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S Meri

Publications and source records attributed to S Meri.

At least 37 records · Page 2Linked to original sources
PubMed2001

Nerve growth factor and brain-derived neurotrophic factor mRNAs are regulated in distinct cell populations of rat heart after ischaemia and reperfusion.

Neurotrophins play a crucial role in the development of the peripheral nervous system and their mRNAs are often regulated after several types of tissue injury. This study has investigated the regulation of nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and neurotrophin-3 (NT-3) mRNAs 30 min after myocardial ischaemia followed by reperfusion, by northern blotting, and in situ hybridization in a rat model. Between 2 and 120 h of reperfusion, Ngf mRNA levels showed two- to four-fold up-regulation compared with sham-operated hearts. Scattered Ngf-expressing cells, probably pericytes, were detected in the viable border zone of the myocardium in close association with capillaries, venules, and arterioles. In addition, diffuse Ngf expression was seen in the infarct area after 120 h of reperfusion. Bdnf mRNA showed transient up-regulation after 2 and 5 h of reperfusion and remained at control levels thereafter. Bdnf was expressed in the myocytes of the viable border zone. Nt-3 expression showed no significant changes compared with sham-operated hearts. These results suggest a role for NGF and/or BDNF in the pathogenesis of reperfusion injury or in the alterations of cardiac sensory and sympathetic neuronal function after myocardial ischaemia and reperfusion.

Animals↗
PubMed2001

Complement-regulator factor H and related proteins in otitis media with effusion.

Otitis media with effusion (OME) is a common disease in childhood. It is characterized by chronic inflammation in which the proinflammatory activity of the complement (C) system is one of the underlying factors. The C system becomes strongly activated in the middle ear effusion (MEE) fluid, but the reasons for this are not known. Here we demonstrate by using complement Bb fragment ELISA that MEE specimens strongly activate the alternative C pathway (AP) in normal human serum (NHS). Some of the MEEs were also found to promote lysis of rabbit erythrocytes by NHS. These findings indicated a disturbance in the fluid-phase regulation of the AP in MEE. The main regulator of the AP, factor H (FH), and proteins structurally related to it (FHL-1, FHR-1, -2, -3, and -4) were present in the MEE fluids of OME patients. Relative to serum, the FHR proteins were more abundant in the MEEs. In addition, we detected the recently discovered 65-kDa FH-related protein FHR-5 in the MEE. The FHR proteins share binding sites with FH in the C3d region of C3b. Thus they may compete with FH in binding to C3b and interfere with the regulatory activity of FH. Consequently, a disturbance in AP control in the MEE may lead to an ongoing excessive C activation and inflammation in OME.

Apolipoproteins↗
PubMed2001

Proteomics: posttranslational modifications, immune responses and current analytical tools.

The publication of the human genome sequence enables most of the still unknown protein sequences to be added to the current databases. A sequence alone does not, however, give information about the possible expression level of the corresponding protein, neither does it inform about the possible posttranslational modifications, like phosphorylation, glycosylation or changes in individual amino acids. Thus, the human proteome project, a large scale analysis of the functions of gene products, will have an enormous impact on our understanding of the biochemistry of proteins, processes and pathways they are involved in. The diversity in proteins is considerably expanded by various posttranslational modifications. These also pose problems to the investigators, but their careful analysis often pays back because they can reveal important properties in proteins or peptides--like an increased antigenicity leading to (auto)immune responses or an active form of a signaling protein. Immune tolerance usually exists towards self-proteins, but in specific cases it may be broken by posttranslational modifications in the proteins. Novel mass spectrometric, affinity and display techniques offer valuable tools for the large-scale analysis of proteomes. In the present paper we discuss their use for the detection of posttranslational modifications, functional interactions and possible disease-associated abnormalities in proteins.

Electrophoresis, Gel, Two-Dimensional↗
PubMed2001

Complement C3b interactions studied with surface plasmon resonance technique.

