scholarly journals High doses of intravenous immunoglobulin do not affect the recognition phase of the classical complement pathway

Blood ◽  
1991 ◽  
Vol 78 (3) ◽  
pp. 700-702 ◽  
Author(s):  
M Basta ◽  
LF Fries ◽  
MM Frank
Keyword(s):  
Human Serum ◽  
Intravenous Immunoglobulin ◽  
Normal Serum ◽  
Normal Human Serum ◽  
Complement Pathway ◽  
Classical Complement Pathway ◽  
Recognition Phase ◽  
Normal Human ◽  
Classical Complement

Abstract We have recently found that intravenous immunoglobulin (IVIg) prevents deposition of C3 and C4 fragments onto antibody sensitized erythrocytes. To find out if such an effect results from the blockade of the recognition phase of the classical complement cascade, we investigated the ability of human serum containing high concentrations of IVIg to deposit the recognition subunit of the first complement component (C1q) onto targets. Normal human serum supplemented in vitro with IVIg did not demonstrate reduced C1q binding to targets as determined by radiolabeled antihuman C1q antibody uptake. Similarly, methylamine-treated normal human serum to which IVIg was added was equally effective in terms of C1q binding as the same serum without IVIg. At increasing doses of sensitizing antibody, C1q uptake decreased proportionally; however, at all antibody dilution points C1q uptake was not significantly different in the serum with IVIg in comparison with normal serum. Serum from a patient treated with IVIg did not differ in its capacity to deposit C1q from the same patient's serum before therapy. Our data suggest that IVIg does not interfere with the recognition step of classical complement pathway. This is a US government work. There are no restrictions on its use.

scholarly journals High doses of intravenous immunoglobulin do not affect the recognition phase of the classical complement pathway

Blood ◽  
1991 ◽  
Vol 78 (3) ◽  
pp. 700-702
Author(s):  
M Basta ◽  
LF Fries ◽  
MM Frank
Keyword(s):  
Human Serum ◽  
Intravenous Immunoglobulin ◽  
Normal Serum ◽  
Normal Human Serum ◽  
Complement Pathway ◽  
Classical Complement Pathway ◽  
Recognition Phase ◽  
Normal Human ◽  
Classical Complement

We have recently found that intravenous immunoglobulin (IVIg) prevents deposition of C3 and C4 fragments onto antibody sensitized erythrocytes. To find out if such an effect results from the blockade of the recognition phase of the classical complement cascade, we investigated the ability of human serum containing high concentrations of IVIg to deposit the recognition subunit of the first complement component (C1q) onto targets. Normal human serum supplemented in vitro with IVIg did not demonstrate reduced C1q binding to targets as determined by radiolabeled antihuman C1q antibody uptake. Similarly, methylamine-treated normal human serum to which IVIg was added was equally effective in terms of C1q binding as the same serum without IVIg. At increasing doses of sensitizing antibody, C1q uptake decreased proportionally; however, at all antibody dilution points C1q uptake was not significantly different in the serum with IVIg in comparison with normal serum. Serum from a patient treated with IVIg did not differ in its capacity to deposit C1q from the same patient's serum before therapy. Our data suggest that IVIg does not interfere with the recognition step of classical complement pathway. This is a US government work. There are no restrictions on its use.

scholarly journals The UspA2 Protein of Moraxella catarrhalis Is Directly Involved in the Expression of Serum Resistance

2005 ◽  
Vol 73 (4) ◽  
pp. 2400-2410 ◽  
Author(s):  
Ahmed S. Attia ◽  
Eric R. Lafontaine ◽  
Jo L. Latimer ◽  
Christoph Aebi ◽  
George A. Syrogiannopoulos ◽  
...  
Keyword(s):  
Human Serum ◽  
Resistant Strain ◽  
Moraxella Catarrhalis ◽  
Normal Human Serum ◽  
Serum Resistance ◽  
Complement Pathway ◽  
Classical Complement Pathway ◽  
Amino Acid Region ◽  
Normal Human ◽  
Classical Complement

