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PubMed1991

IgA-mediated inhibition of human leucocyte function by interference with Fc gamma and C3b receptors.

The inhibitory effects of IgA from human colostrum, and IgA1 and IgA2 from human serum on the chemiluminescence (CL) response and phagocytosis of polymorphonuclear leucocytes (PML) to Staphylococcus epidermidis and the CL response to formylmethionyl-leucyl-phenylalanine (FMLP) were studied. The dose-dependent inhibition of the luminol-mediated CL response of human PML to the bacteria was observed in the presence of more than 0.1 mg/ml IgA from both colostrum and serum. The preincubation of PML with a solution of IgA enhanced the suppressive effect of IgA on the cells. Removal of IgA from the reaction mixture after preincubation resulted in recovery, with time, of the response of PML to the bacteria. The bacteria treated with IgA did not give rise to any inhibition of the response. The CL response of PML to FMLP was not affected by the presence of IgA in the reaction mixture. The decrease of phagocytic activity of PML in the presence of IgA resulted in a decrease of NADPH oxidase activity of PML after stimulation with the bacteria as compared with the absence of IgA. The effect of IgA on the receptors of Fc and C3b (CR1) on the surface of PML was measured by monitoring erythrocyte-antibody (EA) or erythrocyte-antibody-complement (EAC) rosette formation and by direct and indirect immunofluorescence techniques using anti-CR1 antibody and Fc-specific antibodies. The presence of IgA in the reaction mixture led to a quantitative decrease in CR1 and the ability to bind IgG to the surface of PML.

Antigens, Differentiation↗
PubMed1991

Participation of protein kinases in complement C5b-9-induced shedding of platelet plasma membrane vesicles.

The formation of membrane microparticles through vesiculation of the platelet plasma membrane is known to provide catalytic surface for several enzyme complexes of the coagulation system, and to underlie the procoagulant responses elicited with platelet activation. This induced shedding of vesicles from the plasma membrane is most prominent when platelets are activated by the terminal complement proteins, C5b-9, by a Ca2+ ionophore, or by the combination of thrombin plus collagen. Although shown to require elevated [Ca2+], the cellular events that initiate plasma membrane evagination and fusion to form the shed vesicles remain unresolved. To gain additional insight into the cellular events that regulate membrane microparticle formation, we have examined how this process is influenced by the activity of cellular protein kinases. Cytoplasmic [Ca2+] of gel-filtered platelets was increased by membrane assembly of the terminal complement proteins C5b-9 in the presence of selective inhibitors of protein kinase or phosphatase reactions, and resulting microparticle formation was quantitated by fluorescence-gated flow cytometry. Pre-equilibration of the phosphatase inhibitor vanadate into the platelet cytosol increased microparticle formation by as much as 40%, suggesting that vesiculation of the platelet plasma membrane is influenced by the state of phosphorylation of a cellular constituent. By contrast to the stimulatory effects of vanadate, microparticle formation was partially inhibited in platelets treated with the protein kinase inhibitor sphingosine, the myosin light chain kinase inhibitor ML-7, the calmodulin-antagonist W-7, and under conditions of elevated cytosolic concentration of cyclic adenosine monophosphate. These results indicate that complement-induced platelet microparticle formation is influenced by one or more protein kinase(s) as well as by calmodulin, and suggest a role for the platelet myosin light chain kinase or another Ca(2+)-pluscalmodulin-regulated membrane component.

Blood Platelets↗
PubMed1990

Inhibition of leukocyte chemotaxis by serum factor in diabetes mellitus: selective depression of cell responses mediated by complement-derived chemoattractants.

