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Biomedical subjects

C Erik Hack

Publications and source records attributed to C Erik Hack.

At least 55 records · Page 3Linked to original sources
PubMed2004

Inhibition of plasmin activity by tranexamic acid does not influence inflammatory pathways during human endotoxemia.

OBJECTIVE: Plasmin activates several proinflammatory pathways at the cellular level in vitro. Lipopolysaccharide (LPS) administration to healthy humans results in a rapid generation of plasmin activity, accompanied by activation of a number of inflammatory systems. METHODS AND RESULTS: To determine the role of early plasmin activity in LPS-induced inflammation in vivo, 16 healthy males received an intravenous bolus injection with LPS (from Escherichia coli, 4 ng/kg) directly preceded by a 30-minute intravenous infusion of tranexamic acid (2 g, n=8), a plasmin activation inhibitor, or placebo (n=8). LPS injection induced marked increases in the plasma levels of D-dimer and plasmin-alpha2-antiplasmin complexes, indicative of plasmin activation and generation, respectively, which were strongly attenuated by tranexamic acid (both P<0.01 versus placebo). However, tranexamic acid did not influence LPS-induced coagulation activation, granulocytosis, neutrophil activation (expression of CD11b, CD66b, and L-selectin) or degranulation (plasma concentrations of elastase-alpha1-antitrypsin and bactericidal permeability-increasing protein), endothelial cell activation (plasma levels of von Willebrand factor and soluble E-selectin), or cytokine release. CONCLUSIONS: These data argue against a role of early plasmin generation in the subsequent activation of other inflammatory pathways during human endotoxemia.

Adult↗
PubMed2004

Complement activation by necrotic cells in normal plasma environment compares to that by late apoptotic cells and involves predominantly IgM.

Necrotic cells are generally considered to stimulate inflammation, whereas apoptotic cells should not. However, apoptotic cells have pro-inflammatory properties since they can activate complement. To what extent this activation compares to that by necrotic cells is not known. We compared complement activation by necrotic cells and apoptotic cells in plasma. Jurkat cells were made apoptotic or necrotic by incubation with etoposide or by heat shock, respectively. Cells incubated in recalcified plasma were tested for C3 and C4 fixation and fluid phase generation of complement activation products. Fixation of C3 and C4 to necrotic cells occurred mainly via the classical pathway, independent from the method of necrosis induction and the cell type. Depletion of IgM from plasma almost completely abrogated complement fixation by necrotic cells, which was restored by supplementation with purified IgM. Complement activation by late apoptotic cells was comparable to that by necrotic cells regarding the extent and dependence on IgM. Moreover, incubation of plasma with necrotic or late apoptotic cells led to the generation of comparable amounts of complement activation products. These results indicate that late apoptotic and necrotic cells employ similar complement activation mechanisms in the plasma environment.

Apoptosis↗
PubMed2004

Lymphocytic inflammation in childhood bronchiolitis obliterans.

Childhood bronchiolitis obliterans (CBO) is an infrequent, severe disorder characterized by persistent obstructive respiratory symptoms after an acute episode of bronchiolitis. The viral etiology is most common, and adenovirus is the most frequently identified causative agent. Pathologically, the disease is characterized as constrictive type BO, with variable degrees of chronic inflammation and fibrosis in the bronchioles. The nature of the cellular infiltrate is largely unknown, and its characterization may provide better understanding of the disease and offer clues for therapy. Therefore, the aim of the present study was to characterize the inflammatory infiltrate in the bronchioles of 23 open lung biopsies of children with CBO and to compare this to the infiltrate in histologically normal airways. Our results show that CD3+ T cells were the most frequent cell type observed in CBO, with a predominance of the CD8+ T-cell subtype. When compared to the control group, there was a larger number of CD8+, CD4+, CD20+, granzyme B+, and perforin+ lymphocytes in the CBO group. Further studies are needed to address the role of different cell types in the development of CBO.