ABSTRACT Many strains of Moraxella catarrhalis are resistant to the bactericidal activity of normal human serum. Previous studies have shown that mutations involving the insertion of an antibiotic resistance cartridge into the M. catarrhalis uspA2 gene resulted in the conversion of a serum-resistant strain to a serum-sensitive phenotype. In the present study, the deletion of the entire uspA2 gene from the serum-resistant M. catarrhalis strain O35E resulted in a serum-sensitive phenotype and did not affect either the rate of growth or the lipooligosaccharide expression profile of this mutant. Inactivation of the classical complement pathway in normal human serum with Mg2+ and EGTA resulted in the survival of this uspA2 mutant. In contrast, blocking of the alternative complement pathway did not protect this uspA2 mutant from complement-mediated killing. To determine whether the UspA2 protein is directly involved in serum resistance, transformation and allelic exchange were used to replace the uspA2 gene in the serum-resistant strain O35E with the uspA2 gene from the serum-sensitive M. catarrhalis strain MC317. The resultant O35E transformant exhibited a serum-sensitive phenotype. Similarly, when the uspA2 gene from the serum-resistant strain O35E was used to replace the uspA2 gene in the serum-sensitive strain MC317, the MC317 transformant acquired serum resistance. The use of hybrid O35E-MC317 uspA2 genes showed that the N-terminal half of the O35E protein contained a 102-amino-acid region that was involved in the expression of serum resistance. In addition, when the uspA2 genes from strains O35E and MC317 were cloned and expressed in Haemophilus influenzae DB117, only the O35E UspA2 protein caused a significant increase in the serum resistance of the H. influenzae recombinant strain. These results prove that the UspA2 protein is directly involved in the expression of serum resistance by certain M. catarrhalis strains.

scholarly journals Phosphoethanolamine Substitution of Lipid A and Resistance of Neisseria gonorrhoeae to Cationic Antimicrobial Peptides and Complement-Mediated Killing by Normal Human Serum

2008 ◽  
Vol 77 (3) ◽  
pp. 1112-1120 ◽  
Author(s):  
Lisa A. Lewis ◽  
Biswa Choudhury ◽  
Jacqueline T. Balthazar ◽  
Larry E. Martin ◽  
Sanjay Ram ◽  
...  
Keyword(s):  
Neisseria Gonorrhoeae ◽  
Human Serum ◽  
Lipid A ◽  
Polymyxin B ◽  
Normal Human Serum ◽  
Bactericidal Action ◽  
Complement Pathway ◽  
Classical Complement Pathway ◽  
Normal Human ◽  
Classical Complement

ABSTRACT The capacity of Neisseria gonorrhoeae to cause disseminated gonococcal infection requires that such strains resist the bactericidal action of normal human serum. The bactericidal action of normal human serum against N. gonorrhoeae is mediated by the classical complement pathway through an antibody-dependent mechanism. The mechanism(s) by which certain strains of gonococci resist normal human serum is not fully understood, but alterations in lipooligosaccharide structure can affect such resistance. During an investigation of the biological significance of phosphoethanolamine extensions from lipooligosaccharide, we found that phosphoethanolamine substitutions from the heptose II group of the lipooligosaccharide β-chain did not impact levels of gonococcal (strain FA19) resistance to normal human serum or polymyxin B. However, loss of phosphoethanolamine substitution from the lipid A component of lipooligosaccharide, due to insertional inactivation of lptA, resulted in increased gonococcal susceptibility to polymyxin B, as reported previously for Neisseria meningitidis. In contrast to previous reports with N. meningitidis, loss of phosphoethanolamine attached to lipid A rendered strain FA19 susceptible to complement killing. Serum killing of the lptA mutant occurred through the classical complement pathway. Both serum and polymyxin B resistance as well as phosphoethanolamine decoration of lipid A were restored in the lptA-null mutant by complementation with wild-type lptA. Our results support a role for lipid A phosphoethanolamine substitutions in resistance of this strict human pathogen to innate host defenses.

scholarly journals Complement Interaction with Trypanosomatid Promastigotes in Normal Human Serum

2002 ◽  
Vol 195 (4) ◽  
pp. 451-459 ◽  
Author(s):  
Mercedes Domínguez ◽  
Inmaculada Moreno ◽  
Margarita López-Trascasa ◽  
Alfredo Toraño
Keyword(s):  
Human Serum ◽  
Binding Capacity ◽  
Normal Human Serum ◽  
Leishmania Infection ◽  
Classical Pathway ◽  
Classical Complement Pathway ◽  
Ethylenediaminetetracetic Acid ◽  
Human Sera ◽  
Normal Human

In normal human serum (NHS), axenic promastigotes of Crithidia, Phytomonas, and Leishmania trigger complement activation, and from 1.2 to 1.8 × 105 C3 molecules are deposited per promastigote within 2.5 min. In Leishmania, promastigote C3 binding capacity remains constant during in vitro metacyclogenesis. C3 deposition on promastigotes activated through the classical complement pathway reaches a 50% maximum after ∼50 s, and represents >85% of total C3 bound. In C1q- and C2-deficient human sera, promastigotes cannot activate the classical pathway (CP) unless purified C1q or C2 factors, respectively, are supplemented, demonstrating a requirement for CP factor in promastigote C3 opsonization. NHS depleted of natural anti-Leishmania antibodies cannot trigger promastigote CP activation, but IgM addition restores C3 binding. Furthermore, Leishmania binds natural antibodies in ethylenediaminetetracetic acid (EDTA)-treated NHS; after EDTA removal, promastigote-bound IgM triggers C3 deposition in natural antibody-depleted NHS. Serum collectins and pentraxins thus do not participate significantly in NHS promastigote C3 opsonization. Real-time kinetic analysis of promastigote CP-mediated lysis indicates that between 85–95% of parasites are killed within 2.5 min of serum contact. These data indicate that successful Leishmania infection in man must immediately follow promastigote transmission, and that Leishmania evasion strategies are shaped by the selective pressure exerted by complement.

