and breaks down to NO 3 - 11 Sepsis Immunohistochemical endothelial nitrotyrosine≠ [176] 3 Septic shock NO 2 -/NO 3 -, plasmatic nitrotyrosine≠ [177] 3 Septic lung injury Immunohistochemical endothelial nitrotyrosine≠ Antioxidant capacity was defined as the ability of plasma to inhibit * ferryl myoglobin production by hydrogen peroxide addition to metmyoglobin or ** oxo-iron induced damage to deoxyribose, phospholipids and DNA. TBARS, thiobarbituric acid reactive substances; XOD, xanthine oxidase. models and therefore question the view that this molecule solely is detrimental. Toxicity induced by an interaction between hydrogen peroxide and nitric oxide has led to conflicting results. Rat lung microvascular endothelial cells and porcine pulmonary artery endothelial cells exposed to H 2 O 2 have been reported to be protected by NO˙donors [76, 77] , whereas toxicity in bovine aortic endothelial cells [78] and in rat liver microvascular endothelial cells [79] was increased in the presence of NO˙donors. NO˙in the presence of H 2 O 2 may also produce OH˙like molecules independent of the presence of iron [80] . Neutrophils may add to the complexity of toxic reactions. Myeloperoxidase from activated neutrophils, which produces HOCl and OH˙molecules in the presence of O 2 -˙, has recently been demonstrated to convert NO 2 into NO 2 , thereby damaging endothelial cells [81] . High doses of reactive oxygen species (including NO˙) have been shown to cause apoptosis of endothelial cells, whereas low doses were protective [82] . In mice disseminated endothelial apoptosis has been suggested to be responsible for organ failure and shock induced by endotoxin or TNF-a [83] . Both LPS or TNF-a usually do not cause endothelial cell death unless protein synthesis is blocked probably due to simultaneous increases in antiapoptotic protein synthesis [84] . However, postmortal investigation in humans deceased from or with sepsis did not confirm these results [85] . Most of the genes activated in vitro during the endothelial stress response are controlled by at least two transcription factor families: activator protein 1 (AP-1) and nuclear factor kappa B (NFkB). NFkB has gained wide interest as a target in inflammatory diseases as it seems to be invariably upregulated [86] . A variety of agents including cytokines and reactive oxygen stress cause IkB to dissociate from the complex after phosphorylation by IkB-kinase complex. H 2 O 2 has been reported to activate transcription factor NFkB in porcine aortic endothelial cells [87] whereas HUVEC were unresponsive [88] . NO˙has in most cases been shown to inhibit transcription factor NFkB and its influence on transcription factor AP-1 is unclear. Inhibition of NFk-B as a therapeutic means has been suggested. However, as this transcription factor is also involved in protective gene regulation, its inhibition can make cells sensitive to e.g. TNF-a [89] . Data on activated intracellular signalling pathways in sepsis patients are scarce but include NFkB within mononuclear cells [90] . Usually any blood cell is kept off endothelial surfaces. This is believed to be accomplished by net electrical charge, biomechanical characteristics of flowing blood and the secretion of NO˙. If blood cells touch the endothelium a Ca 2+ -signal is induced [91] , but it is unclear at present whether this has any functional consequence. It is tempting to speculate that Ca 2+signals may then in a context sensitive manner augment proadhesive processes or antiadhesive endothelial properties. Activated endothelial cells are potent producers of cytokines like IL-1, a major proinflammatory cytokine, and chemotactic peptides including IL-8 for neutrophils, macrophage inflammatory protein-1 alpha (MIP-1a), monocyte chemoattractant proteins 1-4 and RANTES (regulated on activa-tion, normal T cell expressed and secreted) for monocytes, T-lymphocytes and dendritic cells, growth related protein (GRO) and gamma-interferon-inducible protein (IP-10) for activated T-lymphocytes, epithelial neutrophil activating peptide 78 (ENA-78), vascular monocyte adhesion-associated protein (VMAP-1) and endothelial monocyteactivating polypeptide II (EMAP-II) for monocytes [92] . Many adhesion molecules are expressed on the surface of endothelial cells in a highly complex yet regulated manner. P-Selectin, E-Selectin, ICAM-1, vascular cell adhesion molecule 1 (VCAM-1), PECAM-1 are well known for their stimulus-, cell-, time-and organ specific dependence of expression and their importance in regulation of leukocyteendothelial interactions. Moreover, apart from being passive adhesion molecules, all of the above mentioned molecules have been shown to signal inside endothelial cells upon receptor engagement. Shed receptors from endothelial surfaces may also serve as endogeneous antiadhesive molecules demonstrating even more the dynamic and complex nature of these processes. Soluble forms of adhesion molecules have been shown to be present in high amounts in human sepsis (Table 3) , however whether this is beneficial or detrimental is not known and the assumption that these parameters may serve as practical indexes remains to be established. Particularly IFN-g has been repeatedly shown to increase endothelial HLA-DR expression rendering them capable of MHC class II restricted interactions with CD4 + T-cells. Costimulatory CD80 (B7-1) and CD86 (B7-2) are usually not present on endothelial surfaces leading to the conventional view that endothelial cells are semiprofessional antigen presenting cells. However, under certain conditions both costimulatory molecules and CD40 can be upregulated on endothelial surfaces indicating excessive antigen presentation [93] . Migration of lymphocytes into inflamed mouse tis-Vol. 49, 2000 Endothelial function in sepsis 189 sues has been reported to depend on PSGL-1 and ESL-1 binding