Adolescent↗
PubMed2004

N(omega)-(carboxymethyl)lysine depositions in human aortic heart valves: similarities with atherosclerotic blood vessels.

Recent studies indicate a role of atherosclerosis-like changes involved in the pathogenesis of aortic valve stenosis. Interestingly, one of the major advanced glycation end products (AGEs), N(omega)-(carboxymethyl)lysine (CML) has been related to the process of atherosclerosis in blood vessels. In the present study, we have analyzed the presence of CML in degenerative altered aortic valves with atherosclerosis-like changes, and in degenerated mitral valves without atherosclerosis-like changes, derived from patients suffering from acute rheumatism during childhood. Degenerated and non-degenerated valves were derived from autopsy or obtained during cardiac surgery. The presence of CML was examined by immunohistochemistry. CML was found on the endothelium and fibroblasts in control aortic and mitral valves. Minor differences in CML staining were observed between control and degeneratively affected mitral valves. In contrast, in degenerated aortic valves, CML accumulation was found in macrophages and on calcification sites, comparable to that in atherosclerotic arteries, while the presence of CML staining on the endothelium and fibroblasts was significantly less as compared with control aortic valves. Our data support the hypothesis that the process of degeneration of aortic valves resembles that of atherosclerosis in blood vessels. They suggest that CML also plays a role in the process of atherosclerosis in aortic valves.

Adult↗
PubMed2004

Hereditary and acquired angioedema: problems and progress: proceedings of the third C1 esterase inhibitor deficiency workshop and beyond.

Hereditary angioedema (HAE), a rare but life-threatening condition, manifests as acute attacks of facial, laryngeal, genital, or peripheral swelling or abdominal pain secondary to intra-abdominal edema. Resulting from mutations affecting C1 esterase inhibitor (C1-INH), inhibitor of the first complement system component, attacks are not histamine-mediated and do not respond to antihistamines or corticosteroids. Low awareness and resemblance to other disorders often delay diagnosis; despite availability of C1-INH replacement in some countries, no approved, safe acute attack therapy exists in the United States. The biennial C1 Esterase Inhibitor Deficiency Workshops resulted from a European initiative for better knowledge and treatment of HAE and related diseases. This supplement contains work presented at the third workshop and expanded content toward a definitive picture of angioedema in the absence of allergy. Most notably, it includes cumulative genetic investigations; multinational laboratory diagnosis recommendations; current pathogenesis hypotheses; suggested prophylaxis and acute attack treatment, including home treatment; future treatment options; and analysis of patient subpopulations, including pediatric patients and patients whose angioedema worsened during pregnancy or hormone administration. Causes and management of acquired angioedema and a new type of angioedema with normal C1-INH are also discussed. Collaborative patient and physician efforts, crucial in rare diseases, are emphasized. This supplement seeks to raise awareness and aid diagnosis of HAE, optimize treatment for all patients, and provide a platform for further research in this rare, partially understood disorder.

Angioedema↗
PubMed2004

Levels of natural IgM antibodies against phosphorylcholine in healthy individuals and in patients undergoing isolated limb perfusion.