scholarly journals Simple Method To Distinguish between Primary and Secondary C3 Deficiencies

2003 ◽  
Vol 10 (2) ◽  
pp. 216-220
Author(s):  
Marlene Pereira de Carvalho Florido ◽  
Patrícia Ferreira de Paula ◽  
Lourdes Isaac
Keyword(s):  
Human Serum ◽  
Complement System ◽  
Factor H ◽  
Normal Human Serum ◽  
Alternative Complement Pathway ◽  
Complement Pathway ◽  
Factor B ◽  
Alternative Complement ◽  
Normal Human ◽  
The Complement System

ABSTRACT Due to the increasing numbers of reported clinical cases of complement deficiency in medical centers, clinicians are now more aware of the role of the complement system in the protection against infections caused by microorganisms. Therefore, clinical laboratories are now prepared to perform a number of diagnostic tests of the complement system other than the standard 50% hemolytic component assay. Deficiencies of alternative complement pathway proteins are related to severe and recurrent infections; and the application of easy, reliable, and low-cost methods for their detection and distinction are always welcome, notably in developing countries. When activation of the alternative complement pathway is evaluated in hemolytic agarose plates, some but not all human sera cross-react to form a late linear lysis. Since the formation of this linear lysis is dependent on C3 and factor B, it is possible to use late linear lysis to routinely screen for the presence of deficiencies of alternative human complement pathway proteins such as factor B. Furthermore, since linear lysis is observed between normal human serum and primary C3-deficient serum but not between normal human serum and secondary C3-deficient serum caused by the lack of factor H or factor I, this assay may also be used to discriminate between primary and secondary C3 deficiencies.

scholarly journals Herpes Simplex Virus Type 1 and 2 Glycoprotein C Prevents Complement-Mediated Neutralization Induced by Natural Immunoglobulin M Antibody

2006 ◽  
Vol 80 (8) ◽  
pp. 4038-4046 ◽  
Author(s):  
Lauren M. Hook ◽  
John M. Lubinski ◽  
Ming Jiang ◽  
Michael K. Pangburn ◽  
Harvey M. Friedman
Keyword(s):  
Herpes Simplex ◽  
Human Serum ◽  
Immunoglobulin M ◽  
Complement Pathway ◽  
Glycoprotein C ◽  
Classical Complement Pathway ◽  
Simplex Virus ◽  
Classical Complement ◽  
Hsv 1

ABSTRACT Glycoprotein C (gC) of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) binds complement component C3b and protects virus from complement-mediated neutralization. Differences in complement interacting domains exist between gC of HSV-1 (gC1) and HSV-2 (gC2), since the amino terminus of gC1 blocks complement C5 from binding to C3b, while gC2 fails to interfere with this activity. We previously reported that neutralization of HSV-1 gC-null virus by HSV antibody-negative human serum requires activation of C5 but not of downstream components of the classical complement pathway. In this report, we evaluated whether activation of C5 is sufficient to neutralize HSV-2 gC-null virus, or whether formation of the membrane attack complex by C6 to C9 is required for neutralization. We found that activation of the classical complement pathway up to C5 was sufficient to neutralize HSV-2 gC-null virus by HSV antibody-negative human serum. We evaluated the mechanisms by which complement activation occurred in seronegative human serum. Interestingly, natural immunoglobulin M antibodies bound to virus, which triggered activation of C1q and the classical complement pathway. HSV antibody-negative sera obtained from four individuals differed over an approximately 10-fold range in their potency for complement-mediated virus neutralization. These findings indicate that humans differ in the ability of their innate immune systems to neutralize HSV-1 or HSV-2 gC-null virus and that a critical function of gC1 and gC2 is to prevent C5 activation.

