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Biomedical subjects

G R Adolf

Publications and source records attributed to G R Adolf.

At least 55 records · Page 3Linked to original sources
PubMed1988

Cloning of human lysozyme gene and expression in the yeast Saccharomyces cerevisiae.

cDNA clones encoding human lysozyme were isolated from a human histiocytic cell line (U-937) and a human placenta cDNA library. The clones, ranging in size from 0.5 to 0.75 kb, were identified by direct hybridization with synthetic oligodeoxynucleotides. The nucleotide sequence coding for the entire protein was determined. The derived amino acid sequence has 100% homology with the published amino acid (aa) sequence; the leader sequence codes for 18 aa. Expression and secretion of human lysozyme in Saccharomyces cerevisiae was achieved by placing the cloned cDNA under the control of a yeast gene promoter (ADH1) and the alpha-factor peptide leader sequence.

Amino Acid Sequence↗
PubMed1987

Characterization of the human erythrocyte complement receptor CR1 (C3b receptor) by epitope mapping.

Monoclonal antibodies and an anti-idiotypic serum against human complement receptor CR1 (C3b receptor, immune adherence receptor) were used to identify CR1 and some of its proteolytic fragments by an immunoblotting technique. The anti-idiotypic serum had a specificity for the C3b-binding site, as could be shown by its cross-reactivity with complement factor H. The monoclonal antibodies GARP-4 and GARP-37 were specific for epitopes located nearby the ligand-binding site, because they blocked the immune adherence reaction. For the immunoblotting technique, it was essential to use non-reducing conditions, since reduction of CR1 destroyed the epitopes. Therefore, mainly large (disulphide-linked) fragments of CR1 were obtained. A chymotryptic fragment of Mr 56,000 identified by GARP-4, was the smallest cleavage product to be associated with the C3b-binding domain. Different proteases gave CR1 degradation products of similar Mr, indicating the presence of distinct domains, three of which had a Mr approximately 38,000. A schematic model of CR1 substructure was deduced from the epitope mapping data.

Antibodies, Monoclonal↗
PubMed1987

Structure and expression in Escherichia coli of canine interferon-alpha genes.

Using a human interferon-alpha (IFN-alpha) cDNA probe, several recombinant phages containing type I IFN genes were isolated from a canine genomic library. One of these phages contains two complete CaIFN-alpha genes with identical coding sequences, and a second one a slightly different IFN-alpha gene. The IFN-alpha protein sequences contain six cysteine residues as well as two or three potential N-glycosylation sites. Expression of mature CaIFN-alpha 1 in E. coli results in antiviral activity on dog cells. Genomic analysis using an equine IFN-omega probe and DNA sequencing suggests the deletion of IFN-omega genes from canine genome.

Amino Acid Sequence↗
PubMed1987

Antigenic structure of human interferon omega 1 (interferon alpha II1): comparison with other human interferons.

Recombinant human interferon (IFN) omega 1 (IFN-alpha II1) was compared with other human IFNs with regard to antigenic structure, using polyclonal and monoclonal antibodies. Antisera to recombinant IFN-alpha 1 or IFN-alpha 2c cross-neutralized the antiviral activity of other species of IFN-alpha, but not that of IFN-omega 1. Antisera to IFN-beta or IFN-gamma likewise had no effect on IFN-omega 1. Conversely, antiserum to IFN-omega 1 did not neutralize any of the other IFNs examined (IFN-alpha 1, -alpha 2c, -alpha 10, -alpha B, -alpha F, -beta, -gamma). None of four monoclonal antibodies to IFN-alpha, including a broad-spectrum antibody that neutralized all five recombinant species of IFN-alpha tested, had any effect on the biological activity of IFN-omega 1. Various immunoassays for IFN-alpha did not recognize IFN-omega 1. In immunoprecipitation experiments, radiolabelled IFN-alpha 2c was not precipitated by antiserum to IFN-omega 1. In contrast, antisera to human leukocyte IFN or 'lymphoblastoid' IFN did neutralize IFN-omega 1, although with titres significantly lower than those towards the immunogen or recombinant species of IFN-alpha. In conclusion, human IFN-omega 1 is antigenically different from human IFN-alpha, IFN-beta or IFN-gamma, but is a component of 'natural' mixtures of IFN species produced by virus-induced leukocytes or Burkitt's lymphoma cells.