The surface plasmon resonance (SPR) phenomenon is utilized in a number of new real time biosensors. In this study, we have used this technique to study interactions between the central complement component C3b and its multiple ligands by using the Biacore equipment. The SPR technique is particularly suitable for analysis of the alternative complement pathway (AP) because the inherent nature of the latter is to amplify deposition of C3b on various surfaces. C3b was coupled onto the sensor surface and the coupling efficiency was compared under various conditions on both polystyrene and carboxymethylated dextran surfaces. After enzymatic C3b coupling or standard amine C3b coupling, we analyzed and compared the binding of four C3b ligands to the surface: factor B, factor H, C5 and the soluble complement receptor 1 (sCR1, CD35). Binding of each ligand to C3b was detected when C3b had been coupled either enzymatically or using the amine coupling, but the half-lives of the interactions were found to vary depending on the coupling procedure. Factor H binds to C3b via three interaction sites. The target sites are exposed on the C3b, C3c and C3d fragments of C3, respectively. Therefore, we also tested by using the Biacore whether factor B, C5 and sCR1 bind to C3c and/or C3d. It was found that factor B bound to C3d, but not to C3c. On the other hand, both C5 and sCR1 bound to C3c, but not to C3d. In conclusion, this study shows that SPR is a powerful tool in analyzing and mapping the interactions of C3b with its multiple ligands.

Binding Sites↗
PubMed2001

Cytolytic complement activity in otitis media with effusion.

Otitis media with effusion (OME) is a chronic inflammation persisting in the middle ear cavity of at least 8 weeks duration. Middle ear effusion (MEE; n = 38), samples from children suffering from OME were investigated for their direct cytolytic activity or an ability to enhance complement lysis of unsensitized bystander cells. Thirteen of the 38 MEEs had direct endogenous haemolytic activity and 27 samples had an ability to enhance serum-initiated lysis. Using an enzyme immunoassay, high levels of terminal complement complexes (TCC) were detected in the MEE samples (mean 34.1 microg/ml, range 5--89 microg/ml). This indicated strong local complement activation that had progressed to the terminal stage. As one potential factor promoting complement activation we identified both monomeric and trimeric properdin in MEE by Western blotting. By stabilizing C3 and C5 convertases properdin accelerates the alternative and terminal pathways of complement. On the other hand, the membrane attack complex (MAC) inhibitor CD59, which was found to be extensively shed into the MEE in a functionally active form, may control excessive cytotoxicity of the MEE. In conclusion, intense complement activation, up to the terminal level, maintains ongoing inflammation in the middle ear cavity and can pose a threat to the local epithelium.

CD59 Antigens↗
PubMed2001

Survival of Helicobacter pylori From complement lysis by binding of GPI-anchored protectin (CD59).

BACKGROUND & AIMS: Although Helicobacter pylori is sensitive to complement lysis in vitro, chronic infection persists for years. We tested whether H. pylori acquires complement resistance by binding glycolipid-tailed inhibitors from the host. METHODS: Gastric biopsy specimens from H. pylori-infected patients (n = 10) and noninfected controls (n = 6) were analyzed for complement deposition and expression of the complement regulators protectin (CD59) and DAF. Protectin binding and complement sensitivity analyses were performed with the NCTC strain 11637 (CagA(+)) and 2 clinical isolates 9:0 (CagA(+)) and 67:20 (CagA(-)). RESULTS: In the noninfected mucosa, protectin was strongly expressed on the membranes of epithelial cells, but in the infected epithelia the expression was granular and more focused to the mucus. H. pylori bacteria in the gastric pits were often positive for protectin but negative for C5b-9. An opposite pattern was seen on the surface mucosa. In vitro analyses using (125)I-CD59 and bacteriolysis assays showed that protectin bound to H. pylori and protected CagA(+) strains against complement killing. In an enzyme-linked immunosorbent assay, the binding of CD59 correlated inversely with the appearance of the C5b-9 neoantigen. CONCLUSIONS: Binding of protectin inhibits membrane attack complex assembly on H. pylori and may thereby contribute to their survival on the gastric mucosa.

Adult↗
PubMed2001

Complement evasion by Borrelia burgdorferi: serum-resistant strains promote C3b inactivation.

