Biomedical subjects
J Kopecek
Publications and source records attributed to J Kopecek.
Macrophages of athymic nude mice: Fc receptors, C receptors, phagocytic and pinocytic activities.
Expression of Fc receptors (FcR) for IgG1, IgG2A, IgG2B, IgM, IgA and IgE, binding of C3 and C5 complement components and phagocytic and pinocytic activities were determined in peritoneal and omental macrophages of nu/nu, nu/+ and +/+ Balb/c mice. nu/nu mice showed a higher proportion of FcR and complement receptor-bearing peritoneal macrophages along with a significantly higher phagocytic activity of peritoneal macrophages both in vitro and in vivo. Tests of pinocytic activity in these cells and phagocytic activity in omental phagocytes yielded similar results. We conclude that athymic mice compensate their immune defects by a higher phagocytic activity of their professional phagocytes and a higher expression of receptors mediating this process.
Targeting of N-(2-hydroxypropyl)methacrylamide copolymers to liver by incorporation of galactose residues.
Soluble synthetic polymers have potential as targetable carriers of pharmacological agents. Here we report that incorporation into poly[N-(2-hydroxypropyl)methacrylamide)] of an oligopeptide side-chain terminating in galactose enhanced the polymer's pinocytic uptake from the rat bloodstream by the liver. Within the liver lysosomes enzymic digestion led to the intracellular release of a drug analogue also bound to oligopeptide side-chains of the polymer.
Immunogenicity of N-(2-hydroxypropyl)-methacrylamide copolymers--potential hapten or drug carriers.
After repeated i.p. immunizations of mice with 10 micrograms of homopolymer poly (HPMA) no antibodies were detected by the ELISA test. Immunization with copolymer P-Acap-Leu-HMDA leads to a weak antibody response, while immunization with a copolymer with some side chains modified with ARS or FITC groups (P-Acap-Leu-HMDA-ARS or P-Acap-Leu-HMDA-FITC) leads to a significant antibody response detectable by PFC, ELISA and haemagglutination tests. Most of these antibodies are aimed against the modifying haptenic group, a smaller amount against side oligopeptide sequences of the carrier. Intensity of the antibody response depends on: 1) the antigen dose--the optimal dose was 10 micrograms: both the higher (100 micrograms) and the lower doses (1 and 0.1 micrograms) induced considerably lower antibody responses; 2) molar mass of the immunizing fractions--fractions of high molar mass induced up to five times higher responses than those of a low molar mass; 3) the bound haptenic group--the ARS-copolymers induced ten times lower response than the FITC-copolymers. We detected no difference between capacities of the H-2a, H-2b and H-2d haplotypes to react with anti-ARS antibodies after immunization with P-Acap-Leu-HMDA-ARS.
Synthesis of N-(2-hydroxypropyl)methacrylamide copolymers with antimicrobial activity.
Copolymers of N-(2-hydroxypropyl)methacrylamide which contained up to 20 mol % of p-nitrophenyl esters of N-methacryloylated oligopeptides, and of N-methacryloylaminophenoxyacetic acids (o-, m-, p-) have been prepared. The aminolyses of these polymers with tert. butylamine, ampicillin and 6-aminopenicillanic acid were kinetically characterized. Based on these results polymer bound ampicillin and polymer bound 6-aminopenicillanic acid were prepared. These preparations possessed antimicrobial activity; they inhibited the growth of Staphylococcus aureus 209 P.
An advantageous method for detection of Fc-receptors and for studying Fc-receptor-mediated phagocytosis.
A method employing fluorescent 2-hydroxyethyl methacrylate copolymer particles with dinitrophenyl haptenic group and anti-haptenic murine monoclonal antibodies of different isotypes is described for tracing Fc-receptors and for studies of phagocytosis via these cell surface structures. The possibility of quenching the fluorescence of non-phagocytosed particles enables us to distinguish clearly and reliably between internalized and cell surface-bound particles.
Phagocytosis of 2-hydroxyethylmethacrylate copolymer particles by different types of macrophages.