Amino Acid Sequence↗
PubMed1986

Molecular cloning and expression in Escherichia coli of equine type I interferons.

Using human interferon-alpha 2 (IFN-alpha 2) and IFN-beta DNA to probe an equine genomic library we isolated recombinant phages containing genes for equine interferon-alpha (EqIFN-alpha), interferon-beta (EqIFN-beta), and interferon-omega (EqIFN-omega). Sequence and hybridization analyses of these genes reveal that the equine genome contains gene families of each of these three type I interferon classes. The mature proteins of EqIFN-alpha are 71-77% homologous to human IFN-alpha polypeptides, and, when expressed in E. coli, possess antiviral activity on both equine and human cells. By contrast, EqIFN-beta is only 59% homologous to its human counterpart and shows activity only on equine cells.

Animals↗
PubMed1985

Structure and effects of interferon-gamma.

This brief overview of the structure of interferon-gamma (IFN-gamma) and its biological activities emphasizes recent results with recombinant human and murine IFNs and compares this type II IFN with the type I IFN-alpha and IFN-beta. The amino acid sequence of human IFN-gamma is presented. Its antiviral activity and cytostatic or cytotoxic effects are described. Its effects on the immune system, including enhancement of membrane antigen expression, macrophage activation, effects on B lymphocytes, enhancement of natural killer cell activity and the interaction between IFN-gamma and interleukin 2 are discussed. The evidence for in vivo antitumour activity of IFN-gamma is reviewed and the problem of the molecular mechanisms of IFN action is examined.

Amino Acid Sequence↗
PubMed1985

Immunoassay for the quantitation of human leukocyte interferon.

A sensitive nonradioactive immunoassay based on monoclonal antibodies was developed. A number of monoclonal hybridomas secreting antibodies against human leukocyte interferon (IFN alpha) were generated using mice immunized with purified lymphoblastoid IFN. Although the binding of antibody to IFN alpha was used as one criterium of selection, all antibodies found can neutralize its antiviral activity. A pair of antibodies binding to different regions of IFN alpha was identified. These were incorporated into sensitive sandwich assays of IFN alpha. A microtiter plate assay - using horse radish peroxidase as marker enzyme - is able to detect IFN alpha at a concentration of 30 IU/ml within 5 h. An overnight tube assay can detect approximately 30 pg IFN alpha 2 or 3 IU per ml solution. The 5-h ELISA (enzyme-linked immunosorbent assay) is well suited for the monitoring of the recovery of rIFN alpha from recombinant organism, during purification and refinement of the protein to a therapeutic drug.

Antibodies, Monoclonal↗
PubMed1984

Lymphokines produced by herpesvirus-transformed marmoset monkey lymphoid cell lines. I. Characterization of a constitutively produced interferon.

Conditioned media from cultures of marmoset monkey T-lymphoid cell lines transformed by Herpesvirus saimiri or Herpesvirus ateles were found to contain interferon (IFN) activity. Titers between individual cell lines varied by a factor of 100; large amounts (up to 10(5) units/ml, assayed on human cells) were produced in one of the cell lines. IFN production was enhanced by the diterpene tumor promoters, TPA and mezerein, but not by classical T-cell mitogens. The IFN resembles human IFN-gamma by the following criteria: lability at pH 2, stability against 2-mercaptoethanol, cross-species activity, shape of dose-response curves, and molecular weight determined by size-exclusion chromatography (50,000-55,000). Its activity was not inhibited, however, by antiserum against human IFN-gamma or antisera against human IFN-alpha or IFN-beta.

Animals↗
PubMed1982

Production of monoclonal antibodies to human IFN-alpha and their use for analysis of the antigenic composition of various natural interferons.

