Effect of sequential glycolysis of the lectin of Euonymus europaeus on activation of the classical complement pathway in normal human serum

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.

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

Blood ◽

10.1182/blood.v78.3.700.bloodjournal783700 ◽

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.

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

Blood ◽

10.1182/blood.v78.3.700.700 ◽

1991 ◽

Vol 78 (3) ◽

pp. 700-702 ◽

Cited By ~ 16

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.

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

Infection and Immunity ◽

10.1128/iai.01280-08 ◽

2008 ◽

Vol 77 (3) ◽

pp. 1112-1120 ◽

Cited By ~ 70

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.

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Simple Method To Distinguish between Primary and Secondary C3 Deficiencies

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.10.2.216-220.2003 ◽

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.

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

Journal of Virology ◽

10.1128/jvi.80.8.4038-4046.2006 ◽

2006 ◽

Vol 80 (8) ◽

pp. 4038-4046 ◽

Cited By ~ 48

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.

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In vitro complement binding on cytoplasmic structures in normal human skin: immunoelectronmicroscopic studies.

The Journal of Cell Biology ◽

10.1083/jcb.95.2.543 ◽

1982 ◽

Vol 95 (2) ◽

pp. 543-551 ◽

Cited By ~ 12

Author(s):

G Schuler ◽

H Hintner ◽

K Wolff ◽

P Fritsch ◽

G Stingl

Keyword(s):

Cell Types ◽

Exact Nature ◽

Complement Pathway ◽

Experimental Conditions ◽

Complement Components ◽

Classical Complement Pathway ◽

Pathway Activation ◽

Normal Human ◽

Cytoplasmic Structures ◽

Classical Complement

We have previously provided evidence that suggests that exposure of cryostat skin sections to normal human serum (NHS) results in the antibody-independent Clq binding to cytoplasmic structures of various cell types, leading to classical complement pathway activation as evidenced by cytoplasmic C3 deposition. In the present study, we have employed immunoelectronmicroscopic methods to clarify the exact nature of cytoplasmic C3 binding structures. Incubation of cryostat skin sections with NHS followed by peroxidase-labeled rabbit anti-human C3 serum (HRP-R/Hu C3) revealed that intracytoplasmic binding of C3 occurred in suprabasal keratinocytes, melanocytes, fibroblasts, smooth muscle cells, endothelial cells, pericytes, Schwann cells, and nerve axons, but not in basal keratinocytes, Langerhans cells, and other cellular constituents of the skin. C3 binding, as revealed by the deposition of HRP reaction product, was exclusively confined to intermediate-sized filaments (ISF), which can therefore be considered to represent the subcellular site for classical complement pathway activation. Under experimental conditions that do not allow classical complement pathway activation, ISF were not decorated. Our observation that ISF of ontogenetically different cell types share the capacity of complement fixation is in accordance with the recent finding that different ISF types, despite their biochemical and antigenic heterogeneity, have common alpha-helical domains and may provide a clue to the mechanism and site of interaction between complement components and ISF.

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Kingella kingaeSurface Polysaccharides Promote Resistance to Human Serum and Virulence in a Juvenile Rat Model

Infection and Immunity ◽

10.1128/iai.00100-18 ◽

2018 ◽

Vol 86 (6) ◽

Cited By ~ 7

Author(s):

Vanessa L. Muñoz ◽

Eric A. Porsch ◽

Joseph W. St. Geme

Keyword(s):

Human Serum ◽

Invasive Disease ◽

Complement Pathway ◽

Kingella Kingae ◽

Content Type ◽

Bacterial Surface ◽

Classical Complement Pathway ◽

Bactericidal Effects ◽

Pathogenic Process ◽

Classical Complement

ABSTRACTKingella kingaeis a Gram-negative coccobacillus that is increasingly being recognized as an important cause of invasive disease in young children. The pathogenesis ofK. kingaedisease begins with colonization of the oropharynx, followed by invasion of the bloodstream, survival in the intravascular space, and dissemination to distant sites. Recent studies have revealed thatK. kingaeproduces a number of surface factors that may contribute to the pathogenic process, including a polysaccharide capsule and an exopolysaccharide. In this study, we observed thatK. kingaewas highly resistant to the bactericidal effects of human serum complement. Using mutant strains deficient in expression of capsule, exopolysaccharide, or both in assays with human serum, we found that elimination of both capsule and exopolysaccharide was required for efficient binding of IgG, IgM, C4b, and C3b to the bacterial surface and for complement-mediated killing. Abrogation of the classical complement pathway using EGTA-treated human serum restored survival to wild-type levels by the mutant lacking both capsule and exopolysaccharide, demonstrating that capsule and exopolysaccharide promote resistance to the classical complement pathway. Consistent with these results, loss of both capsule and exopolysaccharide eliminated invasive disease in juvenile rats with an intact complement system but not in rats lacking complement. Based on these observations, we conclude that the capsule and the exopolysaccharide have important redundant roles in promoting survival ofK. kingaein human serum. Each of these surface factors is sufficient by itself to fully prevent serum opsonin deposition and complement-mediated killing ofK. kingae, ultimately facilitating intravascular survival and promotingK. kingaeinvasive disease.

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