Search PubMed⌕ Search

Biomedical subjects

F Stirpe

Publications and source records attributed to F Stirpe.

At least 127 records · Page 7Linked to original sources
PubMed1985

Properties of volkensin, a toxic lectin from Adenia volkensii.

Volkensin, a highly toxic protein from the roots of Adenia volkensii (kilyambiti, kinoria), was purified by affinity chromatography on acid-treated Sepharose 6B. The toxin is a glycoprotein (Mr 62,000, neutral sugar content 5.74%) consisting of an A subunit (Mr 29,000) and of a B subunit (Mr 36,000) linked by disulfide and noncovalent bond(s). The amino acid, amino sugar, and neutral sugar composition of the protein were determined. Volkensin is a galactose-specific lectin and is a potent inhibitor of eukaryotic protein synthesis in whole cells as well as in a cell-free system (a rabbit reticulocyte lysate). The inhibitory and the lectin activities are functions of the A and B subunits, respectively. Volkensin can be included amongst the ricin-like toxins and resembles most closely modeccin, the toxin of Adenia digitata.

Amino Acids↗
PubMed1985

Characterization of a Saponaria officinalis seed ribosome-inactivating protein: immunoreactivity and sequence homologies.

A ribosome inactivating protein from Saponaria officinalis, SO-6, was purified and the N-terminus sequenced. The sequence shows extensive homology with Pokeweed antiviral protein, Pokeweed antiviral protein II, Pokeweed antiviral seed protein and dodecandrin. SDS gel electrophoresis in the Laemmli system revealed two bands of similar intensities with a smear between them, probably an artifact due to the high pI of the protein. Use of a harsher denaturing gel system resulted in one band in electrophoresis. Immune antisera was raised in rabbits against this protein and it cross reacted with other proteins (SO-5, SO-8 and SO-9) from seeds of Saponaria officinalis, but not with gelonin, Momordica charantia inhibitor and dianthin 32.

Amino Acid Sequence↗
PubMed1985

Modification of the carbohydrate in ricin with metaperiodate-cyanoborohydride mixtures. Effects on toxicity and in vivo distribution.

Attempts to target antibody-ricin conjugates (immunotoxins) to designated cell types in vivo may be thwarted by their rapid clearance by hepatic reticuloendothelial cells which have receptors that recognise oligosaccharide side chains on the toxin. The B-chain of ricin contains high mannose type oligosaccharides and the A-chain contains a complex unit (GlcNAc)2-Fuc-Xyl-(Man)4-6, all of which potentially could be recognised by the reticuloendothelial system. Treatment of ricin with a mixture of sodium metaperiodate and sodium cyanoborohydride at pH 3.5 resulted in oxidative cleavage of the carbohydrates and reduction of the aldehyde groups thus formed to primary alcohols. By conducting the modification procedure at acidic pH, both the possibility of Schiff's base formation between the aldehyde groups and amino groups in the protein and the possibility of non-specific oxidation of amino acids were minimised. The extent of the carbohydrate modification depended on the duration of treatment, resulting maximally in the destruction of 13 of the 18 mannose residues and of all xylose and fucose. The toxicity of the modified toxin to cells in culture declined by up to 90% as the carbohydrate was destroyed. This was not due to a reduced ability of the B-chain to bind to cells or of the A-chain to inactivate ribosomes. In contrast to the in vitro results, the toxicity of the modified toxin to mice and rats was elevated by up to fourfold. The modification greatly reduced the clearance of the toxin by non-parenchymal cells in the liver and prevented the damage to hepatic Kupffer and sinusoidal cells and to the red pulp of the spleen that is inflicted by the native toxin. The elevated toxicity to animals appears to be because the modified toxin evades the reticuloendothelial system and persists in the bloodstream for longer periods, thus resulting in lethal damage to vital tissues in the animal at lower dosage. The results suggest that immunotoxins prepared from modified ricin would not be readily cleared by the reticuloendothelial system and so be more effective at killing their target cells.

Amino Acids↗
PubMed1985

The immunological activity of plant toxins used in the preparation of immunotoxins--II. The immunodepressive activity of gelonin.

The immunological activity of Gelonin, a 30,000 dalton plant protein possessing close similarity to Ricin chain A as a protein synthesis inhibitor which may be of interest for the preparation of antibody-toxin conjugates, was studied in mice. At in vitro concentrations not affecting baseline radioactivity uptake, this substance reduced mitogen responses with the following order of sensitivity PHA less than ConA less than LPS. In microgram/ml concentrations it also markedly reduced macrophage-dependent cytotoxicity while not affecting NK activity. Macrophagic (but not NK) cytotoxicity and mitogen responses were similarly depressed after in vivo treatment. When given before (but not after) stimulus, Gelonin also reduced the primary responses to a T-dependent and, although to a lower degree, to a T-independent antigen, and decreased resistance to allogeneic tumor grafts and L. monocytogenes challenges. The immunopharmacological activity of this and similar substances should be considered in the design of antibody-toxin conjugates and in the evaluation of their therapeutic activity.