We have established three hybridoma cell lines (designated EBI-1, -2, and -3) secreting immunoglobulin G (IgG) antibodies specific for human alpha-interferon (HuIFN-alpha) by fusing P3X63Ag8.6.5.3 myeloma cells with spleen cells of mice immunized with human "lymphoblastoid" (NC-37) interferon (IFN). The antibodies were used to study the antigenic diversity of a number of IFNs derived from blood leukocytes and lymphoid cell lines. When incubated with 10 U of IFN, EBI-1 antibodies completely neutralized the antiviral activity of HuIFN-alpha rA produced in E coli as well as of IFN produced spontaneously in several lymphoblastoid cell lines. In contrast, only 60-70% of alpha-interferon (IFN-alpha) induced in leukocytes, lymphoma, or lymphoblastoid cell lines by Sendai virus or poly (I):(C) could be neutralized by the antibody; furthermore, IFN-alpha produced spontaneously in bromodeoxyuridine-treated lymphoma cells was also only partially neutralized. EBI-2 and EBI-3 antibodies were tested against a smaller set of samples and found to react similarly. Our results indicate that human IFN-alpha subtypes can be grouped into at least two antigenically distinguishable subsets, one of which being represented by IFN-alpha A, and that these subsets can be expressed differently under different conditions in a given cell type.

Animals↗
PubMed1982

Spontaneous production of alpha- and beta-interferon in human lymphoblastoid and lymphoma cell lines.

A large number of human haematopoietic cell lines was examined for spontaneous production of interferon. Unconcentrated culture supernatants from 70 out of 71 B-lymphoblastoid cell lines contained considerable amounts of interferon (median titer 22 units per ml); a few lines produced more than 100 units/ml with peak values up to 500 units/ml. In contrast, only one B-lymphoma line out of 18 genuine lymphoma, myeloma, and leukaemia cell lines tested spontaneously produced small amounts of interferon. Following treatment with 5-bromodeoxyuridine (BrdUrd), interferon was produced without further induction in most B-lymphoid cell lines, but not in any of the non-B, non-T, T-lymphoid or myeloid lines examined. Modulation of spontaneous interferon production by chemicals (sodium butyrate, dexamethasone, dimethylsulfoxide, a phorbol ester, and BrdUrd) was studied in more detail in three B-lymphoblastoid and four B-lymphoma cell lines. The patterns of responses observed were different for the action of different chemicals on a given cell line as well as between lymphoblastoid and lymphoma lines in general; furthermore, several lines of evidence suggest that chemicals can differentially influence spontaneous and virus-induced interferon production in an given cell line. The composition of spontaneously produced interferon was analysed using antisera specific for HuIFN-alpha and HuIFN-beta. Interferons produced by untreated as well as BrdUrd-treated lymphoblastoid cells contained more than 95 per cent IFN-alpha, whereas BrdUrd-treated lymphoma cells produced IFN-alpha as well as minor amounts (cell lines Namalwa and NC-37) or even over 90 per cent of IFN-beta (Daudi).

Bromodeoxyuridine↗
PubMed1982

Interferon production in human hematopoietic cell lines: response to chemicals and characterization of interferons.

We have surveyed interferon production and its modulation by 5-bromodeoxyuridine, butyrate, dexamethasone, dimethylsulfoxide and tetradecanoylphorbolacetate in 20 human hematopoietic cell lines derived from leukemias, lymphomas, myelomas and normal leukocytes, representing various maturation stages of lymphoid and myeloid cells. Sendai virus-induced interferon production was enhanced by at least one of the chemicals in 13 out of 14 B-type lymphoid cell lines, whereas no enhancement was observed in any of the non-B, non-T-, T-lymphoid, or myeloid cell lines tested. Interferon produced by 11 cell lines was partially characterized using antisera specific for HuIFN-alpha and HuIFN-beta. Six cell lines produced both IFN-alpha and IFN-beta, two lines produced by IFN-alpha and three lines (including both T-cell lines tested) produced only IFN-beta. In all cases examined, enhancement by chemicals of total interferon yields was due to selective stimulation of production of IFN-alpha. Poly (I):(C) induction of interferon was studied in a number of B-cell lines. In general, a similar pattern of IFN-alpha and -beta synthesis was observed as in virus-induced cells, but the proportion of IFN-beta was relatively smaller. Treatment with butyrate enhanced interferon production to a similar extent accompanied by a similar shift in composition as in virus-induced cells. Our results demonstrate that even in closely related cell types, production of IFN-alpha and IFN-beta can be regulated differently in response to the same inducer. In a single cell type in response to a single inducer, expression of IFN-alpha and IFN-beta can be differentially affected by chemicals.