The most characteristic features of the Lyme disease pathogens, the Borrelia burgdorferi sensu lato (s.l.) group, are their ability to invade tissues and to circumvent the immune defenses of the host for extended periods of time, despite elevated levels of borrelia-specific antibodies in serum and other body fluids. Our aim in the present study was to determine whether B. burgdorferi is able to interfere with complement (C) at the level of C3 by accelerating C3b inactivation and thus to inhibit the amplification of the C cascade. Strains belonging to different genospecies (Borrelia garinii, B. burgdorferi sensu stricto, and Borrelia afzelii) were compared for their sensitivities to normal human serum and abilities to promote factor I-mediated C3b degradation. B. burgdorferi sensu stricto and B. afzelii strains were found to be serum resistant. When the spirochetes were incubated with radiolabeled C3b, factor I-mediated degradation of C3b was observed in the presence of C-resistant B. afzelii (n = 3) and B. burgdorferi sensu stricto (n = 1) strains but not in the presence of C-sensitive B. garinii (n = 7) strains or control bacteria (Escherichia coli, Staphylococcus aureus, and Enterococcus faecalis). Immunoblotting and radioligand binding analyses showed that the C-resistant strains had the capacity to acquire the C inhibitors factor H and factor H-like protein 1 (FHL-1) from growth medium and human serum. A novel surface protein with an apparent molecular mass of 35 kDa was found to preferentially bind to the N terminus region of factor H. Thus, the serum-resistant B. burgdorferi s.l. strains can circumvent C attack by binding the C inhibitors factor H and FHL-1 to their surfaces and promoting factor I-mediated C3b degradation.

Animals↗
PubMed2001

Serum C3 but not plasma acylation-stimulating protein is elevated in Finnish patients with familial combined hyperlipidemia.

A trapping defect of fatty acids due to impaired function of acylation-stimulating protein (ASP) has been suggested as one mechanism underlying the metabolic abnormalities in familial combined hyperlipidemia (FCHL). The study aimed at defining the role of ASP and complement C3 in 35 Finnish FCHL families. There was no difference in plasma ASP levels between the 66 hypertriglyceridemic FCHL patients and their 84 normotriglyceridemic relatives. No response in plasma ASP could be observed after a fatty meal in 10 FCHL patients or in 10 control subjects. In familial correlation analyses, C3 exhibited a significant sibling-sibling correlation. The FCHL patients had higher serum C3 levels than their unaffected relatives (P<0.001). Furthermore, serum C3 levels correlated significantly with several lipid parameters. The correlations between ASP and lipid variables were weaker than those of C3. These analyses suggest that common genes might contribute to the regulation of serum C3, triglycerides, HDL-C, free fatty acids, and insulin. The present data do not support the hypothesis that defects of the ASP pathway are reflected in plasma lipoproteins or in impaired plasma lipid clearance postprandially.

Adult↗
PubMed2001

Complement activation in circulation and central nervous system after rituximab (anti-CD20) treatment of B-cell lymphoma.

Rituximab (IDEC-C2B8, Mabthera, Rituxan), a chimeric monoclonal antibody against the B-cell specific CD20-antigen, has been demonstrated to be effective in the treatment of non-Hodgkin's B-cell lymphoma (B-NHL). Previous in vitro studies have shown that direct complement-dependent cytotoxicity (CDC), ADCC and apoptosis are important in the rituximab-induced killing of lymphoma cells. It is, however, unknown whether rituximab penetrates the blood-brain barrier. Therefore, we studied rituximab levels and complement (C) activation in blood and cerebrospinal fluid (CSF) following intravenous rituximab therapy in a patient with relapsing non-Hodgkin's lymphoma with central nervous system (CNS) involvement. Longitudinal samples from blood and CSF were taken at 13 time-points during the treatment period. The results show that the C cascade becomes activated in blood during the first mAb infusion (C3a-desArg concentration rose from 55 to 138 microg/ml during the first 2 hours). After the first infusion the proportions of lymphocytes positive for the CD19- and CD20-antigens in the peripheral blood were reduced from 41% and 35%, respectively, to a level of 2% (for both). In CSF the rituximab concentration increased after successive infusions, but remained below 0.55 microg/ml (compared to a Cmax of 400 microg/ml in peripheral blood). Although a minor and delayed C activation response was seen in the CSF the treatment did not clear CD20-positive cells away from the CNS. Thus, it appears that an intact blood-brain barrier restricts the entry of rituximab into the CNS. Possible options to circumvent this would be dose escalation or intrathecal rituximab treatment.

