SUMMARYThe complement (C)-activating capabilities in human serum of 32 mouse and 10 mouse/human ehimeric MoAbs of different isotypes. and their fragments, v^ere tested in vitro. Aetivation of C via the classical pathway (CP) was F>erformed in 1 % faetor D-defieient serum in gelatin containing Veronal bufTer in the presenee of ealeium and magnesium (GVB'^*). while activation of the alternative pathway of C (AP) was assessed in iO'> Clq-depleted serum in the presenee of 5 mM MgCl; in GVB^ •, The C-aetivating ability of MoAbs was expressed relative to the degree of activation of complement by aggregated IgG for the CP and relative to mouse tgG I for the AP. Att of seven mouse IgG2a MoAbs were potent activators of the CP. The results of CP aetivation by IgG I, IgG2b and IgG3 isotypes were dilTerent for individual MoAbs. Only three (two IgG! and one IgG3)of32 mouse MoAbs were potent activators ofthe AP. lgG2a and IgG2b were relativety poor AP activators. There were a few MoAbs whieh aetivated both the AP and CP. Of tO ehimeric MoAbs. two IgGt. one IgG2 and one lgG4 were poor or non-activators ofthe CP. On the other hand, igG2 and IgG4 were good APacIivalors. IgG3 was the mcsl polent AP activator. Mosl of IheF(ab); rragmenis wereaclivators ofthe AP and displayed no activation ofthe CP. Fe tragments onty aetivated the CP, whereas Fab' did not activate the CP or the AP. These studies suggest that the route of comptement activation by etass and subelass MoAbs ean not atways be predicted in advance and based only on their subclass identity.
In this article we describe the isolation of a 4-carboxyglutamic acid (Gla)-containing protein from calcified human atherosclerotic plaques. The protein was extracted from pulverized calcified plaques by demineralization with ethylenediaminetetraacetate and was subsequently purified by anion-exchange fast protein and highperformance liquid chromatography by using ion-exchange and gel-filtration columns. The protein was designated as plaque Gla protein (PGP) and has an apparent mass of 23 kD as estimated from sodium dodecyl sulfate-polyacrylamide gel analysis. By determining the sequence of its first six amino acid residues, the protein was unequivocally demonstrated to be not related to any other known protein.Moreover, no immunological relationship (as tested by Western blot analysis) was found between PGP and other known Gla-containing proteins. (Arteriosclerosis 10:991-995, November/December 1990)
SummaryIn this paper we present the following observations:1) In sheep vitamin K-antagonists like phenprocoumon induce a decrease of the serum levels of osteocalcin (bone Gla-protein) and of the affinity of the circulating osteocalcin for hydroxyapatite.2) In sheep vitamin K counteracts the effect of phen-procoumon on the blood coagulation system, but not that on the osteocalcin production.3) In human subjects vitamin K-antagonists also lead to decreased levels of serum osteocalcin and a low affinity of the protein for hydroxyapatite.4) These two variables reached steady-state levels within 24 h after the start of oral anticoagulant treatment and - at continu-ation of the therapy - they remained low for at least several years.
In the liver vitamin K epoxide, which is produced during the posttranslational carboxylation of proteinbound glutamic acid residues, is recycled by the action of one or more dithiol‐dependent reductases. In vitro synthetic dithiols may serve as a cofactor for these enzymes, but the physiological reluctant has not yet been found. In this paper we report that in vitro the commercially available thioredoxin/thioredoxin reductase from E. coli can replace the synthetic dithiols during the various reactions of the vitamin K cycle. Based on the assumption that in vivo thioredoxin also plays a role in the regeneration of vitamin K hydroquinone from the epoxide, an extension of the generally accepted vitamin K cycle is proposed.
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