endothelial P-selectin and E-selectin, respectively [94] . This phenotype was found to be restricted to Th1 lymphocytes, but human sepsis seems to be predominated by a Th2 type [95] . Both TNF-a and IFN-g are able to promote transmigration of leukocytes, whereas coapplication of both cytokines inhibit this process [96] . T-cells migrating through the endothelial barrier in a PECAM-1-dependent manner are subject to inhibitory signals which may limit their activation in tissues [97] . TNF bound to monocytes has been shown to inhibit endothelial apoptosis, whereas this process is promoted in the presence of lymphocytes [98] . Unperturbed endothelial cells express Fas-ligand which has been implicated as a means of signalling apoptosis to constitutively Fas receptor (CD95) bearing cells, whereas TNF-a treated endothelial cells decrease Fas ligand expression thereby allowing leukocyte survival during extravasation [99] . How the endothelium might interact specifically with lymphocytes or monocytes in septic patients can only be speculated on. Neutrophils interacting with endothelial cells have repeatedly been shown to cause toxicity due to the release of enzymes and reactive oxygen species. If uncontrolled, these cells are believed to mediate significant tissue damage. However whether uncontrolled activation or rather deactivation is present in human sepsis is a matter of debate. Endothelial cells have been reported to inhibit some neutrophil functions [100] . Transmigrated neutrophils may actively participate in the endothelial resealing process by the secretion of adenosin precursors [101] . However, massive leukocyte extravasation as one would expect from most animal studies has never been shown in septic patients [102] . Endothelium is known to regulate transvascular fluid flux, flux of nutrients, mediators and cells by either paracellular or transcellular vacuolar channel related pathways. Lateral junction proteins including the vascular endothelial cadherin-complex, platelet-endothelial cell adhesion molecule-1, occludin, zona occludens-1, recently described junctional adhesion protein, CD151/platelet endothelial tetraspan antigen and CD81/target of antiproliferative antigen are known to participate in this process. Paracellular permeability is achieved by either an active contraction or the controlled release of an intrinsic tone mediated in most cases by the action of myosin light chain kinase (MLCK) acting on non muscle myosin. Generally an increase in the concentration of cAMP keeps cultured endothelial monolayers tight and cGMP has been reported to assist this function in human aortic and foreskin vessels [103] . Endothelial retraction may be initiated by increases in intracellular Ca 2+ concentration, but elevation of Ca 2+ in the presence of maintained cAMP-kinase dependent phosphorylation is not edemagenic. These antagonistic effects of Ca 2+ and cAMP in endothelial permeability regulation have recently been reviewed by Moore et al. [104] . Many reports documented increases in endothelial permeability involving exotoxins like S. aureus alpha-toxin and P. aeruginosa cytotoxin [105, 106] or endotoxins [107] . For example, P. multocida toxin has been shown to activate Rho/Rho kinase, which inactivates MLC phosphatase. The resulting increase in MLC phosphorylation caused endothelial cell retraction and a rise in endothelial permeability [51] . LPS induced increases in paracellular permeability by caspase activated cleavage of adherens junction proteins [108] . Counteracting lipid peroxidation during LPS activation may inhibit increases in permeability [109] . TNF-a stimulated endothelial cadherin complex is disrupted in a proteasome dependent manner [110] . The resulting increase in permeability has been reported to lower cAMP and activate phosphediesterase II and IV [111] . H 2 O 2 induced hyperpermeability of porcine pulmonary endothelial cells has been reported to be effectively reduced by cGMP elevating drugs including phosphodiesterase II inhibition or NO˙donators [112] . The electroneutral Na-K-Cl cotransport system is thought to function in the maintenance of a selective permeability. IL-1, TNF-a and LPS upregulate the expression of a bumetanide-sensitive Na-K-Cl cotransporter subtype in HUVEC and in murine lung and kidney endothelial cells [113] . Septic rats show different increases in albumin flux accross several endothelial beds [114] . Increases in venular permeability have been shown to be preventable by the antioxidants N-acetyl-cystein or Tirilazad mesylate in E. coli infused rats [115, 116] . IL-10, an antiinflammatory cytokine not produced by endothelial cells, was shown to participate as an inhibitor of endothelial permeability induced by LPS in mice [117] . Clinically, increases in endothelial permeability may be obvious in many septic patients, but only recently venous congestion plethysmography showed a selectively elevated filtration capacity as a measure of endothelial dysfunction in septic patients [118] . Which of the many pathways of increased permeability might be turned on and whether it persists remains unknown. In principle endothelial cells are believed to be anticoagulatory by virtue of their surface expression of glycosaminoglycan-antithrombin III complex, thrombomodulin, heparin releasable tissue factor pathway inhibitor and production of adenosine by ecto-ADPases, their secretion of protein S, prostacyclin and NO˙. NO˙production was shown to participate in heparan sulfate preservation in porcine aortic endothelial cells [119] and may be responsible for prostacyclin secretion by activating cyclooxygenase-1 under resting conditions [120] . Endothelial release of plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) may determine the fibrinolytic potential of plasma. Endothelial cells specifically bind coagulation factors XII, IIa, IX, VIIa