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L A Retegui

Publications and source records attributed to L A Retegui.

34 records · Page 2Linked to original sources
PubMed1989

[The antigenic topography of human growth hormone and the humoral expression of its various epitopes].

Twenty monoclonal antibodies (MAb) against human growth hormone (hGH) were used to establish the antigenic topography of this protein. Mapping experiments were carried out by testing the ability of paired MAb to bind simultaneously or separately to 126I-hGH. Since the specificity of the MAb versus hGH, human placental lactogen, animal prolactins and growth hormones was known, consideration of the whole set of results obtained supported the proposal of a tridimensional model for the antigenic structure of hGH. Further work correlated some of the mapped epitopes with particular regions within the primary structure of the hormone. In addition, it could be demonstrated that certain MAb may induce conformational changes in the antigen as a result of its binding. The humoral expression of various hGH epitopes during the maturation of the immune response was studied in human patients therapeutically treated with hGH and in mice and hamsters submitted to different immunization schemes: chronic administration of the antigen, secondary response and conventional hyperimmunization. The results indicated temporal and individual variations in the titers of each class of Ab as well as the existence of enhancer and heteroclitic Ab. Hence, the expression of the various epitopes of a protein antigen during the immune response is quite probably a stochastic process which depends on individual variations.

Antibodies, Monoclonal↗
PubMed1988

Stochastic humoral expression of human growth hormone epitopes.

Competition experiments between insolubilized monoclonal antibodies (mAb) and polyclonal antisera has led to the description of the humoral expression of human growth hormone (hGH) epitopes. This study was carried out with sera from mice and hamsters submitted to different immunization schedules: chronic administration of the antigen, secondary response and conventional hyperimmunization. The results indicated the absence of a unique immunodominant epitope in hGH; a significant individual variation of antibody (Ab) population titres with time; changes with time in the relative proportion of one Ab population with respect to the others; and the occurrence of Ab enhancing the 125I-hGH binding to five mAb depending upon the individuals and the time of immunization. Heterocliticity towards non-human GH was also detected. Although most of the animals showed cross-reacting Ab, two out of 12 mice, chronically injected, developed heteroclitic Ab. The data suggest that the humoral response to different epitopes of a protein antigen during the maturation of the immune response is a stochastic process leading to transient humoral immunodominance, enhancing Ab populations and heterocliticity, depending upon individual characteristics, either in outbred or inbred populations.

Animals↗
PubMed1987

Proteolytic modification of growth hormone by a rat kidney lysosomal protease.

A protease activity has been detected in rat kidney which has some degree of specificity to cleave native bovine growth hormone (bGH). The enzyme has an optimum pH of about 5.0, is dependent on thiol reagents and non-inhibited by EDTA or Aprotinin. Controlled digestion of bGH leads to a derivative retaining growth promoting activity, while showing a different physicochemical behavior from that of the native hormone. The data support the hypothesis of the kidney, among other tissues, originating active forms of growth hormones.

Animals↗
PubMed1986

Heteroclitic behaviour of some monoclonal antibodies against bovine growth hormone.

Five monoclonal antibodies (MAbs) prepared against bovine growth hormone (bGH) were found to be directed against an immunodominant antigenic domain in bGH. MAbs D8 and H3 reacted equally well with bGH and ovine growth hormone and to a lesser extent with equine (eGH) or porcine (pGH) growth hormones while MAbs H1, H2 and C12 behaved as heteroclitic antibodies, i.e. they bound better a cross-reacting antigen (eGH on pGH) than the immunogen. The reactivity of bGH with the heteroclitic MAbs increased proportionally to the time that the native protein was kept frozen. Deamidation of bGH by treatment with alkali also increased its reactivity. Circular dichroism measurements indicated the occurrence of changes in the conformation of the bGH molecule by these treatments which presumably uncover normally buried or non-accesible epitopes. bGH shares epitopes with eGH and pGH which are immunologically expressed by bGH only when its native conformation is modified.

Animals↗
PubMed1986

Relative distribution of various antigenic determinants on the human growth hormone surface.

The relative distribution of 12 antigenic determinants on the surface of the human growth hormone (hGH) molecule has been established. The necessary information was obtained by testing the ability of paired monoclonal antibodies (MAb) to bind simultaneously or not, to 125I-hGH which leads to the formation of 1:2 or 1:1, Ag-Ab complexes, respectively. The results obtained indicate that the epitopes occupy a large percentage of the total hGH molecular surface and revealed the existence of; an antigenic region specific for hGH; at least two independent domains of immunological identity between hGH and human placental lactogen (hPL), one of them also shared by heterologous GH; and other independent areas of partial cross reactivity with hPL. MAb competition experiments in a solid-phase RIA showed the unreliability of this technique for mapping purposes. The distribution of the hGH epitopes suggested in this work is in accord with present views on protein antigenicity and also explains data existing in the literature concerning the behavior of some of the MAb tested here.

Antibodies, Monoclonal↗
PubMed1985

Comparison of human lactoferrins from milk and neutrophilic leucocytes. Relative molecular mass, isoelectric point, iron-binding properties and uptake by the liver.