Animals↗
PubMed1985

An immunotoxin composed of monoclonal anti-Thy 1.1 antibody and a ribosome-inactivating protein from Saponaria officinalis: potent antitumor effects in vitro and in vivo.

The ribosome-inactivating protein saporin, from Saponaria officinalis, was coupled by a disulfide bond to monoclonal anti-Thy 1.1 antibody (OX7) and to its F(ab')2 fragment. The immunotoxins were at least as toxic as the plant toxin ricin to the Thy 1.1-expressing cell lines AKR-A and BW5147 in tissue culture. They reduced the rate at which the cells incorporated [3H]leucine into protein by 50% at cell concentrations of 1.5-3 X 10(-11) and 3 X 10(-12) M, respectively. The toxic effect was specific. No toxicity was seen when the immunotoxins were applied to Thy 1.2-expressing EL 4 lymphoma cells at 3 X 10(-8) M, and a control immunotoxin made from an antibody (R10) of irrelevant specificity was without effect on AKA-A cells. Further, the treatment of spleen cells from AKR mice with OX7-saporin at 10(-8) M abolished their response to the T-lymphocyte mitogen concanavalin A, without impairing their response to the B-lymphocyte mitogen lipopolysaccharide. A single iv injection of OX7-saporin into nu/nu randombred mice bearing peritoneal AKR-A lymphoma cells prolonged the survival time of the animals by an extent corresponding to that expected if 99.999% of the tumor cells had been eradicated by the immunotoxin. None of the control materials (unconjugated OX7, unconjugated saporin, OX7 plus saporin, or R10-saporin) delayed tumor growth. The OX7 F(ab')2-saporin conjugate was also highly effective as an antitumor agent, although significantly less so than the conjugate made with intact OX7. Unexpectedly, the acute toxicity of saporin to mice (median lethal dose = 6.8 mg/kg) was elevated eightfold to sixteenfold by conjugation to OX7, R10, or OX7 F(ab')2. Histologic examination of recipients of the immunotoxin revealed gross damage to hepatic parenchymal cells and to the white pulp of the spleen, neither of which was caused by unconjugated saporin. Ricin A-chain coupled to OX7 antibody was one hundredfold to one thousandfold less effective than OX7-saporin as an antitumor agent in vivo, although the two immunotoxins were equally cytotoxic to AKR-A cells in vitro.

AKR murine leukemia virus↗
PubMed1984

Cytoxicity of erythrocyte ghosts loaded with ribosome-inactivating proteins following fusion with CHO cells.

The ribosome-inactivating proteins gelonin, Momordica charantia inhibitor, pokeweed antiviral protein, and one from Saponaria officinalis were enclosed in human erythrocyte ghosts. The proteins once trapped in ghosts and fused with CHO cells inhibited colony formation at concentrations of approximately 1 ng/ml (3 X 10(-11) M), whereas the free proteins only had an effect at concentrations of greater than 1 microgram/ml.

Animals↗
PubMed1983

Ribosome-inactivating proteins from the seeds of Saponaria officinalis L. (soapwort), of Agrostemma githago L. (corn cockle) and of Asparagus officinalis L. (asparagus), and from the latex of Hura crepitans L. (sandbox tree).

Ribosome-inactivating proteins, similar to those already known [Barbieri & Stirpe (1982) Cancer Surveys 1, 489-520] were purified from the seeds of Saponaria officinalis (two proteins), of Agrostemma githago (three proteins), and of Asparagus officinalis (three proteins), and from the latex of Hura crepitans (one protein). The yield ranged from 8 to 400 mg/100 g of starting material. All proteins have an Mr of approx. 30000 and an alkaline isoelectric point. Their sugar content varies from 0 (proteins from S. officinalis) to 40% (protein from H. crepitans). The ribosome-inactivating proteins inhibit protein synthesis by rabbit reticulocyte lysate, the ID50 (concentration giving 50% inhibition) ranging from 1 ng/ml (a protein from S. officinalis) to 18 ng/ml (a protein from A. githago). Those which were tested (the proteins from S. officinalis and from A. githago) also inhibit polymerization of phenylalanine by isolated ribosomes, acting in an apparently catalytic manner. The protein from H. crepitans inhibited protein synthesis by HeLa cells, with an ID50 of 4 micrograms/ml, whereas the proteins from S. officinalis and from A. githago had an ID50 of more than 50-100 micrograms/ml. The ribosome-inactivating proteins from S. officinalis and from A. githago reduced the number of local lesions by tobacco-mosaic virus in the leaves of Nicotiana glutinosa.