and Xa. Xa was shown to bind to endothelial effector cell protease receptor-1 and thereby cause release of NO˙, IL-6, IL-8, MCP-1 and functional upregulation of ICAM-1, E-Selectin and VCAM-1 [121] . When endothelium is perturbed by physical or chemical factors transformation to a prothrombotic surface is invariably seen in in vitro models. Thrombomodulin surface expression on HUVEC can easily be downregulated by LPS, IL-1 and TNF-a and upregulated by increasing cAMP [122] . A TNF-a induced decrease in surface Thrombomodulin has been suggested via activation of phosphodiesterase II and IV thereby decreasing cAMP in BAEC [111] . Vascular endothelial growth factor may counteract IL-1, TGF-b and LPS induced suppression of both thrombomodulin surface antigen and mRNA [123] . Adenosin nucleotides are released from damaged as well as LPS stimulated and shear stressed HUVEC [124] . Endothelial cells have ATP diphosphohydrolase (CD 39) on their surface to degrade ATP via ADP and AMP to Adenosine. Adenosine is known to have antiaggregatory properties due to stimulation of prostacyelin and NO˙production. However, activating endothelial cells with TNF-a has been reported to cause loss of ATP diphosphohydrolase activity, which was preventable in the presence of antioxidants [125] . Tissue factor (TF) expression on endothelial cells by bacteria, LPS, IL-1 and TNF-a is well known. Tissue factor pathway inhibitor (TFPI), a serine protease inhibitor of Xa and Xa/VIIa/TF complex on endothelial surfaces, which immediately blocks tissue factor activation, has been shown to be decreased under proinflammatory conditions. Another counterbalancing mechanism includes shear stress in TNF-a stimulated HUVEC [126] . However, an anticipated increase in endothelial tissue factor expression has not convincingly been demonstrated in animal or human sepsis [127, 128] . Fibrinolytic systems on endothelial surfaces are also believed to be altered in sepsis. Increases in PAI-1 has been reported after stimulation with IL-1 or LPS [129, 130] . However soluble PAI-1 in septic patients was not found to be different from nonseptic patients [131] . Once thrombin formation has occured, its cleavage of endothelial proteinase activated receptor (PAR) in turn may lead to secretion of IL-8 and IL-6 [132] , upregulation of ICAM-1 and VCAM-1 [133] , relaxation via NO˙production or to vasoconstriction by an as yet unidentified factor [134] . These data may demonstrate an interconnection between coagulation activation, inflammation and vasoregulation mediated by the endothelium. Coagulation activation is clearly present in septic patients, however endothelial participation in this process is unclear. Potent procoagulatorv sources may well include bacterial surfaces per se [135] or monocytes [128] . Up to the late 60ies the endothelium was viewed as a passive organ, which at best was able to remove vasoactive hormones in the lung. In 1976-1978 Sir John Vane's group (noble laureate 1982) reported that endothelial cells can synthesize I series prostaglandins (like prostacyclin) and thereby relax arteries and inhibit platelet aggregation. However, whether PGI 2 regulates basal vascular tone is unclear. After a new technician used an unintended vessel preparation Robert Furchgott realized after a series of contradicting results that acetylcholine (ACh) was no longer able to relax precontracted arteries when endothelium was removed and termed the nonprostanoid mediator an endothelium derived relaxing factor (EDRF). In 1986 superoxide was shown to participate in vasoregulation. The presence of superoxide dismutase prolonged the action of EDRF whereas addition of O 2 -˙i nactivated EDRF. Even direct effects of several reactive oxygen species have been suggested to be relevant in cerebral or coronary circulation. Collectively Robert Furchgott, Louis Ignarro and Ferrid Murad received the noble laureate in 1998 for demonstrating that NO˙is a major EDRF. Endothelial cells produce more relaxing factors which pharmacologically can be separated from nitric oxide action and these were termed endothelium derived hyperpolarizing factors (EDHFs). These factors may particularly be important in coronary and gastrointestinal vessels. Epoxyeicosatrienoic acids, anandamide, the endogenous ligand of cannabinoid receptors or simply the release of K + may constitute EDHFs. Conceptually shear may in larger vessels primarily determine production of NO˙, whereas cyclic strain determines physiological EDHF release. Soon after the discovery of EDRF endothelin-1 (ET-1), a vasoconstricting peptide produced from endothelial cells was isolated. Low doses of ET-1 can induce NO˙release and subsequent relaxation via ET Breceptors on endothelial cells. Endothelin secretion is thought to occur abluminally leading to ET A -receptor activation on smooth muscle cells and subsequent vasoconstriction. Many other factors clearly contribute to endothelial control of vasoregulation by e.g. transcellular production of vasoconstricting prostanoids like thromboxane A 2 or prostaglandin H 2 or the enzymatic conversion of angiotensin I to angiotensin II by angiotensin converting enzyme. A hallmark of sepsis is the heterogeneous pattern of vasoconstriction and vasodilatation in different organs, culminating in a fall in total peripheral vascular resistance concomitant with regional maldistribution of blood flow. Vasoactive substances produced by the endothelium under experimental septic conditions are known to be altered by factors such as NO˙, PGI 2 , angiotensin converting enzyme (ACE) activity, endothelin and adrenomedullin. Endothelium dependent vasoregulation has largely been studied in animal models (Table 4 ). In 1985 endothelium dependent vasoregulatory failure was seen as a defect in reactive hyperemia related vasodilator release [136] and decreased dilatation of