Natural IgM antibodies against phosphorylcholine (anti-Pc IgM) resemble C-reactive protein (CRP) regarding specificity and have gained increasing attention because of their supposed role in clearance of damaged cells and in cardiovascular disease. In order to quantify these antibodies in human plasma, we have developed an ELISA system, in which p-aminophenylphosphorylcholine (PCH) coupled to human serum albumin (HSA) was coated on microtiters plates. Human plasma or serum samples were incubated in the plates, after which bound anti-Pc IgM was detected with mouse anti-human IgM-HRP. Pre-incubation of plasma with competitors such as phosphorylcholine, phosphorylethanolamine, phosphorylserine or glycine-HSA, confirmed that the ELISA was specific for anti PC IgM. Levels of anti Pc IgM in a cohort of healthy donors differed by more than 100-fold, whereas the fluctuation of anti-Pc IgM levels in individuals over time was small (coefficient of variation between 6% to 25%). Furthermore, there was no correlation between CRP and anti-Pc IgM in this cohort. Levels of anti-Pc IgM in the normal donors correlated significantly with IgM binding to apoptotic cells. To test the hypothesis that anti-Pc IgM can bind to neo-antigens expressed on necrotic or apoptotic cells, anti-Pc IgM was also quantified in patients with tumors undergoing isolated limb perfusion with tumor necrosis factor-alpha (TNF-alpha). Following this procedure, a significant decrease of circulating anti-Pc IgM relative to total IgM was found in all five patients tested. In conclusion, we have developed a specific and reproducible ELISA for anti Pc IgM quantification. Fluctuation of levels of these natural antibodies over time in healthy individuals was limited, although the variation among individuals was large. Significant decreases of levels of anti-Pc IgM were found to occur during tissue damage.

Animals↗
PubMed2004

Towards functional transplant donor matching by measurement of granzyme A and granzyme B production levels.

Graft-versus-host disease (GvHD) can be a major complication after allogeneic stem cell transplantation (SCT) especially when donor and recipient are unrelated. The latter serious complication, together with the growing number of available unrelated stem cell donors, demand a simple in vitro assay for functional stem cell donor selection. Activated donor cytotoxic T lymphocytes (CTLs) and natural killer cells produce granzymes (Gr) that are involved in the pathogenesis of GvHD. We measured granzymes A and B (GrA and GrB) production levels in the supernatants of 96 h pretransplant mixed lymphocyte cultures (MLC) of 26 sibling and 31 unrelated patient/donor pairs by enzyme-linked immunosorbent assay (ELISA). In detail, the GrA and GrB production levels from a selected cohort of 37 potential patient/donor pairs were correlated with relative responses (RR) of MLC and with human leukocyte antigen (HLA) class II mismatches and with the development of acute GvHD in a second, consecutive cohort of 20 sibling SCT recipients. In vitro measurement of GrA and GrB production levels significantly correlated with the RR of pretransplant MLC (r=0.492, p< or =0.01 and r=0.853, p< or =0.01, respectively) and increased with the number of HLA class II mismatches between patient and donor. Pretransplant GrA production levels were significantly associated with the in vivo development of acute GvHD grades II-IV in patients transplanted with an HLA-identical sibling donor (p< or =0.001). In conclusion, in vitro GrA and GrB production levels can be measured by a quantitative and sensitive ELISA. This novel and simple method may be used for functional selection of unrelated stem cell donors and for the identification of patients who are at risk for acute GvHD grades II-IV.

Donor Selection↗
PubMed2004

Production, characterization, and use of serpin antibodies.

Serine protease inhibitors (serpins) constitute a still expanding superfamily of structural similar proteins, which are localized extracellularly and intracellularly. Serpins play a central role in the regulation of a wide variety of (patho) physiological processes including coagulation, fibrinolysis, inflammation, development, tumor invasion, and apoptosis. Serpins have a unique mechanism of inhibition that involves a profound change in conformational state upon interaction with their protease. This conformational change enables the production of monoclonal antibodies specific for native, complexed, and inactivated serpins. Antibodies, and assays based on these antibodies, have been helpful in elucidating the (patho) physiological function of serpins in the last decade. Serpin-specific antibodies can be used for: (1) structure-function studies such as detection of conformational changes; (2) identification of target-proteases; (3) the detection and quantification of serpin and serpin-protease complexes in bodily fluids by immunoassays such as ELISA, RIA or FACS; (4) detection of serpins in tissues by immunohistochemistry; and (5) possible therapeutical interventions. This review summarizes the techniques we have used to obtain and screen antibodies against extra- and intracellular serpins, as well as the use of these antibodies for some of the above-mentioned purposes.