Bromodeoxyuridine↗
PubMed1981

Accumulation of histone H1(0) during chemically induced differentiation of murine neuroblastoma cells.

A basic nuclear protein with properties similar to lysine-rich histones accumulates during differentiation in vitro of C1300 murine neuroblastoma cells (clone Neuro-2a) induced by either n-butyrate, dimethyl sulfoxide, hexamethylene bisacetamide or 1,6-dibutyryl-adenosine 3',5'-monophosphate. A protein with the same electrophoretic properties, present in adult mouse brain cell nuclei in amounts similar to those in the induced neuroblastoma cells, has been characterized as histone H1(0). Digestion of the two proteins with Staphylococcus aureus V8 protease gives identical peptide maps (sharing no common peptides with those of histones H1A or H1B), which indicates that the induced protein qualifies as H1. Accumulation of histone H1(0) in neuroblastoma cell nuclei accompanies shutoff of DNA synthesis and transgression of the cells into a G1-phase resting state. Upon removal of n-butyrate the cells resume proliferation and their H1 content decreases, indicating an association of H1 with the resting state during which differentiated cellular functions are expressed.

Acetamides↗
PubMed1980

Tumour-promoting phorbol esters inhibit DNA synthesis and enhance virus-induced interferon production in a human lymphoma cell line.

12-O-tetradecanoyl-phorbol-13-acetate (TPA), a potent tumour promoter, was tested for its effects on the proliferation of the human Burkitt lymphoma cell line, Namalwa, and the synthesis of interferon by these cells. At nanomolar concentrations, TPA blocked thymidine incorporation into cellular DNA by more than 90% within 24 h. TPA-treated cells produced about 20-fold more interferon in response to Sendai virus than did untreated controls and simultaneous treatment with TPA and sodium n-butyrate gave a further two- to three-fold enhancement. Neither of these effects of TPA was reversed on removal of the compound; furthermore, exposure of Namalwa cells to TPA for only 1 h was sufficient for full activity. 4-O-methyl-TPA, a compound only marginally active as a tumour promoter, showed effects similar to TPA, but only at concentrations 300-fold higher. In contrast to its effects on Namalwa cells, TPA did not affect synthesis of interferon in response to Sendai virus in two other Burkitt lymphoma lines (Raji and Daudi) nor in the Epstein-Barr virus (EBV)-negative lymphoma line, BJAB; it inhibited interferon production in the human myeloid cell line, HL-60.

Burkitt Lymphoma↗
PubMed1979

Glucocorticoid hormones inhibit DNA synthesis and enhance interferon production in human lymphoid cell line.

Although buffy coat leukocytes have been the prime source of human interferon, cells of the Burkitt lymphoma line Namalwa are increasingly used for the large scale production of interferon. On induction with Sendai virus or Newcastle disease virus, Namalwa cells produce a substantial quantity of interferon which contains predominantly the leukocyte antigenic species and minor amounts of fibroblast-type interferon. We have recently demonstrated that inducers of erythropoietic differentiation in Friend cells are able to enhance interferon synthesis in Namalwa cells when added to cultures larger than or equal to 24 h before interferon induction by Sendai virus. The most potent compounds, n-butyrate, stimulated interferon production about 30-fold and has also been independelty described by others. All active compounds inhibited DNA synthesis in Namalwa cells and the extent of inhibition apparently paralleled the stimulatory potency of the respective compound. Induction of differentiation of Friend cells can be antagonised by various steroid hormones, which by themselves have no measurable effects on these cells. In contrast, we report here that glucocorticoid hormones inhibit DNA synthesis in Namalwa cells and augment Sendai virus-induced interferon synthesis.

Burkitt Lymphoma↗