State of iron(III) in normal human serum: low molecular weight and protein ligands besides transferrin

1970 ◽  
Vol 48 (12) ◽  
pp. 1339-1350 ◽  
Author(s):  
Bibudhendra Sarkar
Keyword(s):  
Molecular Weight ◽  
Human Serum ◽  
Binding Capacity ◽  
Normal Serum ◽  
Normal Human Serum ◽  
Void Volume ◽  
Antibody Concentration ◽  
Low Molecular Weight ◽  
Protein Ligands ◽  
Normal Human

A fraction of Fe(III) in normal human serum is bound to both low molecular weight as well as protein ligands besides transferrin. Citrate was shown to be the major Fe(III)-binding substance in the low molecular weight fraction. Amino acids, sugars, and organic acids, such as ascorbate, pyruvate, and lactate, showed very little or no binding to Fe(III) in normal serum. Iron(III)-binding proteins other than transferrin were shown to be present in normal serum when the native serum with [59Fe(III)] was fractionated by (NH4)2SO4 and Sephadex G-150. The presence of these proteins was observed when trace amounts of Fe(III) were added to the normal serum and when the iron-binding capacity was saturated with Fe(III) to 50% and 100%. These proteins were eluted in the void volume of Sephadex G-150 and none of them corresponded electrophoretically to transferrin. The results of the gel filtration of a mixture of [131I]-transferrin and the proteins eluted in the void volume of Sephadex G-150 were strongly in favor of the Fe(III)-proteins as being neither transferrin aggregates nor transferrin adducts with other proteins. Immunoelectrophoresis of the Sephadex G-150 void volume proteins on agar gel against the antibody to transferrin revealed the absence of transferrin. The presence of at least six proteins in this fraction was shown by immunoelectrophoresis. Positive precipitin reactions were obtained with the antibodies to α2-macroglobulin, γG-globulin, γA-globulin, and γM-globulin. At least two more proteins in this fraction remained unidentified. When the same fraction containing [59Fe(III)] was treated with the whole antisera and the precipitates were counted for radioactivity, a typical antigen-antibody reaction curve was obtained as the antibody concentration was increased. Similar experiments with this fraction and antibodies to α2-macroglobulin, γG-globulin, γA-globulin, and γM-globulin failed to show any significant radioactivity in the precipitate. Since this fraction did not contain any transferrin, it was concluded that there are proteins besides transferrin which can act as ligands for Fe(III) in normal blood plasma.

The Binding of Lithium Carmine to a2-Macroglobulin

1969 ◽  
Vol 24 (11) ◽  
pp. 1442-1447 ◽  
Author(s):  
J. J. Picard ◽  
J. F. Heremans
Keyword(s):  
Human Serum ◽  
Density Gradient ◽  
Protein Complex ◽  
Gel Filtration ◽  
Heavy Fraction ◽  
Normal Human Serum ◽  
Density Gradient Ultracentrifugation ◽  
Γ Globulins ◽  
Normal Human

The colloidal dye lithium carmine was added in vitro to normal human serum. Electrophoretic experiments showed that the dye was associated mainly with α2-globulins, small amounts with the albumin and only traces with the γ-globulins. The main complex was eluted with the macroglobulin peak obtained by gel filtration on Sephadex G-200 and sedimented in the heavy fraction on density gradient ultracentrifugation. The dye-protein complex could be precipitated with an antiserum specific for a2-macroglobulin. Gel filtration of a solution of pure a2-macroglobulin, to which lithium carmine was added, demonstrated that the dye was bound to this protein.

scholarly journals FORMATION OF C3a AND C5a ANAPHYLATOXINS IN WHOLE HUMAN SERUM AFTER INHIBITION OF THE ANAPHYLATOXIN INACTIVATOR

1973 ◽  
Vol 137 (5) ◽  
pp. 1109-1123 ◽  
Author(s):  
Enrique H. Vallota ◽  
Hans J. Müller-Eberhard
Keyword(s):  
Human Serum ◽  
Active Form ◽  
Aminocaproic Acid ◽  
Classical Complement Pathway ◽  
Effective Doses ◽  
Epsilon Aminocaproic Acid ◽  
Antigen Antibody ◽  
Minimal Effective Concentration ◽  
Classical Complement

Two biologically and chemically distinct anaphylatoxins (ATs) could be generated in whole human serum after removal of the AT inactivator (AI) by immune-absorption or after inhibition of AI with 1 M epsilon-aminocaproic acid (EACA). Both human ATs could be generated by treatment of serum with antigen-antibody complexes, which activate the classical complement pathway, and with inulin or yeast, both of which trigger the alternate pathway. The ATs were isolated from serum in active form and characterized as C3a and C5a. Although human C3a had been characterized previously, C5a had not. The molecular weight of human C5a AT was 17,500; its electrophoretic mobility at pH 8.5 was –1.7 x 10–5 cm2 V–1 s–1. The minimal effective concentration in vitro was 7.5 x 10–10 M. The minimal effective doses of human C5a in producing a wheal and erythema in the human skin was 1 x 10–15 mol. The results strongly suggest a biological function for both ATs and indicate that the expression of their activity is controlled by the AI of normal blood plasma.

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