Antibodies↗
PubMed2004

Complement activation by apoptotic cells occurs predominantly via IgM and is limited to late apoptotic (secondary necrotic) cells.

Apoptotic cells activate complement via various molecular mechanisms. It is not known which of these mechanisms predominate in a physiological environment. Using Jurkat cells as a model, we investigated complement deposition on vital, early and late apoptotic (secondary necrotic) cells in a physiological medium, human plasma, and established the main molecular mechanism involved in this activation. Upon incubation with recalcified plasma, binding of C3 and C4 to early apoptotic cells was similar to background binding on vital cells. In contrast, late apoptotic (secondary necrotic) cells consistently displayed substantial binding of C4 and C3 and low, but detectable, binding of C1q. Binding of C3 and C4 to the apoptotic cells was abolished by EDTA or Mg-EGTA, and also by C1-inhibitor or a monoclonal antibody that inhibits C1q binding, indicating that complement fixation by the apoptotic cells was mainly dependent on the classical pathway. Late apoptotic cells also consistently bound IgM, in which binding significantly correlated with that of C4 and C3. Depletion of plasma for IgM abolished most of the complement fixation by apoptotic cells, which was restored by supplementation with purified IgM. We conclude that complement binding by apoptotic cells in normal human plasma occurs mainly to late apoptotic, secondary necrotic cells, and that the dominant mechanism involves classical pathway activation by IgM.

Animals↗
PubMed2004

Hyperexpression of the granzyme B inhibitor PI-9 in human renal allografts: a potential mechanism for stable renal function in patients with subclinical rejection.

BACKGROUND: Granzyme B-positive T lymphocytes infiltrate renal allografts during acute cellular rejection and cause graft injury by inducing apoptosis of tubular cells. Protease inhibitor 9 (PI-9), an intracellular serpin that inhibits granzyme B, is known to protect cells from the action of cytotoxic T lymphocytes. METHODS: Expression of granzyme B and PI-9 in transplant biopsies from patients with acute cellular rejection (N= 18), subclinical rejection showing a mononuclear cell infiltrate without deterioration of renal function (N= 15), or stable transplant function (N= 13) were studied. Immunohistochemical stainings were analyzed and scored semiquantitatively by two independent observers who were not aware of clinical results. RESULTS: Granzyme B was expressed by mononuclear cells in all biopsies with cellular infiltrates. PI-9 was diffusely expressed by tubular cells in the allografts of all patients with subclinical rejection. In contrast, PI-9 expression was only focally in the patients with clinical rejection or without rejection. Although no difference was observed in granzyme B levels between acute and subclinical rejection, in subclinical rejection tubular epithelial cells showed significantly stronger expression of PI-9 than in acute rejection (P= 0.011). CONCLUSION: These data suggest that a high expression of PI-9 by tubular epithelial cells can serve as one of the factors protecting renal allografts from rejection in spite of the presence of inflammatory cell infiltrates.

Acute Disease↗
PubMed2003

Intracellular serpin SERPINB6 (PI6) is abundantly expressed by human mast cells and forms complexes with beta-tryptase monomers.

SERPINB6 (PI6) is a member of the intracellular serine protease inhibitors (serpins). Previous studies showed that SERPINB6 is localized mainly in the cytoplasm of endothelial cells, some epithelial cells, monocytes, and neutrophils. In these cells SERPINB6 is thought to prevent cellular damage by scavenging leaking lysosomal proteases. We show here, using novel, well-defined monoclonal antibodies, that SERPINB6 is abundantly expressed by mast cells in all organs and by the human mast cell line HMC-1. Gel filtration experiments revealed that the latter cells contain a high-molecular-weight form of SERPINB6, which consists of sodium dodecyl sulfate (SDS)-stable complexes of this inhibitor with monomeric beta-tryptase. Expression of SERPINB6 by mast cells was compared with those of tryptase and CD117 (c-kit) in biopsies from patients with different forms of mast cell disease. In all cases the lesional mast cells expressed SERPINB6, and, in diffuse cutaneous mastocytosis and mastocytoma, SERPINB6 was expressed by a substantially higher number of mast cells when compared with tryptase. In conclusion, SERPINB6 is abundantly expressed by normal mast cells and by mast cells in mastocytoma lesions. We suggest that in mast cells, SERPINB6 serves to regulate the activity of endogenous beta-tryptase in the cytoplasm.