Phagocytic activity (for 2-hydroxyethylmethacrylate particles) in normal and thioglycollate- or proteose peptone-stimulated peritoneal macrophages, Peyer's patches macrophages, bone marrow macrophages and macrophage-like cell lines BWM, DCH 5, PU-5-lR and RAW 264.7, in suspension and after adherence to a substrate, was tested. Results of testing showed that 2-hydroxyethylmethacrylate particles are advantageous for following the phagocytic activity of the above cell types.
Degradation of side-chains of N-(2-hydroxypropyl)methacrylamide copolymers by lysosomal thiol-proteinases.
N-(2-Hydroxypropyl)methacrylamide copolymers bearing oligopeptide side-chains terminating in p-nitroaniline (NAp) were incubated with rat liver lysosomal enzymes in the presence of the thiol glutathione, and the rate of p-nitroaniline release was measured. Twelve of the 16 side-chains investigated were hydrolysed to release p-nitroaniline and in all but one case degradation was partially or totally inhibited by leupeptin. The effect of substrate concentration on the degradation of the most readily cleaved side-chain, -Ala-Gly-Val-Phe-NAp, was measured.
Polymers containing enzymatically degradable bonds. VI. Hydrophilic gels cleavable by chymotrypsin.
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Phagocytosis of human blood leukocytes: a simple micromethod.
A simple micromethod for testing human blood leukocyte phagocytosis employing synthetic hydrophilic particles based on 2-hydroxyethylmethacrylate is described. The normal level of phagocytosing leukocytes in healthy children was 20.1 +/- 2.7%, and in healthy adult donors 34.3 +/- 6.1%. The method was found suitable for routine testing in both clinical and laboratory practice.
Pinocytic uptake and intracellular degradation of N-(2-hydroxypropyl)methacrylamide copolymers. A potential drug delivery system.
Synthetic 125I-labelled N-(2-hydroxypropyl)methacrylamide copolymers containing four different, potentially degradable peptidyl side chains were incubated with rat visceral yolk sacs cultured in vitro. All copolymers were captured by fluid-phase pinocytosis and three of the side chains were susceptible to lysosomal hydrolysis, resulting in release of [125I]iodotyrosine back into the culture medium. Uptake and degradation was completely inhibited by 2,4-dinitrophenol. The thiol-proteinase inhibitor leupeptin did not affect the rate of pinocytosis, but caused different degrees of inhibition of hydrolysis depending on side chain composition.
Differences in phagocytic activity of methacrylate copolymer particles in normal and stimulated macrophages.
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[Duxon--a new Czechoslovak--made infusion solution--an experimental contribution to biological evaluation].
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Degradation of side chains of N-(2-hydroxypropyl) methacrylamide copolymers by lysosomal enzymes.
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Polymers containing enzymatically degradable bonds V. Hydrophilic polymers degradable by papain.
Copolymers of N-(2-hydroxypropyl)methacrylamide were prepared, in which synthetic polymer chains are joined by crosslinks containing oligopeptidic sequences degradable with papain, with the general structure P-(Gly)n-X-Y-NH-(CH2)6-NH-Y-X-(Gly)n-P (P is the polymer chain, n = 1,2; X...Phe, Val, Gly; Y...Lys, Gly, Tyr, Ala, Phe). The relationship between the structure of these polymeric substrates and their degradability with papain was investigated viscometrically. It was shown that -Phe-Lys- was the most suitable -X-Y- sequence. Extension of the oligopeptidic sequence by one amino acid residue causes a pronounced rise in the rate of cleavage of the polymeric substrates.
Immunosuppressive effects of a synthetic polymer poly N-(2-hydroxypropyl)methacrylamide (Duxon).
Poly N-(2-hydroxypropyl)methacrylamide (Duxon) was studied as a substitute of blood plasma and was found to prolong significantly the survival of semiallogeneic skin grafts in mice and rats and to reduce MLC reactions of human lymphocytes. It did not inhibit the growth of lymphocytes and the growth of the fibroblast-like (diploid) or epithelioid (heteroploid) cell lines in tissue culture.
[New types of synthetic infusion solutions. The effect of Duxon on cell proliferation in vitro].
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Comparison of the functional activity of normal and stimulated peritoneal exudate cells in suspension and after adherence to the substrate.
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