arterioles induced by ACh [137] . In a rat model of cecal ligation and puncture decreased vasoconstriction was found after administration of norepinephrine in septic animals which was largely reversible by removal of the endothelium [138] . Parker et al. [139] showed in explanted coronary arteries and aortas of guinea pigs treated with LPS intraperitoneally for 16 h that endothelium dependent relaxation induced by acetylcholine and ADP was depressed, whereas relaxation induced by substance P or receptor independent relaxations by Ca 2+ -ionophore A23187 was unaffected. EDRF release and bioactivity from explanted aortas of these animals was decreased after ADP or ACh stimulation, whereas A23187 induced EDRF release was unaltered [140] . This group also demonstrated reduced ADP and ACh responses after 4 h of LPS endotoxemia in guinea pigs and that ADP may produce constricting thromboxane in septic animals [141] . In contrast, coronary arteries of rabbits treated for 5 weeks with low doses of LPS stimulation with ACh but not ADP showed increased relaxation of explanted vessels [142] . Wang et al. [143] isolated subepicardial arterioles from rats treated 48 h intraperitoneally with feces containing life E. coli and showed in a pressurized no-flow chamber that relaxation by alpha 2 agonist clonidine and ADP was reduced in an endothelium dependent manner, which could be inhibited by the NOS inhibitor LNMA. Also, in this model mesenteric arter-Vol. 49, 2000 Endothelial function in sepsis iolar relaxation after ADP and clonidine was decreased, whereas in skeletal muscle these agonists caused vasoconstriction [144] . Pulmonary arteries of rats treated with LPS showed depressed endothelin-1 induced contractions which were even augmented in endothelium denuded vessels. The authors [145] concluded that a vasoconstrictor eicosanoid is produced in LPS treated animals by pulmonary endothelium upon ET-1 stimulation. Swine infused with live P. aeruginosa showed no alteration in endothelium dependent bradykinin and endothelium independent nitroprusside relaxation, whereas ACh induced relaxation was found to be reduced in explanted peripheral arteries [146] . Chaudry's group investigated endothelium dependent relaxation in rats treated with cecal ligation and puncture. A time dependent alteration was demonstrated with increased ACh induced vasorelaxation early after challenge whereas depressed vasodilatation after 5-20 h was found in explanted aortas with no alteration in nitroglycerine induced relaxation [147] . The decreased endothelium dependent response to ACh was also found in superior mesenteric arteries and small intestinal arteries [148] . In the same model this group demonstrated a reduction of immunodetectable NOS III in explanted aortas [149] . Porcine coronary arterioles incubated for 20 h with E. coli LPS (100 mg/ml) decreased bradykinin induced EDHF secretion [150] . Carotid and coronary arteries from rabbits also showed decreases in EDHF-release in an ex-vivo assay after treatment with LPS, TNF-a and IL-1 [151] . Collectively these data indicate that the majority of sepsis models consistently show a disruption of receptor coupled relaxation mechanism leading to an intraendothelial signalling deficit. To quantify endothelial function in humans several methods are available. Endothelium dependent relaxation after pharmacological stimulation or flow dependent relaxation after vessel obstruction are frequently quantified by high resolution ultrasound techniques [152] , alternatively flow and size can be determined angiographically. However, definite functional measurements in human sepsis are scarce. Endothelium dependent relaxation has been investigated in isolated superficial hand veins of healthy volunteers after LPS exposure. Reduced vasorelaxation by bradykinin and arachidonic acid in noradrenalin precontracted vessels were noted which persisted for more than 2 days [153] . Reactive hyperemia is believed to mainly test endothelial NO˙production upon shear stress if vessel diameters and flow is monitored. Implications from indirect measurements support an endothelial dysfunction in septic patients [154] [155] [156] . However, data on pharmacological stimulation of endothelium dependent relaxation in humans are currently not available. 192 T. Volk +/-Ec: presence or absence of endothelium; NE: norepinephrine; ACh: acetylcholine; ADP, adenosine diphosphate; SP, substance P; A23187, Ca 2 + -ionophore; COX, cycloogygenase; SNP, sodium nitroprusside; NTG, nitroglycerine; ET, endothelin; BK, bradykinin; EDHF, endothelium derived hyperpolarizing factor; pres., preserved. Table 4 . Endothelium dependent relaxation is impaired in animal sepsis models. A normal response to infection or other insults is a self limiting process that through temporal expression of regulators and effector molecules causes resolution. The failure to resolve the causative infection may lead to sepsis. Cellular and animal models of sepsis using bacteria, endotoxins, exotoxins, cytokines or some peptides all consistently produce endothelial impairment which is usually regarded as dysfunctional. Blocking the majority of pathways used by these inducing agents has often lead to the inhibition of such endothelial alterations. Many aspects of these induced alterations can be expected to reveal exciting new pathways and complex interactions at various molecular and cellular level. The past has taught us that inhibitors of presumably activated pathways consistently failed to improve survival in septic patients. This has stimulated many researchers to reconcile the results of experimental and clinical models in sepsis. Compared to activating pathways, considerably less is known about how an inflammatory response is endogenously counterregulated. Cytokines induce a whole host of signal inhibiting proteins and endogenous