Antibodies, Monoclonal↗
PubMed2000

The complement regulator factor H binds to the surface protein OspE of Borrelia burgdorferi.

Spirochete bacteria of the Borrelia burgdorferi sensu lato complex cause Lyme borreliosis. The three pathogenic subspecies Borrelia garinii, Borrelia afzelii, and Borrelia burgdorferi sensu stricto differ in their disease profiles and susceptibility to complement lysis. We investigated whether complement resistance of Borreliae could be due to acquisition of the main soluble inhibitors of the alternative complement pathway, factor H and the factor H-like protein 1. When exposed to nonimmune EDTA-plasma, the serum-resistant B. afzelii and B. burgdorferi sensu stricto strains bound factor H/factor H-like protein 1 to their surfaces. Assays with radiolabeled proteins showed that factor H bound strongly to the B. burgdorferi sensu stricto strain. To identify factor H ligands on the borrelial surface, we analyzed a panel of outer surface proteins of B. burgdorferi sensu stricto with the surface plasmon resonance technique. The outer surface lipoprotein OspE was identified as a specific ligand for factor H. Using recombinant constructs of factor H, the binding site for OspE was localized to the C-terminal short consensus repeat domains 15-20. Specific binding of factor H to B. burgdorferi sensu stricto OspE may help the pathogen to evade complement attack and phagocytosis.

Antigens, Bacterial↗
PubMed2000

Deficiency of complement defense protein CD59 may contribute to neurodegeneration in Alzheimer's disease.

Complement defense 59 (CD59) is a cell surface glycophosphoinositol (GPI)-anchored protein that prevents complement membrane attack complex (MAC) assembly. Here, we present evidence from ELISA assays that CD59 protein levels are significantly decreased in the frontal cortex and hippocampus of Alzheimer's disease (AD) compared with nondemented elderly (ND) patients, whereas complement component 9, a final component to form MAC, is significantly increased. To further confirm the CD59 deficit, PI-specific phospholipase C (PIPLC) was used to cleave the CD59 GPI anchor at the cell surface in intact slices from AD and ND cortex. CD59 released by PIPLC cleavage was significantly reduced in AD compared with ND samples. By the use of a ribonuclease protection technique, amyloid beta-peptide was found to downregulate CD59 expression at the mRNA level, suggesting a partial explanation of CD59 deficits in the AD brain. To evaluate the pathophysiological significance of CD59 alterations in neurons, we exposed cultured NT2 cells, which normally underexpress CD59, and NT2 cells transfected to overexpress CD59 to homologous human serum. Lactic acid dehydrogenase assays revealed significant complement-induced cell lysis in CD59-underexpressing NT2 cells and significant protection from such lysis in CD59-overexpressing NT2 cells. Moreover, cells expressing normal levels of CD59 showed no evidence of MAC assembly or damage after exposure to homologous serum, whereas pretreatment of these cells with a CD59-neutralizing antibody resulted in MAC assembly at the cell surface and morphological damage. Taken together, these data suggest that CD59 deficits may play a role in the neuritic losses characteristic of AD.

Aged↗
PubMed2000

Each of the three binding sites on complement factor H interacts with a distinct site on C3b.

Factor H (fH) restricts activation of the alternative pathway of complement at the level of C3, both in the fluid phase and on self-structures, but allows the activation to proceed on foreign structures. To study the interactions between fH and C3b we used surface plasmon resonance analysis (Biacore(R)) and eight recombinantly expressed fH constructs containing fragments of the 20 short consensus repeat domains (SCRs) of fH. We analyzed the binding of these constructs to C3b and its cleavage products C3c and C3d. Three binding sites for C3b were found on fH. Site 1 was localized to the five amino-terminal SCRs (SCR1-5), and its reciprocal binding site on C3b was found to be lost upon the cleavage of C3b to C3c and C3d. Site 2 on fH was localized by exclusion probably within or near SCRs 12-14 (fragment SCR8-20 bound to C3b, C3c, and C3d; SCR8-11 did not bind to C3b at all; and SCR15-20 bound only to the C3d part of C3b). Site 3 on fH for C3b was localized to the carboxyl-terminal SCRs 19-20, and its reciprocal binding site was mapped to the C3d part of C3b. In conclusion, we confirmed and mapped three binding sites on fH for C3b and demonstrated that the three binding sites on fH interact with distinct sites on C3b. Multiple reciprocal interactions between C3b and fH can provide a basis for the different reactivity of the alternative pathway with different target structures.