Rat neutrophil chemotactic responses to N-formyl-methionyl-leucyl-phenylalanine (FMLP), leukotriene (LT) B4, and lipopolysaccharide-activated serum (LPS-AS) were quantitatively assessed using the micropore filter system. Cells were suspended in either normal or diabetic rat serum for testing. Diabetic donor serum did not affect migration of neutrophils in a concentration gradient of the synthetic chemotactic agents. In contrast, the migratory responses to LPS-AS were significantly less than normal in this circumstance. Summation of effects was observed when FMLP and LPS-AS, or LTB4 and LPS-AS were simultaneously added to the test chamber, with cells suspended in normal serum. Suspended in diabetic rat serum neutrophils responded normally to the synthetic chemoattractants but the response to the activated serum was blocked. Cells previously incubated in the presence of diabetic donor serum then transferred to a culture medium for testing, presented reduced migratory responses to LPS-AS. Supramaximal, inhibitory concentrations of FMLP and LTB4, did not influence the response of neutrophils to LPS-AS. In vivo, suppression of cellular emigration to an inflamed area was observed from the early stages of the diabetic state. The inhibitory activity of chemotaxis in diabetes mellitus was previously reported to be associated with a protein factor in plasma of the animals. It is suggested that the inhibitory factor of chemotaxis in diabetes mellitus interacts with neutrophil receptors for complement-derived chemoattractants to induce blockade of cell-oriented locomotion either in vitro or in vivo.

Amino Acid Sequence↗
PubMed1981

Mechanism of the serum defect in yeast opsonization in children with fulminant hepatic failure (FHF).

Defective opsonization of heat-killed baker's yeast was found in all 14 children with fulminant hepatic failure (FHF) but returned to normal in two who recovered and was normal in six mothers of patients. Yeast opsonization was significantly correlated with factor B activity of FHF serum but not with other components of classical or alternative pathways of complement. Reconstitution and family studies suggested that defective yeast opsonization in FHF is secondary and, although dependent on factor B activity, is qualitatively and quantitatively similar to that of primary yeast opsonization deficiency. These findings suggest that the factors responsible for opsonization of yeasts, or their regulation, are synthesized or controlled by the liver.

Child↗
PubMed2000

The revision of the air quality legislation in the european union related to ground-level ozone.

Complying with the obligation in the current ozone directive, the European Commission came forward in 1999 with a strategy to combat tropospheric ozone together with a proposed revision of the air quality legislation for this pollutant. As a daughter legislation under the 1996 Framework Directive on Air Quality, the proposed ozone daughter directive defines for the first time (interim) air quality targets for ozone to be attained by 2010, complemented by long-term objectives for ozone based on the guideline values of the World Health Organisation. It also sets out enhanced requirements for monitoring and assessment of ozone concentrations, as well as minimum criteria for appropriate information of the public about the measured air pollution. In the past, abatement strategies against air pollution consisted of concrete obligations for controlling emissions derived solely on the basis of technical and economic aspects, covering specific types of installations or activities, thus with no direct quantitative relationship to the level of air pollution let alone to its effects. In compensating this deficit, the Commission presented, as a complement to the existing sectoral legislation, a proposal for a directive on national emission ceilings (NEC) which quantifies emission targets for every Member State to bring its total precursor emissions by 2010 down to levels being considered as necessary to achieve everywhere on a regional scale the air quality targets set in the ozone daughter directive. As the core element of the ozone abatement strategy, the national ceilings for emissions of sulfur dioxide (SO(2)), nitrogen oxides (NO(x)), ammonia (NH(3)) and volatile organic compounds (VOC) were derived from a cost-effectiveness analysis integrating information on economic, technical, physical and biological aspects of ozone pollution and abatement. This integrated assessment considers the potential and costs for further emission control in the various economic sectors in the Member States and combines this with information on ozone formation and transport processes in the atmosphere and with indicators for the impact of ozone on human health and environmental. Reflecting the discussions with Member States and stakeholders, a number of decisive steps in the process of deriving the national emission ceilings are presented here: the way of framing interim objectives, how to choose an appropriate ambition level, aspects of how to cope with uncertainties in the model and the input data and how to treat extreme meteorological situations and resolve problems in the spatial distribution of the interim objectives given the different size of countries. Finally, the paper explains the scenario underpinning the proposed national emission ceilings, its environmental gains and the distribution of cost incurring for emission control measures in Member States.