counterregulating systems are just beginning to be elucidated. Activation of endogenous counterregulatory systems may become the predominant feature of the so-called compensatory antiinflammatory response syndrome. Endothelial responses to endogenously present antiinflammatory mediators have hardly been investigated in sepsis models. Almost all endothelial cell studies in sepsis indicated that an imbalance in reactive oxygen species production is associated with the above described dysfunctions. Animal studies in which endothelial function could be improved pharmacologically also consistently indicate that reversal of imbalanced reactive oxygen production may be a common link (Table 4 ). It seems that at times an adequate production of nitric oxide is lacking whereas superoxide and/or derivatives are overproduced. However, as endothelial functional measurements in septic humans become available, we will hopefully get a clearer picture of what might happen in our patients. [199] CLP, cecal ligation and puncture; LPO, lipid peroxidation; ROS, reactive oxygen species. Table 5 . Treatment of endothelial vasoregulatory dysfunction in animal sepsis models. Bacterial pathogens isolated from patients wit bloodstream infection: frequencies of occurrence and antimicrobial susceptibility patterns from the SENTRY antimicrobial surveillance program (United States and Canada, 1997) Staphylococcus aureus infections Internalization of Staphylococcus aureus by endothelial cells induces apoptosis Staphylococcus aureus small colony variants are induced by the endothelial cell intracellular milieu Genetic inactivation of the extracellular cysteine protease enhances in vitro internalization of group A streptococci by human epithelial and endothelial cells Interaction of viable group A streptococci and hydrogen peroxide in killing of vascular endothelial cells Group B streptococci invade endothelial cells: type III capsular polysaccharide attenuates invasion Invasion of brain microvascular endothelial cells by group B streptococci Streptococcus pneumoniae anchor to activated human cells by the receptor for platelet-activating factor Pneumococcal trafficking across the blood-brain barrier. Molecular analysis of a novel bidirectional pathway Two distinct phospholipases C of Listeria monocytogenes induce ceramide generation, nuclear factor-kappa acivation, and E-selectin expression in human endothelial cells Internalin B is essential for adhesion and mediates the invasion of Listeria monocytogenes into human endothelial cells Activation of human endothelial cells by viable or heat-killed gram-negative bacteria requires soluble CD14 Interaction of Neisseria maningitidis with the components of the blood-brain barrier correlates with an increased expression of PilC The Ndomain of the human CD66a adhesion molecule is a target for Opa proteins of Neisseria meningitidis and Neisseria gonorrhoeae Human microvascular endothelial tissue culture cell model for studying pathogenesis of Brazilian purpuric fever Pseudomonas aeruginosa selective adherence to and entry into human endothelial cells Endothelial function in sepsis 193 Cincomitant endosome-phagosome fusion and lysis of endosomal membranes account for Pseudomonas aeruginosa survival in human endothelial cells Endothelial cell GlcNAc beta 1-4GlcNAc epitopes for outer membrane protein A enhance traversal of Escherichia coli across the blood-brain barrier Escherichia coli invasion of brain microvascular endothelial cells in vitro and in vivo: molecular cloning and characterization of invasion gene ibe10 Characterization of a strain of Chlamydia pneumoniae isolated from a coronary atheroma by analysis of the omp1 gene and biological activity in human endothelial cells Chlamydia species infect human vascular endothelial cells and induce procoagulant activity Signal Transduction Pathways Activated in Endothelial Cells Following Infection with Chlamydia pneumoniae Bartonella (Rochalimaea) quintana endocarditis in three homeless men Bartonella quintana invades and multiplies within endothelial cells in vitro and in vivo and forms intracellular blebs Interaction of Bartonella henselae with endothelial cells results in bacterial aggregation on the cell surface and the subsequent engulfment and internalisation of the bacterial aggregate by a unique structure, the invasome NF-kappa B-dependent inhibition of apoptosis is essential for host cellsurvival during Rickettsia rickettsii infection Rickettsia conorii infection enhances vascular cell adhesion molecule-1-and intercellular adhesion molecule-1-dependent mononuclear cell adherence to endothelial cells IL-6 and IL-8 production from cultured human endothelial cells stimulated by infect on with Rickettsia conorii via a cell-associated IL-1 alpha-dependent pathway The role of CD14 in signaling mediated by outer membrane lipoproteins of Borrelia burgdorferi Integrins alpha(v)beta3 and alpha5beta1 mediate attachment of lyme disease spirochetes to human cells Different classes of proteoglycans contribute to the attachment of Borrelia burgdorferi to cultured endothelial and brain cells Borrelia burgdorferi upregulates the adhesion molecules E-selectin, P-selectin, ICAM-1 and VCAM-1 on mouse endothelioma cells in vitro Characterization of Plasmodium falciparum-infected erythrocyte and P-selectin interaction under flow conditions Intercellular adhesion molecule-1 and CD36 synergize to mediate adherence of Plasmodium falciparum-infected erythrocytes to cultured human microvascular endothelial cells PECAM-1/CD31, an endothelial receptor for binding Plasmodium falciparum-infected erythrocytes Candida albicans stimulates cytokine production and leukocyte adhesion molecule expression by endothelial cells Secreted aspartyl proteinases and interactions of Candida albicans with human endothelial cells Dengue virus infection of human