Cell Line↗
PubMed2003

Type II secretory phospholipase A2 in cardiovascular disease: a mediator in atherosclerosis and ischemic damage to cardiomyocytes?

Inflammatory reactions contribute to the pathogenesis of cardiovascular conditions such as atherosclerosis and ischemic damage in acute myocardial infarction (AMI). Among the mediators involved in inflammation are secretory phospholipase A2 group II (sPLA2-II) enzymes. Though some cells constitutively express sPLA2-II, the synthesis by cells such as hepatocytes is typical for an acute-phase reactant. Recent literature suggests multiple roles for sPLA2-II in cardiovascular disease. In this review we discuss the role of sPLA2-II in various in vivo and in vitro models of atherosclerosis or AMI, including the therapeutic perspective of sPLA2-II inhibitors. It was concluded that sPLA2-II appears to be an important inflammatory mediator of cardiovascular disease.

Arteriosclerosis↗
PubMed2003

Recombinant human C1-inhibitor produced in Pichia pastoris has the same inhibitory capacity as plasma C1-inhibitor.

Therapeutic application of the serpin C1-inhibitor (C1-Inh) in inflammatory diseases like sepsis, acute myocardial infarction and vascular leakage syndrome seems promising, but large doses may be required. Therefore, a high-yield recombinant expression system for C1-Inh is very interesting. Earlier attempts to produce high levels of C1-Inh resulted in predominantly inactive C1-Inh. We describe the high yield expression of rhC1-Inh in Pichia pastoris, with 180 mg/l active C1-Inh at maximum. On average, 30 mg/l of 80-100% active C1-Inh was obtained. Progress curves were used to study the interaction with C1s, kallikrein, coagulation factor XIIa and XIa, and demonstrated that rhC1-Inh had the same inhibitory capacity as plasma C1-Inh. Structural integrity, as monitored via heat stability, was comparable despite differences in extent and nature of glycosylation. We conclude that the P. pastoris system is capable of high-level production of functionally and structurally intact human C1 inhibitor.

Cloning, Molecular↗
PubMed2003

The functional integrity of the serpin domain of C1-inhibitor depends on the unique N-terminal domain, as revealed by a pathological mutant.

C1-inhibitor (C1-Inh) is a serine protease inhibitor (serpin) with a unique, non-conserved N-terminal domain of unknown function. Genetic deficiency of C1-Inh causes hereditary angioedema. A novel type of mutation (Delta 3) in exon 3 of the C1-Inh gene, resulting in deletion of Asp62-Thr116 in this unique domain, was encountered in a hereditary angioedema pedigree. Because the domain is supposedly not essential for inhibitory activity, the unexpected loss-of-function of this deletion mutant was further investigated. The Delta 3 mutant and three additional mutants starting at Pro76, Gly98, and Ser115, lacking increasing parts of the N-terminal domain, were produced recombinantly. C1-Inh76 and C1-Inh98 retained normal conformation and interaction kinetics with target proteases. In contrast, C1-Inh115 and Delta 3, which both lack the connection between the serpin and the non-serpin domain via two disulfide bridges, were completely non-functional because of a complex-like and multimeric conformation, as demonstrated by several criteria. The Delta 3 mutant also circulated in multimeric form in plasma from affected family members. The C1-Inh mutant reported here is unique in that deletion of an entire amino acid stretch from a domain not shared by other serpins leads to a loss-of-function. The deletion in the unique N-terminal domain results in a "multimerization phenotype" of C1-Inh, because of diminished stability of the central beta-sheet. This phenotype, as well as the location of the disulfide bridges between the serpin and the non-serpin domain of C1-Inh, suggests that the function of the N-terminal region may be similar to one of the effects of heparin in antithrombin III, maintenance of the metastable serpin conformation.