Air Pollution↗
PubMed1978

The measurement of complement as an aid in diagnosis: 1. Nephritis and collagen disease.

A selection of patients with nephritis and collagen diseases were studied for complement abnormalities. This communication briefly reviews the current comcepts of complement physiology, details standard assay used in this laboratory and describes clinical examples where these tests have provided significant clinical information. Quantitative measurements of individual components, the presence of circulating immune complexes, and the demonstration of in vivo C3 cleavage products were assessed simultaneously. The importance of measuring cleavage products and immune complexes is stressed. Clinical examples are given where misleading or erroneous interpretation of results might have occurred if these tests had not been performed.

Adolescent↗
PubMed1976

[Possible role of immune-phenomena in the pathogenesis of rheumatoid arthritis (author's transl)].

The R.A. is accompanied by a variety of different immune-phenomena. These are not restricted to the well known and diagnostically exploited IgM-rheumatoid factors in serum, but involve also rheumatoid factors of the Ig-classes G and A, which, at least partially are produced by lymphoid cells, which infiltrate the synovial membranes. These cellular infiltrations consist of macrophages, B and T lymphocytes, the latter being in excess. There is now good evidence that locally deposited, complement activating immune-complexes participate in the inflammatory lesions of the R.A.-joints by inducing the release of hydrolases from phagocytic cells. This event is also mediated by lymphokines and complement factors. Is is suggested that the pathogenicity of the immunoreactions occuring within the synovial tissue and fluid is a quantitative rather than a qualitative problem. In contrast to other tissues, the anatomy of the joint may favour the local accumulation of the immune-complexes. Moreover abundant immuno-reactions may be provided by a defective T-cell mediated control. Thus there is strong indication that immune-phenomena play an important role in the pathogenesis of R.A. This does not exclude the significance of metaboltic disturbances of the connective tissues for this disease.

Antigen-Antibody Complex↗
PubMed1977

F-bodies as Y chromosome markers in mature human sperm heads: a quantitative approach.

It is generally accepted that at least some Y chromosomes in human mature sperm heads are represented in preparations by fluorescent F-bodies. From data consisting solely of F-body scorings and the visually estimated size of sperm heads from normal donors, a quantitative model is developed which permits simultaneous estimation of six parameters representing biological factors (the proportions of ploidy classes and of their sex-chromosome complements) and biassing factors (not all Y chromosomes represented by F-bodies, not all F-bodies representing Y chromosomes, and error in scoring ploidy from visual size). The analysis is statistically adequate and succeeds in predicting a variety of independently validated phenomena. The work may be regarded as a contribution to the integration of qualitative and quantitative evidence in the study of F-bodies. In all, 21,700 one-headed spermatozoa from 12 donors were scored. A mean of 54% spermatozoa lacking F-bodies was subject to a standard deviation between donors of 4.8, over and above binomial error. Analysis yielded the following estimates: 83.3% of diploid spermatozoa are XY, the remainder XX and YY in assumed equal number (this information assists analysis of the origin of embryonic triploidy); 2.4% of spermatozoa are diploid (this estimate does not compete in accuracy with direct estimates from DNA absorbance); only 83% of Y chromosomes are represented by F-bodies; 7% of haploid heads and 14% of diploids contain one or more "adventitious bodies" indistinguishable from true F-bodies. The visually scored head sizes "Small" and "Medium" correspond approximately with haploidy and diploidy; nearly all haploids are scored as Small, and 72% of the Medium are diploid. F-bodies are not thought at present to give a useful basis for estimating the presumably low incidence of sex-chromosome aneuploidy and nondisjunction. No such estimate has been attempted. The problem is overshadowed by not fully understood complicating factors of greater magnitude, which now appear to be a major subject of enquiry. Some implications and possible future developments are discussed.

Humans↗
PubMedDate not supplied

[Detection of IgG and IgM antibodies to herpes simplex virus type 1 and type 2 in human serum using an immunoenzyme method (ELISA) and the complement fixation reaction].

