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Biomedical subjects

N R Cooper

Publications and source records attributed to N R Cooper.

At least 127 records · Page 7Linked to original sources
PubMed1976

Physicochemical and functional characterization of the C1r subunit of the first complement component.

C1r was isolated from serum by an improved method and found to be a glycoprotein with a sedimentation coefficient of 7.0S. Under conditions of physiologic ionic strength and pH, C1r consist of two apparently identical noncovalently linked 95,000 dalton polypeptide chains. Antisera to C1r detected a protein of gamma-mobility on electrophoresis of serum in agarose in the presence of calcium, and a Beta-mobility protein when the electrophoretic separation was carried out in EDTA. On sucrose gradient ultracentrifugation of normal human serum in the presence of calcium, C1r antigenicity was found in the 19 S region of the gradient. On the other hand, when the gradient contained EDTA, C1r antigenicity was found in the 7 S region. No reaction of anti-C1r with C1r-deficient sera was observed. C1r had a high affinity for active C1s or proenzyme C1s in the presence of calcium and was able to activate C1s and to form C1 in conjunction with C1q and C1s. Activation of C1s by C1r was inhibited by calcium, C1 inactivator, polyanethol sulfonate, and DFP. Activation of C1s by C1r occurred only after a preliminary incubation of C1r for a brief time at 37 degrees C before addition of C1s. The ability of C1r to form C1 in conjunction with C1q and C1s was, however, progressively lost on incubation at 37 degrees C. Trypsin, although potentiating the activity of crude C1r, did not modify the activity of purified C1r. Its action was on a trypsin-sensitive inhibitor separated from C1r in the final step of the isolation procedure. The binding of 125I-C1r to sensitized sheep erythrocytes required the presence of C1q and calcium but not C1s, whereas the binding of 125I-C1s required C1q, C1r and calcium. Thus, C1r functions as not only the activator of C1s, but also serves as the physical link between C1q and C1s in macromolecular C1.

Animals↗
PubMed1975

Enzymatic activity of the second component of complement.

Isolated C2 and C2i preparations were able to hydrolyze a number of synthetic esters containing basic amino acids, among which N-alpha-acetylglycyl-L-lysine methyl ester (AcGlyLysOMe) was most susceptible. The cleaving activity was a property of the C2 molecule, since it correlated with the presence of C2 on analyses of C2 preparations by ultracentrifugation in sucrose gradients, filtration through Sephadex G-200 columns, and on electrophoresis in acrylamide gels. Furthermore, acrylamide gel electrophoretic studies showed a shift in hydrolytic activity from the position occupied by C2 to that characteristic of C2i after incubation of C2 with C1s. The action was enzymatically mediated as evidenced by a bell-shaped pH activity curve, a linear dependence on C2 concentration, and the presence of Michaelis-Menten kinetics. The Michaelis constant for cleavage of AcGlyLysOMe by C2 was 1.8 X 10(-2) mol. Cleavage of C2 by C1s increased C2 enzymatic activity, yet chemical oxidation of the molecule, although enhancing hemolytic acitivity, failed to increase C2 hydrolytic activity. The observed enzymatic activity of C2 was found to be relevant to the function of C2 in the C42 complex, since AcGlyLysOMe competitively inhibited the C42 mediated cleavage of C3 in free solution and the C42 dependent binding of C3 to cells.

Complement C2↗
PubMed1975

Immunologic injury of cultured cells infected with measles virus. I. role of IfG antibody and the alternative complement pathway.

In these studies, a number of human cell lines including epithelial, neural, glial, and lymphoid cells infected with several strains of measles virus were found to be lysed upon incubation with fresh sera from humans containing antibody measles virus. In all instances, the cytolytic event was mediated by alternative complement (C) pathway without a significant contribution from classical pathway. In contrast, isolated measles virus in conjunction with antibody was found to selectively activate the classical C pathway. Measles antibodies of the IgG class, but not the IgA class, possessed cytolytic potential against cells infected with measles virus. Human IgG antibodies could directly activate the alternative C pathway. No defect was found in cytolytic measles antibody in sera or cerebrospinal fluid from patients with subacute sclerosing panencephalitis, nor was the alternative C pathway impaired in sera from these patients. Sera from newborn humans exhibited a functional alternative C pathway.

Absorption↗
PubMed1975

Isolation and analysis of the mechanism of action of an inactivator of C4b in normal human serum.