Binding Sites↗
PubMed2000

Cellular and humoral immune responses in atrophic lymphocytic pancreatitis in German shepherd dogs and rough-coated collies.

The most common cause for the clinical signs of exocrine pancreatic insufficiency (EPI) in dogs is pancreatic acinar atrophy (PAA). In the subclinical phase of EPI, before total atrophy occurs, exocrine pancreas is affected by infiltrative lymphocytic inflammation, which gradually leads to selective destruction and atrophy of the acinar tissue.Here, we analyzed the role of cell-mediated and humoral immune mechanisms in the pathogenesis of atrophic lymphocytic pancreatitis in German shepherd dogs and rough-coated collies. Pancreas biopsies and serum samples were obtained from 12 dogs with subclinical EPI (SEPI), 13 dogs with clinical EPI and 13 healthy control dogs. Immunohistochemical analysis showed that, in the subclinical phase, the majority of the infiltrating lymphocytes were T-cells with an almost equal number of CD4+ 'T-helper' and CD8+ 'cytotoxic' T-lymphocytes. The distribution of the two lymphocyte subsets was different. Typically, the CD4+ cells were present in large cellular infiltrates in the affected parenchyma, and the scattered CD8+ cells had infiltrated both the affected and the normal parenchyma. In sections where destruction of acinar parenchyma was present, the CD8+ T-cells were predominant. In cases of marked T-cell infiltration, CD79+ B-lymphocytes and plasma cells, and lysozyme-positive macrophages were also detected. Lymphoid follicle germinal centers with a majority of cells staining positively for CD79 were found. The lymphocytic infiltration in the totally atrophic tissue of dogs with clinical EPI was less prominent. Indirect immunofluorescence staining showed serum antibodies reacting weakly with pancreatic acinar cells in five out of nine dogs with subclinical and three out of 10 dogs with clinical EPI, but not in the control dogs. The results suggest that the tissue destruction is largely T-cell-mediated, although the presence of numerous B-lymphocytes and pancreas-specific antibodies in the sera of some dogs indicate that humoral mechanisms are also involved. In conclusion, this study suggests that the atrophic lymphocytic pancreatitis in German shepherds and rough-coated collies is an autoimmune disease.

Animals↗
PubMed2000

Exceptional resistance of human H2 glioblastoma cells to complement-mediated killing by expression and utilization of factor H and factor H-like protein 1.

Of over 20 nucleated cell lines we have examined to date, human H2 glioblastoma cells have turned out to be the most resistant to complement-mediated cytolysis in vitro. H2 cells expressed strongly the membrane attack complex inhibitor protectin (CD59), moderately CD46 (membrane cofactor protein) and CD55 (decay-accelerating factor), but no CD35 (complement receptor 1). When treated with a polyclonal anti-H2 Ab, anti-CD59 mAb, and normal human serum, only 5% of H2 cells became killed. Under the same conditions, 70% of endothelial-like EA.hy 926 cells and 40% of U251 control glioma cells were killed. A combined neutralization of CD46, CD55, and CD59 increased H2 lysis only minimally, demonstrating that these complement regulators are not enough to account for the resistance of H2 cells. After treatment with Abs and serum, less C5b-9 was deposited on H2 than on U251 and EA.hy 926 cell lines. A reason for the exceptional resistance of H2 cells was revealed when RT-PCR and protein biochemical methods showed that the H2 cells, unlike the other cell lines tested, actively produced the soluble complement inhibitors factor H and factor H-like protein 1. H2 cells were also capable of binding human factor H from the fluid phase to their cell surface and promoted the cleavage of C3b to its inactive form iC3b more efficiently than U251 and EA.hy 926 cells. In accordance, anti-factor H mAbs enhanced killing of H2 glioblastoma cells. Taken together, our results show that production and binding of factor H and factor H-like protein 1 is a novel mechanism that these malignant cells utilize to escape complement-mediated killing.