On the basis of borderline optical density values (OD) in an enzyme-linked immunosorbent assay (ELISA) resulting from comparative testing of this method with complement fixation reactions (CF), determination was made of a set criteria for negative, weakly positive and positive results of IgG and IgM antibody detection of Herpes simplex type 1 (HSV 1), respectively type 2 (HSV 2). Of the 192 examined specimens by ELISA, 69.2% and 49.5% IgG HSV 1 and HSV 2 antibodies were found respectively. The number of sera containing IgM HSV 1 od HSV 2 antibodies was low (5.7%), since in the majority of cases the relapsing form of infection was involved. The sensitivity of ELISA to IgG in comparison with CF was slightly higher with the HSV 1 antigen (69.2:68.2%), and apparently lower with HSV 2 (49.5:28.2%). ELISA permits partial differentiation to be made between HSV 1 and HSV 2 antibodies due to a greater degree of antigen purity compared with the unpurified antigen used in CF. In fact, CF is known not to possess the sensitivity necessary to differentiate between HSV 1 and HSV 2 antibodies. For diagnostic purposes, ELISA findings could be more appropriately expressed quantitatively, in corresponding measurement units.

Antibodies, Viral↗
PubMed1992

Demineralization and remineralization evaluation techniques--added considerations.

Methods used for the analysis of tooth de- and remineralization include techniques with various degrees of sophistication and quantitative capabilities, ranging from direct measures of mineral gain/loss (e.g., microradiography) to indirect measures (e.g., iodide permeability) of changes in tooth mineral properties. In all instances, the capabilities of methods for accurate determination of changes in tooth mineral properties are affected by procedures used in the preparation of specimens for analysis, the magnitude of change taking place in the test (vs. the detection limits of the techniques), and protocols for specimen analysis. In specific instances, such as in the case of dentin, unique specimen-handling and analysis procedures must be used to prevent artifacts. The choice of techniques for the assessment of de- and remineralization depends strongly upon study protocols and laboratory capabilities; however, 'quantitative' measures of mineral gain and loss are possible only if direct chemical or radiographic techniques are used. Either radiographic, cross-sectioned microhardness or polarized light can be used for the determination of lesion depth. Porosity, light-scattering, and surface microhardness are indirect techniques which complement direct measures of mineral gain and loss. Whatever methods are used in the analysis of de- and remineralization, researchers must take care to differentiate accurately among the quantitative capabilities of techniques used in analysis.

Dental Enamel↗
PubMed2001

The genetic architecture of quantitative traits.

Phenotypic variation for quantitative traits results from the segregation of alleles at multiple quantitative trait loci (QTL) with effects that are sensitive to the genetic, sexual, and external environments. Major challenges for biology in the post-genome era are to map the molecular polymorphisms responsible for variation in medically, agriculturally, and evolutionarily important complex traits; and to determine their gene frequencies and their homozygous, heterozygous, epistatic, and pleiotropic effects in multiple environments. The ease with which QTL can be mapped to genomic intervals bounded by molecular markers belies the difficulty in matching the QTL to a genetic locus. The latter requires high-resolution recombination or linkage disequilibrium mapping to nominate putative candidate genes, followed by genetic and/or functional complementation and gene expression analyses. Complete genome sequences and improved technologies for polymorphism detection will greatly advance the genetic dissection of quantitative traits in model organisms, which will open avenues for exploration of homologous QTL in related taxa.

Animals↗
PubMed2004

Possible role of factor XIII subunit A in Fcgamma and complement receptor-mediated phagocytosis.