A complement regulatory principle, C4b inactivator, was isolated in a partially purified form from normal human serum. The C4b inactivator, a beta1-globulin with an approximate mol wt of 88,000 daltons, and which may be identical to C3b inactivator, cleaved C4b in free solution or on the surface of cells and rendered it unable to participate in hemolytic reactions or to interact with cells, having receptors for C4b. C/b inactivator functioned by cleaving the alpha-polypeptide chain of C4b at a single site which was sufficient to dissociate the molecule into two fragments, C4c and C4d, and to inactivate it biological function. Certain structural correlates of C4 functions deriving from these studies are discussed and a model for C4 structure based on these findings is presented.

Animals↗
PubMed1975

The relationship between hemolytic and immunodiffusion methods for measurement of C4 in patients with immunologic disorders.

A comparison of radial immunodiffusion and functional (hemolytic) assays for C4 revealed a highly significant correlation between the two methods in patients with a variety of immunologic disorders. Immunodiffusion technics are more convenient than hemolytic assays for most clinical laboratories. C4 assays are useful for the presumptive diagnosis of hereditary angioneurotic edema, whereas C3 is usually normal in such patients. C4 is usually more sensitive than C3 in a variety of immunologic disorders, although in some patients the opposite is true, especially in hypocomplementemic nephritis.

Angioedema↗
PubMed1975

Complement fixation by rheumatoid factor.

The capacity for fixation and activation of hemolytic complement by polyclonal IgM rheumatoid factors (RF) isolated from sera of patients with rheumatoid arthritis and monoclonal IgM-RF isolated from the cryoprecipitates of patients with IgM-IgG mixed cryoglobulinemia was examined. RF mixed with aggregated, reduced, and alkylated human IgG (Agg-R/A-IgG) in the fluid phase failed to significantly reduce the level of total hemolytic complement, CH50, or of individual complement components, C1, C2, C3, and C5. However, sheep erythrocytes (SRC) coated with Agg-R/A-IgG or with reduced and alkylated rabbit IgG anti-SRC antibody were hemolyzed by complement in the presence of polyclonal IgM-RF. Human and guinea pig complement worked equally well. The degree of hemolysis was in direct proportion to the hemagglutination titer of the RF against the same coated cells. Monoclonal IgM-RF, normal human IgM, and purified Waldenström macroglobulins without antiglobulin activity were all inert. Hemolysis of coated SRC by RF and complement was inhibited by prior treatment of the complement source with chelating agents, hydrazine, cobra venom factor, specific antisera to C1q, CR, C5, C6, or C8, or by heating at 56 degrees C for 30 min. Purified radiolabeled C4, C3, and C8 included in the complement source were bound to hemolysed SRC in direct proportion to the degree of hemolysis. These data indicate that polyclonal IgM-RF fix and activate complement via the classic pathway. The system described for assessing complement fixation by isolated RF is readily adaptable to use with whole human serum.

Animals↗
PubMed1975

Studies on human plasma C1 inactivator-enzyme interactions. I. Mechanisms of interaction with C1s, plasmin, and trypsin.

This study has explored the nature of the molecular events which occur when C1 inactivator, a human plasma inhibitor of the complement, kinin-forming, coagulation, and fibrinolytic enzyme systems, interacts with C1s, plasmin, and trypsin. Purified inhibitor preparations demonstrated two bands, when examined by acrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS). The molecular weights of the major and minor bands were 105,000 and 96,000 daltons, respectively. The minor component appeared to be immunologically and functionally identical to the main C1 inactivator component. Loss of C1s and plasmin functional activity was associated with the formation of a 1:1 molar complex between the inhibitor and each enzyme. These complexes were stable in the presence of SDS and urea. The light chain of both these enzymes provided the binding site for C1 inactivator. Complex formation and enzyme inhibition occurred only with native and not with an inhibitor preparation denatured by acid treatment, thereby demonstrating the importance of conformational factors in the enzyme-inhibitor reaction. Although peptide bond cleavage of the C1 inactivator molecule by C1s was not documented, plasmin was found to degrade the inhibitor with the production of several characteristic derivatives. At least one of these products retained the ability to complex with C1s and plasmin. Trypsin, which failed to form a complex with C1 inactivator, degraded the inhibitor in a limited and sequential manner with the production of nonfunctional derivatives one of which appeared structurally similar to a plasmin-induced product. These studies therefore, provide new information concerning the molecular interactions between C1 inactivator and several of the proteases which it inhibits.

Animals↗
PubMed1975

Studies on human plasma C1 inactivator-enzyme interactions. II. Structural features of an abnormal C1 inactivator from a kindred with hereditary angioneurotic edema.