Amino Acid Sequence↗
PubMed2003

Association of mannose-binding lectin genotype with cardiovascular abnormalities in Kawasaki disease.

Kawasaki disease is an acute vasculitis of possible infectious cause, which in particular affects the coronary arteries. Young children rely mostly on their innate immune system for protection against invading microorganisms, of which mannose-binding lectin is an important component. We aimed to investigate the possible role of the gene for this molecule (MBL) in white Dutch patients with Kawasaki disease. In 90 patients, frequency of mutations in the MBL gene was higher than in healthy children. In children younger than 1 year, those with mutations were at higher risk of development of coronary artery lesions than were those without (odds ratio 15.7, 95% CI 1.4-176.5, p=0.026). Our findings suggest that the innate immune system contributes differently to pathophysiology of Kawasaki disease at various ages.

Case-Control Studies↗
PubMed2003

Inhibition of coagulation, fibrinolysis, and endothelial cell activation by a p38 mitogen-activated protein kinase inhibitor during human endotoxemia.

P38 mitogen-activated protein kinase (MAPK) is an important component of intracellular signaling cascades that initiate various inflammatory cellular responses. To determine the role of p38 MAPK in the procoagulant response to lipopolysaccharide (LPS), 24 healthy subjects were exposed to an intravenous dose of LPS (4 ng/kg), preceded 3 hours earlier by orally administered 600 or 50 mg BIRB 796 BS (a specific p38 MAPK inhibitor), or placebo. The 600-mg dose of BIRB 796 BS strongly inhibited LPS-induced coagulation activation, as measured by plasma concentrations of the prothrombin fragment F1 + 2. BIRB 796 BS also dose dependently attenuated the activation and subsequent inhibition of the fibrinolytic system (plasma tissue-type plasminogen activator, plasmin-alpha2-antiplasmin complexes, and plasminogen activator inhibitor type 1) and endothelial cell activation (plasma soluble E-selectin and von Willebrand factor). Activation of p38 MAPK plays an important role in the procoagulant and endothelial cell response after in vivo exposure to LPS.

Adult↗
PubMed2003

C-reactive protein activates complement in infarcted human myocardium.

Circulating levels of C-reactive protein (CRP) constitute a cardiovascular risk marker. Immunohistochemical studies have revealed co-localization of CRP and activated complement in human infarcted myocardium suggesting CRP to enhance inflammation in ischemic myocardium by inducing local complement activation. The aim was to establish whether CRP activates complement in infarcted human myocardium and to assess the relationship between this activation and the duration of infarction. Myocardial tissue samples from 56 patients that had died from acute myocardial infarction were evaluated. Specimens were taken from infarcted as well as noninfarcted sites of the heart. CRP-mediated complement activation was assessed by immunohistochemistry and by measuring levels of complement, CRP, and CRP-complement complexes, specific markers for CRP-mediated activation, in homogenates of the heart. Infarctions of 12 hours to 5 days had significantly more extensive depositions of complement and CRP and contained significantly more CRP, activated complement, and CRP-complement complexes than infarctions that were less than 12 hours old. Levels of CRP complexes correlated significantly with CRP and complement concentrations in the infarctions, as well as with the extent of complement and CRP depositions as measured via immunohistochemistry. Specific activation products of CRP-mediated activation of complement are increased in infarcts of more than 12 hours in duration and correlate with the extent of complement depositions. Hence, CRP seems to enhance local inflammatory reactions ensuing in human myocardial infarcts of more than 12 hours duration.

C-Reactive Protein↗