endothelial cells leads to chemokine production, complement activation, and apoptosis Adhesion molecule expression and lymphocyte adhesion to cerebral endothelium: effects of measles virus and herpes simplex 1 virus Measles virus induction of human endothelial cell tissue factor procoagulant activity in vitro Effects of viral activation of the vessel wall on inflammation and thrombosis Cellular entry of hantaviruses which cause hemorrhagic fever with renal syndrome is mediated by beta3 integrins Ebola virus inhibits induction of genes by double-stranded RNA in endothelial cells Human endothelial cell activation and mediator release in response to the bacterial exotoxins Escherichia coli hemolysin and staphylococcal alpha-toxin Infection by verocytotoxin-producing Escherichia coli Alpha toxin from Clostridium perfringens induces proinflammatory changes in endothelial cells Brain capillary endothelial cells express MBEC1, a protein that is related to the Clostridium perfringens enterotoxin receptors Phospholipase C and perfringolysin O from Clostridium perfringens upregulate endothelial cellleukocyte adherence molecule 1 and intercellular leukocyte adherence molecule 1 expression and induce interleukin-8 synthesis in cultured human umbilical vein endothelial cells Glucosylation of small GTP-binding Rho proteins disrupts endothelial barrier function Pasteurella multocida toxin increases endothelial permeability via Rho kinase and myosin light chain phosphatase Evidence for a structural motif in toxins and interleukin-2 that may be responsible for binding to endothelial cells and initiating vascular leak syndrome The listerial exotoxins listeriolysin and phosphatidylinositol-specific phospholipase C synergize to elicit endothelial cell phosphoinositide metabolism Lipoteichoic acid-induced neutrophil adhesion via E-selectin to human umbilical vein endothelial cells (HUVECs) Endothelial and epithelial cells do not respond to complexes of peptidoglycan with soluble CD14 but are activated indirectly by peptidoglycan-induced tumor necrosis factor-alpha and interleukin-1 from monocytes Cytokines and endothelial cell biology Cytokine regulation of endothelial cell function: from molecular level to he bedside Pseudomonas siderophore pyochelin enhances neutrophil-mediated endothelial cell injury Superoxide dismutase-dependent, catalase-sensitive peroxides in human endothelial cells infected by Rickettsia rickettsii Superoxide release from interleukin-1B-stimulated human vascular cells: in situ electrochemical measuremeut Lipopolysaccharide enhances oxidative modification of low density lipoprotein by copper ions, endothelial and smooth muscle cells E-selectin expression in human endothelial cells by TNF-alpha-induced oxidant generation and NF-kappaB activation Superoxide responses of endothelial cells to C5a and TNF-alpha: divergent signal transduction pathways Lactosylceramide mediates tumor necrosis factor-alpha induced intercellular adhesion molecule-1 (ICAM-1) expression and the adhesion of neutrophil in human umbilical vein endothelial cells Effect of antioxidants on lipopolysaccharide-stimulated induction of mangano superoxide dismutase mRNA in bovine pulmonary artery endothelial cells Ambient but not incremental oxidant generation effects intercellular adhesion molecule 1 induction by tumour necrosis factor alpha in endothelium ICAM-1 and VCAM-1 expression induced by TNF-alpha are inhibited by a glutathione peroxidase mimic Glutathione peroxidase mimics prevent TNFalpha-and neutrophilinduced endothelial alterations Pore-forming bacterial toxins potently induce release of nitric oxide in porcine endothelial cells Cytokine-induced, nitric oxide-dependent, intracellular antirickettsial activity of mouse endothelial cells Endothelial cells are activated by cytokine treatment to kill an intravascular parasite, Schistosoma mansoni, through the production of nitric oxide Escherichia coli endotoxin inhibits agonistmediated cytosolic Ca2 + mobilization and nitric oxide biosynthesis in cultured endothelial cells Expressional control of the "constitutive" isoforms of nitric oxide synthase (NOS I and NOS III) Inducible nitric oxide: an autoregulatory feedback inhibitor of vascular inflammation Apparent hydroxyl radical production by peroxynitrite: implications for endothelial injury from nitric oxide and superoxide Nitric oxide donor prevents hydrogen peroxide-mediated endothelial cell injury Nitric oxide attenuates hydrogen peroxide-mediated injury to porcine pulmonary artery endothelial cells Protective effects of tetrahydrobiopterin against nitric oxide-induced endothelial cell death Endothelial damage induced by nitric oxide: synergism with reactive oxygen species Hydroxyl radical formation resulting from the interaction of nitric oxide and hydrogen peroxide Halliwell B van d V. Formation of nitric oxide-derived inflammatory oxidants by myeloperoxidase in neutrophils Nitric oxide inhibits lipopolysaccharide-induced apoptosis in pulmonary artery endothelial cells Lipopolysaccharide induces disseminated endothelial apoptosis requiring ceramide generation Lipopolysaccharide induces the antiapoptotic molecules, A1 and A20, in microvascular endothelial cells Apoptotic cell death in patients with sepsis, shock, and multiple organ dysfunction Nuclear factor-kappaB: a pivotal transcription factor in chronic inflammatory diseases Oxidant-sensitive and phosphorylation-dependent activation of NF-kappa B and AP-1 in endothelial cells Endothelial activation by hydrogen peroxide. Selective increases of intercellular adhesion molecule-1 and major histocompatibility complex class I Adenovirus-mediated expression of a dominant negative mutant of p65/RelA inhibits proinflammatory gene expression in endothelial cells without sensitzing to apoptosis Role of NFkappaB in the mortality of sepsis Endothelial