Amino Acids↗
PubMed1983

Purification and properties of two lectins from the latex of the euphorbiaceous plants Hura crepitans L. (sand-box tree) and Euphorbia characias L. (Mediterranean spurge).

1. From the latex of two members of the plant family Euphorbiaceae, Hura crepitans L. (sand-box tree) and Euphorbia characias L. (Mediterranean spurge), two lectins were purified by affinity chromatography on acid-treated Sepharose 6B followed by elution with D-galactose. 2. The lectin from E. characias is a single molecular species with Mr 80 000, made up of two identical subunits with Mr 40 000, and is a glycoprotein containing 11% carbohydrate. 3. The lectin from H. creptians appears as a mixture of three isolectins with Mr 140 000, consisting of four different subunits with Mr values 37 500, 35 500, 31 000, and 29 000. 4. Both lectins have haemagglutinating activity, with no specificity for human blood groups. The haemagglutinating activity is inhibited by D-galactose and by galactose-containing oligosaccharides. 5. The lectin from H. crepitans is mitogenic to human T-, but not to B-, lymphocytes. The latex of E. characias is mitogenic to T- and, to a lesser extent, to B-, lymphocytes, but the purified E. characias lectin has no mitogenic activity. 6. The lectin from H. crepitans, but not that from E. characias, inhibits protein synthesis by a rabbit reticulocyte lysate.

Amino Acids↗
PubMed1983

Cytotoxicity acquired by ribosome-inactivating proteins carried by reconstituted Sendai virus envelopes.

Association of the ribosome-inactivating proteins (RIPs): pokeweed antiviral protein (PAP), gelonin, Momordica charantia inhibitor (MCI), with reconstituted Sendai virus envelopes (RSVE) was obtained without detectable loss of activities either of RIPs or of viral envelope glycoproteins. RIPs are inactive towards intact cells, but, once encapsulated in RSVE, they become cytotoxic. The concentration of RSVE-associated PAP, which causes 50% inhibition of protein synthesis by Friend erythroleukemic cells, is 0.5 ng/ml. Substances capable to inhibit the viral activities block the acquired cytotoxicity of RIPs associated to RSVE.

Animals↗
PubMed1983

Mistletoe toxicity.

Explore the source record for details and available documents.

Chemical and Drug Induced Liver Injury↗
PubMed1983

The immunomodulatory activity of the plant proteins Momordica charantia inhibitor and pokeweed antiviral protein.

The immunological activity of Momordica Charantia inhibitor (MCI) and of Pokeweed antiviral protein (PAP-S), 30,000 daltons plant proteins possessing close similarity to Ricin A chain as inhibitor of protein synthesis, was investigated in mice. In vivo, single nontoxic injections of microgram amount of these substances delayed H2-incompatible skin allograft rejection, splenocyte responsiveness to ConA and PHA, but not to LPS, and abrogated the PFC response to a T-dependent (SRBC) antigen while totally sparing that to a T-independent (S III) stimulus. Injection of these substances could also reduce NK cell activity while increasing macrophage-mediated spontaneous cytotoxicity. In vitro, MCI and PAP-S at non-cytotoxic concentrations inhibited lymphoid cell responsiveness to PHA and ConA, but not to LPS, and markedly enhanced macrophage-dependent cytotoxicity.

Animals↗
PubMed1982

Properties of the ribosome-inactivating proteins gelonin, Momordica charantia inhibitor, and dianthins.

The amino acid and sugar compositions of four ribosome-inactivating proteins (gelonin, Momordica charantia inhibitor, dianthin 30 and dianthin 32) were determined. The proteins are all basic glycoproteins (pI greater than 8) containing mannose (more abundant in gelonin), glucose, xylose, fucose (absent from gelonin) and glucosamine. The ribosome-inactivating properties of the proteins examined are not modified by pretreatment with N-ethylmaleimide. Precipitating and inactivating antibodies can be raised against ribosome-inactivating proteins; a weak cross-reaction was observed only between dianthin 30 and dianthin 32.

Amino Acids↗
PubMed1982

Isolation and characterization of viscumin, a toxic lectin from Viscum album L. (mistletoe).