Human lactoferrins isolated from neutrophilic leucocytes and milk by CM-Sephadex chromatography were similar in Mr (76000) and pI (8.7). Upon acidification, both proteins released their two Fe3+ ions/molecule in a similar biphasic way. Both proteins intravenously injected into mice were cleared from plasma at the same rate. The maximal uptakes by the liver, which occurred 5 min after injection, were inhibited to the same extent by milk lactoferrin used as a competitor.

Animals↗
PubMed1985

Specificities of antibodies to human growth hormone (hGH) in patients treated with hGH: longitudinal study and comparison with the specificities of animal antisera.

The specificities of human and animal antibodies (Abs) against human GH (hGH) were analyzed using competition experiments with five monoclonal antibodies to hGH (MAbs). The results indicate that 1) the Abs produced by patients receiving long term hGH therapy as well as Abs of goat, rabbit, and mouse origin recognized the various hGH epitopes defined by the MAbs; 2) the proportion of each Ab population, with a given specificity, differed markedly in different patients and also with time in the same patient; 3) the titer of certain Ab populations was very low in some patients, and 4) polyclonal mouse Abs and some human Abs enhanced the binding of [125I] hGH to insolubilized MAbs.

Adolescent↗
PubMed1984

Monoclonal antibodies and specific cell surface receptors do not discriminate between human growth hormone prepared by DNA recombinant techniques and the native hormone.

Native and E. coli-derived human growth hormone (hGH) have been compared in their ability to react with four different monoclonal antibodies (MAbs) as well as with the specific cell surface receptors of cultured human IM-9 lymphocytes and liver membranes of pregnant rabbit. For all the tests used, the competition curves obtained using both hormone preparations were superimposable. The results suggest a close similarity in the immunological and biological (binding reaction) properties of native and recombinant hGH.

Antibodies, Monoclonal↗
PubMed1984

Uptake of lactoferrin by the liver. I. Role of the reticuloendothelial system as indicated by blockade experiments.

When 125I-labeled human lactoferrin was injected intravenously into mice it was rapidly taken up by the liver, where 75% of the dose was recovered after 15 minutes. The inhibition curve with unlabeled lactoferrin showed that the fates of labeled and unlabeled protein were similar and that the uptake capacity was saturable by milligram quantities of protein per gram of liver, suggesting the existence of numerous specific binding sites. The presence of these sites on the reticuloendothelial system was indicated by the blocking effect of dextran sulfate and latex particles. Fucoidin (fucan sulfate), which is used in the study of receptors specific for fucose, was found to inhibit the uptake of goat red blood cells as well as lactoferrin. Therefore, the inhibition exerted by fucoidin on the uptake of lactoferrin could be mediated by blockade of the reticuloendothelial system and not necessarily by competition for fucose receptors. These data indicate that the report by others that lactoferrin was taken up by a fucosyl receptor on hepatocytes is incorrect. The competition curve obtained when mouse lactoferrin was injected with its human homologue indicated that both proteins reacted with the same binding sites. However, a significant part of mouse lactoferrin was found to be taken up by the kidneys. Hepatic and renal uptakes were both reduced by prior injection of dextran sulfate.

Animals↗
PubMed1984

Particle-counting immunoassay of human somatotropin.

Human somatotropin was assayed by a novel automated nonradioisotopic technique, "particle-counting immunoassay," that requires a 45-min incubation and only 60 microL of 10-fold diluted sample. The principle of the assay is agglutination of antibody-coated latex particle by somatotropin, the reaction being measured by the (instrumented) counting of residual non-agglutinated particles. The dynamic range in serum extends from 0.2 to 40 micrograms/L. The between-assay CV was 12% for a concentration of 3.3 micrograms/L and 8.5% for 31.9 micrograms/L. The coefficient of correlation (r) with radioimmunoassay was 0.97. Curves for various dilutions of the macromolecular and monomeric forms of the hormone were not parallel.

Agglutination Tests↗
PubMed1984

Uptake of lactoferrin by the liver. II. Endocytosis by sinusoidal cells.

Because of the controversy on the cell type involved in lactoferrin (Lf) uptake by the liver we localized injected Lf by immunofluorescence, light microscopic autoradiography, and light and electron microscope cytochemistry with Lf-peroxidase conjugates. These conjugates were essentially taken up by their Lf moiety as shown by competition experiments. By any procedure, Lf was almost exclusively found in the sinusoidal cells as well as in the wall of the central veins. These data directly demonstrate that both endothelial and Kupffer cells are responsible for Lf uptake by the liver.

Animals↗
PubMed1984

Uptake of lactoferrin by the liver. III. Critical role of the protein moiety.