Besides its traditional role in hemostasis, factor XIII subunit A (FXIII-A) is supposed to function as a cellular transglutaminase and to be involved in certain intracellular processes, including cytoskeletal remodeling. To investigate its intracellular role, the aim of the present study was to follow changes in FXIII-A production in combination with the receptor-mediated phagocytic activities of monocytes/macrophages and to examine the phagocytic functions of monocytes in patients with FXIII-A deficiency. Human blood monocytes were isolated from the buffy coats of healthy volunteers and cultured for 4 days. The FcgammaR-mediated phagocytosis of sensitized erythrocytes (EA) and the complement receptor (CR)-mediated phagocytosis of complement-coated yeast particles were studied during monocyte/macrophage differentiation. Changes in the gene expression of FXIII-A were detected by real-time quantitative RT-PCR. FXIII-A protein production was investigated with fluorescent image analysis at single cell level and Western immunoblot analysis. Both the FcgammaR and CR-mediated phagocytosis increased during culturing, which peaked on day 3. The phagocytic activity of the cells could be markedly inhibited with monodansylcadaverine, an inhibitor of the transglutaminase-induced crosslinking of proteins. The phagocytosis of EA, complement-coated and uncoated yeast particles was found to be strongly diminished in monocytes of FXIII-A deficient patients. The phagocytic functions of cultured cells showed a change in parallel with the alterations in FXIII-A mRNA expression, as well as with that in FXIII-A in protein synthesis detected by image and Western immunoblot analyses in concert. Our results suggest that FXIII-A plays a role in the Fcgamma and complement receptor-mediated phagocytic activities of monocytes/macrophages.

Blotting, Western↗
PubMed2005

Carp (Cyprinus carpio L.) innate immune factors are present before hatching.

Expression of the innate immune factors, complement factor 3 (C3), alpha2-macroglobulin (alpha2M), serum amyloid A (SAA) and a complement factor 1 r/s--mannose binding lectin associated serine protease-like molecule (C1/MASP2), was determined with Real Time Quantitative-PCR in carp (Cyprinus carpio L.) ontogeny around hatching. Furthermore, the expression of C3 mRNA and the presence of C3 protein were studied in carp embryos and larvae using In Situ Hybridisation, Western Blotting and Immunohistochemistry. C3, alpha2M, SAA and C1/MASP2 mRNA were produced by embryos from 12 h post-fertilisation, which is relatively long before hatching (2 days post-fertilisation (dpf)), indicating either involvement of these factors in development itself or more probably a preparation of the immune system for the post-hatching period. In addition, maternal mRNA of the aforementioned innate immune factors and maternal C3- and immunoglobulin protein was present in unfertilised eggs. Furthermore, C3 mRNA production was situated in the yolk syncytial layer in embryos from 24 h post-fertilisation to 5 dpf, followed by the liver in larvae, providing a new sequence of C3 production in teleost development.

Animals↗
PubMed1975

Change in the proportion of T and B lymphocytes in human malignant neoplasia in relation to the clinical stage.

Formation of spontaneous (E--erythrocyte) and immune (EAC--erythrocyte, antibody, complement) rosettes of sheep red blood cells (SRBC) with peripheral blood T and B lymphocytes, respectively, was used for quantitative assessment of these lymphocytes populations in tumor patients and control subjects. Relative counts of T and B lymphocytes have been correlated with lymphocyte survival in short-term cultures using standard subtoxic doses of phytohemagglutinin (PHA). The mean values and SE in normal control subjects for E rosettes (T lymphocytes) were 72.8 +/- 1.2%, for EAC rosettes (B lymphocytes) 23.8 +/- 2.5% of lymphocyte population. The survival of lymphocytes in short-term cultures with PHA in control subjects was 55.3 +/- 1.1%. In tumor patients the counts of E, EAC rosettes and percentage of lymphocyte survival in short-term PHA cultures were dependent upon the clinical stage. In patients with localized tumors the values were similar to controls. In patients where dissemination of the tumorous process could be ascertained a decrease of T lymphocyte counts, an increase of B lymphocyte counts and a higher survival of peripheral blood lymphocytes in short-term cultures with PHA was found.

Animals↗
PubMed2004

Zooming in on a quantitative trait for tomato yield using interspecific introgressions.