Antibodies, Monoclonal↗
PubMed2000

Molecular mechanisms of target recognition in an innate immune system: interactions among factor H, C3b, and target in the alternative pathway of human complement.

In the alternative pathway of complement (APC) factor H is the primary control factor involved in discrimination between potential pathogens. The APC deposits C3b on possible Ags, and the interaction with factor H determines whether the initial C3b activates the APC. Factor H is composed of a linear array of 20 homologous short consensus repeats (SCR) domains with many functional sites. Three of these sites are involved in binding C3b and regulating complement activation; others bind to sialic acid and/or heparin and are responsible for host recognition. Using site-directed mutations we have examined the contributions of each of these sites to target discrimination and to functional activities of factor H. Decay acceleration by SCR1-4 of C3/C5 convertases bound to nonactivators was strongly dependent on SCR domains 11-15 and 16-20. Loss of these regions caused a 97% loss of activity, with SCR16-20 being the most critical (>90% loss). On APC activators the pattern of site usage was different and unique on each. On yeast, deletion of the 10 C-terminal domains (SCR11-20) had no effect on specific activity. On rabbit erythrocytes, this deletion caused loss of 75% of the specific activity. An examination of binding affinity to C3b on the four cell types demonstrated that factor H exhibits a unique pattern of SCR involvement on each cell. The results reveal a complex molecular mechanism of discrimination between microbes and host in this ancient innate defense system and help explain the different rates and intensities of APC activation on different biological particles.

Animals↗
PubMed2000

Complement activation after oxidative stress: role of the lectin complement pathway.

The complement system plays an important role in mediating tissue injury after oxidative stress. The role of mannose-binding lectin (MBL) and the lectin complement pathway (LCP) in mediating complement activation after endothelial oxidative stress was investigated. iC3b deposition on hypoxic (24 hours; 1% O(2))/reoxygenated (3 hours; 21% O(2)) human endothelial cells was attenuated by N-acetyl-D-glucosamine or D-mannose, but not L-mannose, in a dose-dependent manner. Endothelial iC3b deposition after oxidative stress was also attenuated in MBL-deficient serum. Novel, functionally inhibitory, anti-human MBL monoclonal antibodies attenuated MBL-dependent C3 deposition on mannan-coated plates in a dose-dependent manner. Treatment of human serum with anti-MBL monoclonal antibodies inhibited MBL and C3 deposition after endothelial oxidative stress. Consistent with our in vitro findings, C3 and MBL immunostaining throughout the ischemic area at risk increased during rat myocardial reperfusion in vivo. These data suggest that the LCP mediates complement activation after tissue oxidative stress. Inhibition of MBL may represent a novel therapeutic strategy for ischemia/reperfusion injury and other complement-mediated disease states.

Acetylglucosamine↗
PubMed2000

Different regulation of factor H and FHL-1/reconectin by inflammatory mediators and expression of the two proteins in rheumatoid arthritis (RA).

Factor H and the FHL-1/reconectin protein are two human plasma proteins that act as important regulators of the alternative complement pathway. Each protein is encoded by a unique transcript, but both mRNAs are derived from the factor H gene by means of alternative processing. In order to address potential functional differences between the two proteins we analysed their expression in hepatic and non-hepatic cells and studied their regulation by inflammatory mediators. We demonstrate that factor H and FHL-1/reconectin transcripts which are regulated by the same gene promoter and are initiated at the same transcription start site are differently expressed. Expression of the molecules is induced and regulated by the inflammatory mediators interferon-gamma (IFN-gamma) and the anti-inflammatory glucocorticoid dexamethasone. Both factor H and FHL-1/reconectin are expressed and secreted by synovial fibroblasts and are present in synovial fluid derived from patients suffering from rheumatoid or reactive arthritis. The local synthesis in synovial fibroblasts and their induction by IFN-gamma and dexamethasone, but not by tumour necrosis factor-alpha, suggests for each of the two complement regulators a protective role in RA.

Alternative Splicing↗