function in sepsis Initial contact and subsequent adhesion of human neutrophils or monocytes to human aortic endothelial cells releases an endothelial intracellular calcium store Chemokines and leukocyte traffic CD40 ligation induced phenotypic and functional expression of CD80 by human cardiac microvascular endothelial cells CD4+ T cells migrate into inflamed skin only if they express ligands for E-and P-selectin T helper cell subset ratios in patients with severe sepsis Cutting edge: combined treatment of TNF-alpha and IFNgamma causes redistribution of junctional adhesion molecule in human endothelial cells A new role for platelet-endothelial cell adhesion molecule-1 (CD31): inhibition of TCRmediated signal transduction Monocytes stimulate expression of the Bcl-2 family member, A1, in endothelial cells and confer protection against apoptosis Negative Regulation of Inflammation by Fas Ligand Expression on the Vascular Endothelium Regulatory effects of endogenous protease inhibitors in acute lung inflammatory injury Neutrophil-derived 5¢-adenosine monophosphat promotes endothelial barrier function via CD73-mediated conversion to adenosine and endothelial A2B receptor activation Neutrophil migration during endotoxemia Expression of cGMP-dependent protein kinase I and phosphorylation of its substrate, vasodilator-stimulated phosphoprotein, in human endothelial cells of different origin Signal transduction and regulation of lung endothelial cell permeability interaction between calcium and cAMP Bacterial exotoxins and endothelial permeability for water and albumin in vitro Effects of Escherichia coli hemolysin on endothelial cell function Endotoxin-neutralizing protein protects against endotoxin-induced endothelial barrier dysfunction Bacterial lipopolysaccharide disrupts endothelial monolayer integrity and survival signaling events through caspase cleavage of adherens junction proteins Endotoxin-induced changes of endothelial cell viability and permeability: protective effect of a 21-aminosteroid Endothelial-dependent mechanisms regulate leukocyte transmigration: a process involving the proteasome and disruption of the vascular endothelial-cadherin complex at endothelial cell-tocell junctions TNF modulates endothelial properties by decreasing cAMP Role of nitric oxide and phosphodiesterase isoenzyme II for reduction of endothelial hyperpermeability Expression of the bumetanide-sensitive Na-K-Cl cotransporter BSC2 is differentially regulated by fluid mechanical and inflammatory cytokine stimuli in vascular endothelium Endothelial barrier resistance in multiple organs after septic and nonseptic challenges in the rat N-acetylcysteine attenuates endotoxin-induced leukocyte-endothelial cell adhesion and macromolecular leakage in vivo Effect of the 21-aminosteroid tirilazad mesylate on leukocyte adhesion and macromolecular leakage during endotoxemia Endogenous interleukin-10 regulates hemodynamic parameters, leukocyte-endothelial cell interactions, and microvascular permeability during endotoxemia Increased microvascular water permeability in patients with septic shock, assessed with venous congestion plethysmography (VCP) Endothelial-derived nitric oxide preserves anticoagulant heparan sulfate expression in cultured porcine aortic endothelial cells Does elevated nitric oxide production enhance the release of prostacyclin from shear stressed aortic endothelial cells? Hypotension and inflammatory cytokine gene expression triggered by factor Xa-nitric oxide signaling Up-regulation of thrombomodulin in human umbilical vein endothelial cells in vitro Thrombomodulin-dependent anticoagulant activity is regulated by vascular endothelial growth factor Increased release of ATP from endothelial cells during acute inflammation Loss of ATP diphosphohydrolase activity with endothelial cell activation Fluid shear stress attenuates tumor necrosis factoralpha-induced tissue factor expression in cultured human endothelial cells Cell biology of tissue factor, the principal initiator of blood coagulation On behalf of the Subcommittee on Tissue factor Pathway Inhibitor (TFPI) of the Scientific and Standardization Committee of the ISTH Effects of lipopolysaccharide on the expression of fibrinolytic factors in an established cell line from human endothelial cells Induction of plasminogen activator inhibitor type 1 and type 1 collagen expression in rat cardiac microvascular endothelial cells by interleukin-1 and its dependence on oxygencentered free radicals Plasminogen: an important hemostatic parameter in septic patients Potential mechanisms for a proinflammatory vascular cytokine response to coagulation activation Thrombin-activated human endothelial cells support monocyte adhesion in vitro following expression intercellular adhesion molecule-1 CD54) and vascular cell adhesion molecule-1 (VCAM-1; CD106) Dual endothelium-dependent vascular activities of proteinase-activated receptor-2-activating peptides: evidence for receptor heterogeneity Activation of the contact-phase system on bacterial surfaces -a clue to serious complications in infectious diseases Reactive hyperemic responses of single arterioles are attenuated markedly after intestinal ischemia, endotoxemia and traumatic shock: possible role of endothelial cells Failure of microscopic metarterioles to elicit vasodilator responses to acetylcholine, bradykinin, histamine and substance P after ischemic shock, endotoxemia and trauma: possible role of endothelial cells Vascular endothelium contributes to decreased aortic contractility in experimental sepsis Selective inhibition of endotheliumdependent vasodilator capacity by Escherichia coli endotoxemia Release of EDRF and NO in ex vivo perfused aorta: inhibition by in vivo E. coli endotoxemia Inhibition of endothelium-dependent vasodilation by Escherichia coli endotoxemia Chronic endotoxemia and