Three possible modes of recognition of human lactoferrin (Lf), which is avidly taken up by the mouse liver, were examined. First, Lf has terminal galactosyl residues for which a receptor exists on hepatocytes. However, when large amounts of Lf were injected with 125I-asialoorosomucoid, no inhibition of asialoorosomucoid uptake by the liver was observed. Second, Lf that exposes fucosyl residues could be recognized by the sugar-polyspecific receptor of liver sinusoidal cells. Digestion of Lf by fucosidase did not affect considerably the uptake of Lf. In addition, bovine Lf, which lacks fucosyl residues, was also avidly taken up by the liver, and this uptake was inhibited by human Lf. Mannan and horseradish peroxidase, which are recognized by the sugar-polyspecific receptor, did not inhibit Lf uptake. These data demonstrate that galactosyl and fucosyl residues are not essential for Lf recognition. Third, Lf could be recognized by its protein moiety. To investigate this possibility, we used two Lf derivatives with intact carbohydrate side chains, carbamylated Lf and the C-terminal half molecule of Lf. Carbamylation reduced the uptake of Lf and its competitive activity toward the uptake of 125I-Lf. The C-terminal fragment, like carbamylated Lf, had a much weaker competitive activity than intact Lf. Therefore, the integrity of the protein moiety of Lf was required for its effective uptake by the liver. As Lf is a cationic protein, competition experiments were also done with lysozyme, another cationic protein which in the form of dimer is taken up by the liver reticuloendothelial system. The strong inhibition by dimerized lysozyme suggests that the liver reticuloendothelial system has common binding sites for certain cationic proteins, as recently shown for isolated macrophages.

Animals↗
PubMed1982

The recognition by monoclonal antibodies of various portions of a major antigenic site of human growth hormone.

The reactivities of five mouse monoclonal antibodies against human growth hormone (hGH) were defined by either a competitive radioimmunoassay with insolubilized antibodies or by an agglutination-inhibition method with hGH-coated polystyrene particles. The five antibodies reacted significantly but to various degrees with human placental lactogen and at least three antibodies reacted with human prolactin and three synthetic peptides extending from residues 19 to 128, 73 to 128 and 98 and 128 of hGH. Four tested monoclonal antibodies failed to react with bovine growth hormone and with hGH oxidized by performic acid. The antibodies were further distinguished by their different reactions with hGH modified by reduction and alkylation or by adsorption on a polystyrene surface. The unique specificity of each antibody was confirmed for most of them by an agglutination method in which the agglutinating activity of hGH was tested on latex particles coated with various paired combinations of the monoclonal antibodies. The lack of agglutination with certain combinations suggested that the specificities of such a pair of antibodies overlapped each other. These results suggest that the sequences corresponding to the synthetic peptides participate in the structure of a major antigenic site of which various portions are recognized by the monoclonal antibodies.

Antibodies, Monoclonal↗
PubMed1982

The same region of human growth hormone is involved in its binding to various receptors.

The Fab fragments of three monoclonal antihuman GH (anti-hGH) antibodies, among five tested, inhibited the binding of the hormone to the receptors of the human lymphoid cell line IM-9 and liver membranes of the pregnant rabbit. The results were similar for the receptors of human lymphocytes and rabbit liver, suggesting that both receptors reacted with the same region of the hormone. The Fab fragments of the most inhibitory antibody also inhibited the down-regulation by the hormone of the receptors on human lymphocytes. The fragments of this antibody completely blocked the binding of the hormone to the receptors of the rabbit liver, despite the fact that this carries two or more classes of receptors. Therefore, all of these various receptors apparently interact with the same region of the hormone. Three synthetic peptides extending from residues 19-128, 73-128, and 98-128 failed to inhibit the binding of hGH to its lymphocyte or liver receptors however, these peptides reacted significantly with the monoclonal antibody, which was the strongest inhibitor of the interaction of hGH with the cellular receptor, confirming that the receptor-binding site of the hormone is in the amino-terminal part of the molecule and suggesting indirectly that the short sequences 98-128 participates in the constitution of the receptor-binding site of the hormone.

Amino Acid Sequence↗
PubMedDate not supplied

MAPAG: a computer program to construct 2- and 3-dimensional antigenic maps.

The contact area between an antibody (Ab) and the antigen (Ag) is called antigenic determinant or epitope. The first step in the characterization of an Ag by using monoclonal antibodies (MAb) is to map the relative distribution of the corresponding epitopes on the Ag surface. The computer program MAPAG has been devised to automatically construct antigenic maps. MAPAG is fed with a binary matrix of experimental data indicating the ability of paired MAb to bind or not simultaneously to the Ag. The program is interactive menu-driven and allows the user an easy data handling. MAPAG utilizes iterative processes to construct and to adjust the final map, which is graphically shown as a 2- or a 3-dimensional model. Additionally, the antigenic map obtained can be optionally modified by the user or readjusted by the program. The suitability of MAPAG was illustrated by running experimental data from literature and comparing antigenic maps constructed by the program with those elaborated by the investigators without the assistance of a computer. Furthermore, since some MAb could present negative allosteric effects leading to misinterpretation of data, MAPAG has been provided with an approximate reasoning module to solve such anomalous situations. Results indicated that the program can be successfully employed as a simple, fast and reliable antigenic model-builder.

Algorithms↗