Biomedical subjects

E Jelezarova

Publications and source records attributed to E Jelezarova.

4 recordsLinked to original sources

PubMed2001

A C3 convertase assay for nephritic factor functional activity.

C3 nephritic factor (C3NeF) is an autoantibody against the C3 convertase which stabilizes this otherwise inherently labile neoenzyme and induces a continuous activation of the alternative pathway with C3 depletion. NeF is found in patients with membranoproliferative glomerulonephritis and/or partial lipodystrpohy. NeF activity is usually detected in plasma by hemolytic tests. In order to obtain reproducible data for the functional activity of purified C3NeF IgG a solid phase assay was developed. C3 convertase was generated on immobilized C3b by incubation with factors B and D in the presence of Ni(2+). Convertase sites were left to decay in the presence of normal IgG or NeF IgG. Residual convertase activity was measured by adding 125I-C3 and capturing nascent 125I-C3b on the plate surface via covalently coupled NH2-Glu-Tyr dipeptide. In the presence of factor H during C3 convertase decay, a dose dependent stabilizing activity was shown for NeF IgG including NeF IgG purified from urine. A second format of the assay was developed in which C3 convertase was assembled on C3b(2)-IgG complexes in the presence of Mg(2+). Since these complexes are more efficient as convertase precursors the signal was five-fold higher than with C3b. Convertase decay, on the other hand, was not influenced by the nature of the precursor and in both systems the stabilizing activity of NeF IgG was similar.

Adult↗

PubMed2000

Interaction of C3b(2)--IgG complexes with complement proteins properdin, factor B and factor H: implications for amplification.

Nascent C3b can form ester bonds with various target molecules on the cell surface and in the fluid phase. Previously, we showed that C3b(2)--IgG complexes represent the major covalent product of C3 activation in serum [Lutz, Stammler, Jelezarova, Nater and Späth (1996) Blood 88, 184--193]. In the present report, binding of alternative pathway proteins to purified C3b(2)--IgG complexes was studied in the fluid phase by using biotinylated IgG for C3b(2)--IgG generation and avidin-coated plates to capture complexes. Up to seven moles of properdin 'monomer' bound per mole of C3b(2)--IgG at physiological conditions in the absence of any other complement protein. At low properdin/C3b(2)--IgG ratios bivalent binding was preferred. Neither factor H nor factor B affected properdin binding. On the other hand, properdin strongly stimulated factor B binding. Interactions of all three proteins with C3b(2)--IgG exhibited pH optima. An ionic strength optimum was most pronounced for properdin, while factor B binding was largely independent of the salt concentration. C3b(2)--IgG complexes were powerful precursors of the alternative pathway C3 convertase. In the presence of properdin, C3 convertase generated from C3b(2)--IgG cleaved about sevenfold more C3 than the enzyme generated on C3b. C3b(2)--IgG complexes could therefore maintain the amplification loop of complement longer than free C3b.

Complement C3-C5 Convertases↗

PubMed1996

High doses of immunoglobulin G attenuate immune aggregate-mediated complement activation by enhancing physiologic cleavage of C3b in C3bn-IgG complexes.

Intravenously applied human IgG has beneficial effects in treating inflammatory diseases, presumably because it has a complement attenuating role. This role of IgG was studied in vitro by following C3 activation and inactivation in sera that were supplemented with exogenous human IgG and incubated with immune aggregates. IgG added at 2 to 10 mg/mL stimulated the physiologic inactivation of C3b-containing complexes twofold to threefold in 20% sera. This, in turn, lowered the overall C3 activation by 28%, as new C3 convertases primarily assembled on C3b-containing complexes. Exogenous IgG (5 mg/mL) also stimulated inactivation of purified C3b2-IgG complexes, whereby their half-life dropped from 3-4 to 1.5 minutes in 20% serum. IgG appeared to act like a modulator of factor H and I because it did not stimulate inactivation of C3b-containing complexes in factor I-deficient serum. Thus, the known partial protection of C3bn-IgG complexes from inactivation by factor H and I was downregulated by high concentrations of IgG. The ability of high doses of IgG to stimulate complement inactivation is a novel regulatory role of IgG. This may be one of the molecular principles for its therapeutic efficacy in treating complement-mediated inflammations.

Antigen-Antibody Complex↗

PubMedDate not supplied

Assembly and regulation of the complement amplification loop in blood: the role of C3b-C3b-IgG complexes.

Amplification of complement activation in blood and serum starts on multi-protein complexes that act as precursors of an alternative C3 convertase. Among these covalently linked C4b-, C3b-, and IgG-containing complexes C3b-C3b-IgG complexes represent the major species containing C3b and IgG. Recent work on their purification and characterization is discussed. Special emphasis is placed on the arrangement of ester bonds in these complexes and their dual type of partial protection from inactivation. Partial protection from inactivation is mediated by properdin which binds to these complexes in the complete absence of any other complement protein. High dose IgG, known to stimulate inactivation of these complexes, appears to lower properdin binding in a process that also involves factor H. Properdin stimulates factor B binding to these complexes and renders them far better precursors of a C3 convertase than C3b. The available information allows a suggestion for a new scheme on how the amplification loop is assembled and regulated in blood and serum.

Antibody Formation↗