endothelium-dependent vasodilation in coronary arteries Chronic septicemia alters alpha-adrenergic mechanisms in the coronary circulation Mesenteric and skeletal muscle microvascular responsiveness in subacute sepsis Contraction to endothelin-1 in pulmonary arteries from endotoxin-treated rats is modulated by endothelium Pulmonary artery endothelial cell function in swine pseudomonas sepsis Nitric oxide. To block or enhance its production during sepsis? Endothelium-dependent relaxation is depressed at the macro-and microcirculatory levels during sepsis Endothelial nitric oxide synthase is downregulated during hyperdynamic sepsis Attenuation of endothelium-dependent hyperpolarizing factor by bacterial lipopolysaccharides Proinflammatory mediators chronically downregulate the formation of the endothelium-derived hyperpolarizing factor in arteries via a nitric oxide/cyclic GMP-dependent mechanism Technical aspects of evaluating brachial artery vasodilatation using high-frequency ultrasound Local venous responses to endotoxin in humans Reactive hyperemia in patients with septic conditions Microvascular function and rheologic changes in hyperdynamic sepsis Peripheral vascular tone in sepsis Decreased antioxidant status and increased lipid peroxidation in patients with septic shock and secondary organ dysfunction Plasma antioxidant potential in severe sepsis: a comparison of survivors and nonsurvivors Complement activation and polymorphonuclear neutrophil leukocyte elastase in sepsis Correlation with severity of disease Xanthine oxidase activity and free radical generation in patients with sepsis syndrome Ascorbyl radical formation in patients with sepsis: effect of ascorbate loading The effects of intravenous antioxidants in patients with septic shock Nitrogen oxide levels in patients after trauma and during sepsis Evidence of increased nitric oxide production in patients with the sepsis syndrome Nitrite/nitrate oxide (NOx) and cytokine levels in patients with septic shock L-arginine: nitric oxide pathway in endotoxemia and human septic shock Relationship between circulating levels of calcitonin gene-related peptide, nitric oxide metabolites and hemodynamic changes in human septic shock Assessment of inflammatory cytokines, nitrate/nitrite, type II phospholipase A2, and soluble adhesion molecules in systemic inflammatory response syndrome Plasma nitrite and nitrate concentrations and multiple organ failure in pediatric sepsis Endothelial function in sepsis Measurements of total plasma nitrite and nitrate in pediatric patients with the systemic inflammatory response syndrome Activation of the L-arginine nitric oxide pathway in severe sepsis Increased serum nitrite and nitrate concentrations in children with the sepsis syndrome Circulating methemoglobin and nitrite/nitrate concentrations as indicators of nitric oxide overproduction in critically ill children with septic shock Effect of L-NAME, an inhibitor of nitric oxide synthesis, on plasma levels of IL-6, IL-8, TNF alpha and nitrite/nitrate in human septic shock Extensive tyrosine nitration in human myocardial inflammation: evidence for the presence of peroxynitrite Clinical evidence of peroxynitrite formation in chronic renal failure patients with septic shock Evidence for in vivo peroxynitrite production in human acute lung injury Elevated von Willebrand factor antigen is an early plasma predictor of acute lung injury in nonpulmonary sepsis syndrome Increased plasma levels of soluble thrombomodulin in patients with sepsis and organ failure Endothelial cell activity varies in patients at risk for the adult respiratory distress syndrome Soluble E-selectin levels in sepsis and critical illness. Correlation with infection and hemodynamic dysfunction Die zirkulierenden Adhäsionsmoleküle sICAM-1 und sB-Selectin bei Patienten mit Sepsis Elevated circulating E-selectin, intercellular adhesion moleculc 1, and von Willebrand factor in patients with severe infection Increased circulating thrombomodulin in children with septic shock Systemic endothelial activation occurs in both mild and severe malaria. Correlating dermal microvascular endothelial cell phenotype and soluble cell adhesion molecules with disease severity Increased plasma von Willebrand factor in the systemic inflammatory response syndrome is derived from generalized endothelial cell activation Blood levels of endothelin-1 and thrombomodulin in patients with disseminated intravascular coagulation and sepsis Plasma levels of endothelial cell protein C receptor are elevated in patients with sepsis and systemic lupus or erythematosus: lack of correlation with thrombomodulin suggests involvement of different pathological processes Demonstration of Rickettsia conorii-induced endothelial injury in vivo by measuring circulating endothelial cells, thrombomodulin, and von Willebrand factor in patients with Mediterranean spotted fever Influence of angiotensin-converting enzyme inhibitor enalaprilat on endothelial-derived substances in the critically ill Mesenteric and skeletal muscle microvascular responsiveness in subacute sepsis Influence of group B streptococci on piglet pulmonary artery response to bradykinin Effects of antisense oligonucleotide to iNOS on hemodynamic and vascular changes induced by LPS Pentoxifylline maintains vascular endothelial cell function during hyperdynamic and hypodynamic sepsis A novel nonanticoagulant heparin prevents vascular endothelial cell dysfunction during hyperdynamic sepsis Effect of endotoxin-enhanced hepatic reperfusion injury on endonthelium-dependent relaxation in rat aorta Splanchnic vascular endothelial dysfunction in rat endotoxemia: role of superoxide radicals Endothelial dysfunction in a rat model of endotoxic shock. Importance of the activation of poly (ADP-ribose) synthetase by peroxynitrite Endothelial cell dysfunction in septic shock