To explore natural biodiversity we developed and examined introgression lines (ILs) containing chromosome segments of wild species (Solanum pennellii) in the background of the cultivated tomato (S. lycopersicum). We identified Brix9-2-5, which is a S. pennellii quantitative trait locus (QTL) that increases sugar yield of tomatoes and was mapped within a flower- and fruit-specific invertase (LIN5). QTL analysis representing five different tomato species delimited the functional polymorphism of Brix9-2-5 to an amino acid near the catalytic site of the invertase crystal, affecting enzyme kinetics and fruit sink strength. These results underline the power of diverse ILs for high-resolution perspectives on complex phenotypes.

Alleles↗
PubMed1988

Synovial fibroblast-like cells synthesize seven proteins of the complement system.

Fibroblast-like cells from synovial tissue obtained during arthroscopy in 4 young adults with recent knee trauma were biosynthetically labeled with 35S-methionine, and protein production was quantitated by immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Synovial fibroblast-like cells synthesized C1r, C1s, C1 inhibitor, C2, C3, factor B, and factor H, all with the same sizes and subunit structures as the proteins synthesized in skin fibroblasts. The capacity to synthesize these proteins was not lost with passages or freeze-thawing. Gamma-interferon stimulation increased synthesis of all 7 proteins. Lipopolysaccharide increased synthesis of only C3 and factor B. Unlike in whole rheumatoid tissue, C4 and C5 were not detected. Synovial lining cells may be an important source of local complement for participation in local defense or development of pathologic states.

Cell Adhesion↗
PubMed1988

Interactions of virulent and avirulent Legionella pneumophila with human polymorphonuclear leukocytes.

Isogenic virulent/avirulent variants of Legionella pneumophila (Lp) were examined in vitro for their effect on PMN function as determined by measurement of peak chemiluminescence and superoxide anion production. When exposed to virulent/avirulent strains of Lp, PMN function was less than that seen following exposure to E. coli, the reduction of the oxidative burst being more pronounced with virulent Lp strains in both assays. To determine whether differences in reduced PMN function correlate with differences in complement component C3 binding by virulent and avirulent Lp, four pairs of organisms were incubated in normal human serum and examined by quantitative immunofluorescence. All strains of Lp bound less C3 than E. coli, although each to different degrees. Avirulent isogenic strains bound more C3 than their virulent counterparts with one exception. Depressed PMN function following exposure to Lp may be due to reduced C3 binding, although other virulence-associated factors may be involved.

Complement C3↗
PubMed1975

Tumor-specific and forssman antigens of guinea-pig hepatoma cells: comparison of tumor cells grown in vivo and in vitro.

Cloned tumor-cell lines were derived from two antigenically distinct ascites variants of diethylnitrosamine-induced guinea-pig hepatomas (designated line 1 and line 10). Cell-surface antigens on the ascites and in vitro-grown tumor cells were analyzed with immunofluorescence, C1 fixation and transfer, and antibody-complement-mediated cytotoxity tests. Tumor-specific and Forssman antigens continued to be expressed during 3-6 months in vitro cultivation. Differences between ascites and cultured cells were noted in the degree of antigen expression and sensitivity to antibody-complement-mediated cytotoxicity. Cells grown in vitro exhibited a greater number of Forssman and tumor-specific antigen sites than in vivo grown cells as determined by the quantitative C1 fixation and transfer test. Immunofluorescent staining indicated that some cloned lines were considerably more homogeneous in terms of antigen expression than were the cultured, non-cloned parent cells or ascites-grown cells. Cloned lines were frequently more sensitive to the cytotoxic action of antibody and complement than were the in vivo grown and cultured non-cloned parent tumor cells. Sensitivity to cytotoxicity did not necessarily correlate, however, with the degree of antigen expression. These results suggest that; (1) the expression of Forssman and tumor-specific antigens does not diminish on cells cultivated in vitro and (2) ascites hepatoma cells in vivo are a heterogeneous population of cells differing in their degree of antigen expression and sensitivity to antibody-complement-mediated cytotoxicity.

Animals↗