A toxic protein, viscumin, was isolated from extracts of mistletoe by affinity chromatography on acid-treated Sepharose 4B. Viscumin was selectively bound to the column and could be eluted with lactose. It migrated in polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate corresponding to Mr = 60,000. In addition, two bands migrating corresponding to Mr = 29,000 and 32,000 were found. After treatment with 2-mercaptoethanol, only 2 bands (Mr = 29,000 and 34,000) were found. Apparently, viscumin consists of two chains which, in some of the molecules, are disulfide-linked. Protection experiments with antiserum against viscumin indicated that the major part of the cytotoxic activity in mistletoe extracts is due to viscumin. Gel filtration experiments on Sephacryl 200 indicated that, at low concentrations, viscumin occurs as a monomer and at higher concentrations as a dimer. Viscumin was found to inhibit protein synthesis in cell-free systems. When the two constituent peptide chains of viscumin were eluted from polyacrylamide gels and tested for ability to inhibit cell-free protein synthesis, this property was found to be associated with the fastest migrating chain, here denoted the A chain. The heavier chain was denoted the B chain. The A chain was found to inhibit protein synthesis by inactivating the ribosomes catalytically. Reconstitution experiments with isolated ribosomal subunits from untreated and A chain-treated ribosomes showed that the 60 S ribosomal subunit was selectively inactivated.

Animals↗
PubMed1982

Action of viscumin, a toxic lectin from mistletoe, on cells in culture.

A toxin from mistletoe, viscumin, inhibited the incorporation of leucine in cells more rapidly than the incorporation of uridine and thymidine, indicating that the toxins act by inhibiting cellular protein synthesis. The presence of galactose, lactose, and melibiose in the medium protected cells against viscumin. The sensitivity to viscumin of different cell lines differed considerably. The cytotoxic properties of viscumin were compared to those of the related toxins abrin, ricin, and modeccin. The profile of the sensitivity of a panel of cell lines to viscumin was dissimilar from those obtained with abrin and modeccin. Cell lines selected for resistance to modeccin and ricin were fully sensitive to viscumin. Ca2+ was required for viscumin to express its toxic effect. In contrast, the Ca2+ ionophore A23187 protected the cells against viscumin. Whereas the sensitivity of cells to viscumin did not vary much between pH 7 and 9, the cells were much less sensitive at pH 6. Also, cells treated with the two metabolic inhibitors, 2-deoxyglucose and NaN3, were insensitive to the toxins. Viscumin taken up by cells in the absence of Ca2+ or at pH 6 was able to intoxicate the cells when Ca2+ was added or when the pH was adjusted to neutrality. In contrast, cells exposed to viscumin in the presence of 2-deoxyglucose and NaN3, which inhibit endocytosis, were not intoxicated when the cells were treated with antiviscumin and then transferred to normal medium. The results indicate that endocytosis is involved in the entry of viscumin.

Animals↗
PubMed1982

Purification and partial characterization of another form of the antiviral protein from the seeds of Phytolacca americana L. (pokeweed).

1. The pokeweed antiviral protein, previously identified in two forms (PAP and PAP II) in the leaves of Phytolacca americana (pokeweed) [Obrig. Irvin & Hardesty (1973) Arch. Biochem. Biophys. 155, 278-289; Irvin, Kelly & Robertus (1980) Arch. Biochem. Biophys. 200, 418-425] is a protein that prevents replication of several viruses and inactivates ribosomes, thus inhibiting protein synthesis. 2. PAP is present in several forms in the seeds of pokeweed. One of them, which we propose to call 'pokeweed antiviral protein from seeds' (PAP-S) was purified in high yield (180 mg per 100 g of seeds) by chromatography on CM-cellulose, has mol.wt. 30 000, and is similar to, but not identical with. PAP and PAP II. 3. PAP-S inhibits protein synthesis in a rabbit reticulocyte lysate with an ID50 (concentration giving 50% inhibition) of 1.1 ng/ml (3.6 x 10(-11) M), but has much less effect on protein synthesis by whole cells, with an ID50 of 1 mg/ml (3.3 x 10(-5) M), and inhibits replication of herpes simplex virus type 1.

Amino Acids↗
PubMed1982

Effect of ribosome-inactivating proteins on virus-infected cells. Inhibition of virus multiplication and of protein synthesis.

HEp-2 cells were infected with herpes simplex virus-1 (HSV-1) or with polio-virus I in the presence of plant proteins which inactivate ribosomes in cell-free systems, while exerting scarce effect on whole cells. Ribosome-inactivating proteins used were gelonin, from seeds of Gelonium multiflorum, an inhibitor from the seeds of Momordica charantia, dianthin 32, from the leaves of Dianthus caryophyllus (carnation), and PAP-S, from the seeds of Phytolacca americana (pokeweed). All proteins tested had the following effects: 1. They reduced viral yield; 2. They decreased HSV-1 plaque-forming efficiency; 3. They inhibited protein synthesis more in infected than in uninfected cells. These results strongly suggest that ribosome-inactivating proteins impair viral replication by inhibiting protein synthesis in virus-infected cells, in which presumably they enter more easily than in uninfected cells.

Cells, Cultured↗