The function and several of the structural features of the C1 inactivator protein isolated from the plasma of a mother and daughter with the variant form of hereditary angioneurotic edema have been examined. These abnormal inhibitors shared immunologic identity with the normal C1 inactivator protein; however, they were inactive in inhibiting the functional activity of C1s. Analysis of the abnormal inhibitors by sodium dodecyl sulfate (SDS) acrylamide gel electrophoresis suggested that each consisted of a single polypeptide chain, the mobility of which was slower than that of the normal C1 inactivator. The apparent molecular weight of the patients' inhibitors was 109,000 daltons as contrasted to 105,000 daltons, that of the normal C1 inactivator. The abnormal inhibitors failed to form a complex with C1s or plasmin as analyzed by SDS-acrylamide gels. The large proteolytic derivatives resulting from the plasmin- and trypsin-induced degradation of the abnormal inhibitors were approximately 3,000 daltons heavier than the corresponding products derived from normal C1 inactivator. Thus, the structural abnormality identified appeared to be a property of the core molecule. Treatment of the inhibitors with neuraminidase failed to demonstrate a difference between the normal and patient-derived C1 inactivator molecule. Neither were major differences found between the amino acid composition of the defective and normal inhibitors; however, the acidic amino acids tended to be higher in the patients' inhibitors, and the phenylalanine content lower. Thus, these studies have identified both structural and functional abnormalities in the C1 inactivator protein isolated from two related patients with hereditary angioneurotic edema. Examination of the interaction between endopeptidases and the inhibitors has further delineated the abnormal structural features.

Amino Acids↗
PubMed1974

Formation and biologic role of polyoma virus-antibody complexes. A critical role for complement.

Interaction of polyoma virus, specific antibody, and complement has been studied. Firm evidence has been gathered that C1 through C3 and not C5 through C9 enhance neutralization of virus-antiviral antibody (V-Ab) complexes. C enhancement of neutralization occurs primarily by agglutination of V-Ab complexes and not by virion lysis or attachment of large protein molecules to the V-Ab complex. In this model, binding of C1, 4, 2, 3 to the V-Ab complex may explain why some viruses concentrate in or infect certain cells bearing C3 receptors such as B lymphocytes, macrophages, and monocytes.

Animals↗
PubMed1974

Variation in susceptibility of a human lymphoid cell line to immune lysis during the cell cycle. Lack of correlation with antigen density and complement binding.

Cultured human lymphoid cells RPMI 8866 at different stages of their growth cycle vary in their susceptibility to lysis by rabbit, human, and guinea pig complement activated by HL-A antibodies or heterologous antibodies directed to membrane antigens; cells in G(1) phase are the least sensitive to lysis. To investigate the cause of differential susceptibility of cells RPMI 8866 to lysis, the expression of HL-A determinants and the ability of cells to react with complement were investigated. No change was detected in the density of HL-A antigens on RPMI 8866 cells in synchronous growth as determined by quantitative microabsorption assays, isotopic antiglobulin tests and yields of soluble HL-A antigens. Cells did not vary during the growth cycle in their ability to interact with complement components and in their capacity to activate the complement system through the classical or alternate pathway. These data suggest that variability in lytic susceptibility is due to changes in the structure of the cell membrane or in its ability to repair complement induced damage at certain intervals during the cell cycle. Therefore, this cell line constitutes a useful model to investigate the final steps of the cytolytic reaction.

Animals↗
PubMed1974

Glomerular complement components in human glomerulonephritis.

154 of 255 individual human renal biopsies studied by immunofluorescence contained varying combinations of immunoglobulins (Ig), complement (C) components C1q, C3, C4, C5, C6, C8, C3 proactivator (C3PA), and/or properdin. 10 patients had linear deposits of Ig in glomeruli characteristic of antiglomerular basement membrane (GBM) antibodies; nine patients had C3 deposits (minimal in three) with generally lesser amounts of C1q, C4, C5, C6, and/or C8. 118 of the patients had granular deposits of Ig, suggesting immune complex glomerulonephritis; 114 of these had deposits of C3, usually accompanied by C1q, C4, C5, and/or C6. These observations indicate that the entire C sequence is deposited in glomeruli in most Ig-mediated glomerulonephritides. However, certain cases of anti-GBM glomerulonephritis with few or no C deposits may utilize pathways of injury independent of C.21 patients had granular C3 deposits without detectable Ig. C5, C6, and C8 were present in the majority of these patients while C1q was absent and scant C4 was observed in only two patients. The presence of only late-acting C components in the absence of Ig, C1q, and C4 suggests selective, possible nonimmune activation of the alternate C pathway. Finally, five patients had granular deposits of C3, C5, C6, and/or C8 diffusely in all or most glomeruli with a lesser number of glomeruli having additional focal granular deposits of Ig, C1q, and C4. This observation suggests that at least two patterns of C activation can occur simulatenously, possibly triggered by antecedent immune complex deposition and then perpetuated by an as yet undetermined mechanism.

Adult↗