Search PubMed⌕ Search

Biomedical subjects

R K Zahn

Publications and source records attributed to R K Zahn.

At least 73 records · Page 4Linked to original sources
PubMed1983

DNA damage by PAH and repair in a marine sponge.

The sponge Tethya lyncurium from the Northern Adriatic has been used as an experimental species. A method is outlined for preparation of DNA which yields a highly purified DNA with a double-strand (ds) molecular weight of 25 M-dalton between single-strand (ss) breaks, which when properly damaged can be cut opposite to ss-breaks with nuclease S1. The molecular weights of the resulting ds-DNA pieces and their distribution has been evaluated by electron microscope photographs. Sponges exposed to benzo[a]pyrene (BaP) in the dark only incorporate BaP-derivatives (BaPD) in small amounts, if any. However, in the presence of light, derivatization to BaP derivatives enables effective coupling to occur, as shown previously (R.K. Zahn et al., 1981). Sponges were exposed to radiolabeled BaP in the presence of light. Coupling of BaPD to the DNA as well as the induction of ss-breaks were measured. Light-mediated coupling is concentration dependent from 0.01-20 ppb BaP with a correlation coefficient of r = 0.84. Under conditions of possible repair, ss-breaks completely disappear from sponge DNA in the course of three weeks while a substantial fraction of the BaP derivatives persists. Double label experiments show that substantial DNA synthesis occurs during this time. Pollution causes a decrease of the molecular weight of unnicked DNA, re-incubation in clean water an increase. A DNA species of 24 M-dalton seems to play a critical role. If its percentage in the DNA population drops below a critical level, recovery is not longer possible. DNA damage by PAH and repair in sponges seems to differ from that of most eucaryotes.

Animals↗
PubMed1983

Activity of benzo[a]pyrene monooxygenase in fish from the Sava River, Yugoslavia: correlation with pollution.

Benzo[a]pyrene monooxygenase (B[a]PMO) activity in non-migratory fish from a given river segment is highly correlated to the recent pollutional history of that part of the river. The enzyme activity level can serve as a relevant measure for the harmful pollutant potential in aquatic ecosystems. Caged experimental fish exposed for about 10 days in river segments show BaPMO activity changes with the same predictive validity as that of the natural population.

Animals↗
PubMed1983

Potentiation of the cytostatic effect of bleomycin on L5178y mouse lymphoma cells by pepleomycin.

Bleomycin (BLM) and pepleomycin (PEP) are two chemically related glycopeptide antitumor antibiotics which differ in their terminal residues only. Studying the growth-inhibitory potencies of BLM (clinical mixture), BLM-A2, BLM-B2 and PEP in the L5178y mouse lymphoma cell culture system, it was elucidated that the slopes of the dose-response curves at the ED50 concentration (around 1 microgram/ml) were steeper for PEP than for BLM. This result together with cytotoxicity determinations revealed a cytostatic action of PEP within a closer concentration range than BLM. Both drugs inhibit cell proliferation during S- and G2-phase. Given in combination, BLM and PEP inhibit cell proliferation in a highly significant synergistic way (FIC indexes: 0.25-0.46). This in vitro result, which might be of therapeutic importance, is correlated with differences on the molecular level. Determinations of the ratio between the number of single- and double-strand breaks in the DNA (the target molecule of the drugs) revealed a considerably lower value for DNA from BLM-treated cells (1.9:1) than for DNA from PEP-treated cells (13:1).

Animals↗
PubMed1983

Partial purification and properties of a chromatin bound endonuclease from the marine sponge Geodia cydonium.

A chromatin bound endonuclease (Mr:107,000) has been extracted and partially purified from the siliceous sponge Geodia cydonium. Disc gel electrophoresis showed that only one enzyme was present in the partially purified preparation which was able to degrade DNA and poly(A). The enzyme liberates oligonucleotides on incubation with poly(A), which are further degraded to yield the 5'-mononucleotide, which has a pI of 6.5 and a pH optimum of 7.5-8.0. Cations are not required for enzymic activity and EDTA does not inhibit the enzyme. Only iodosobenzoic acid was found to completely inhibit the enzyme. The enzyme hydrolysed poly(A), poly(U), poly(C), DNA, poly[d(A-T)], poly[d(G-C)], but not poly (dA) or poly(G).

Animals↗
PubMed1983

Chromatin structure from the marine sponge Geodia cydonium.

The histones isolated from the siliceous sponge Geodia cydonium have been separated using two electrophoretic techniques. A comparison of their mobilities with those of calf thymus and rat liver show that some Geodia histone species (H3, H1 and H1(0) exhibit electrophoretic variance. The results show, that as in other eukaryotic systems the sponge chromatin contains the core histones (H2A, H2B, H3 and H4) and the linker histone (H1). ADP-ribosylation of Geodia histones and separation of the individual histones by electrophoresis resulted in four histones being radiolabeled. Digestion of Geodia chromatin with endogenous endonuclease is shown to result in the formation of nucleosome particles containing approximately 200 base pairs of DNA. A major product of endogenous endonuclease digestion is a relatively stable 110 base pair intermediate. Incubation of chromatin with DNase II and separation of the products under denaturing conditions reveals 20 bands migrating at 10 base intervals.

Adenosine Diphosphate Ribose↗
PubMed1983

Alteration of activity of nuclear envelope nucleoside triphosphatase in quail oviduct and liver in dependence on physiological factors.

The concentrations of the specific proteins ovalbumin and avidin in quail oviduct increase during estrogen and progesterone treatment of immature animals. After 6 days' treatment, values are found which are almost as high as those determined in mature oviduct. During ageing, the concentrations of these proteins decrease by 90%. The alterations in protein content are correlated with the activity of the nuclear envelope nucleoside triphosphatase, an enzyme which is assumed to be involved in the nucleocytoplasmic transport of mRNA. During the process of cell proliferation and differentiation, the activity of this enzyme increases from 0.29 to 2.2 mumol Pi/h X 10(8) nuclear ghosts. It was seen that the liver enzyme was not influenced by hormone treatment and ageing.

Aging↗
PubMed1983

Immunochemical identification of the cell surface bound leucine aminopeptidase, the target enzyme for the immunostimulant bestatin.

The microbial product bestatin is known to inhibit soluble microsomal- and cytosolic leucine aminopeptidase (Leu-APm and Leu-APc) as well as aminopeptidase B (AP-B). To clarify which of these enzymes is the target for bestatin on the cell surface, indirect immunofluorescence studies with antisera raised against purified Leu-APm and AP-B were performed. These antibodies (anti-Leu-APm and anti-AP-B) were found to react with intracellularly localized Leu-APm and AP-B of ethanol-treated L5178y cells. Using non-treated L5178y cells fluorescence was detected only on the cell surface after incubation with anti-Leu-APm. To confirm the supposition that only Leu-APm is present on the cell surface, the AP from the cell membrane was solubilized and analyzed electrophoretically. Based on relative migration data it could be shown, that the cell surface is charged with Leu-APm and not with detectable amounts of Leu-APc or AP-B. Moreover, it could be demonstrated that the solubilized Leu-APm binds to [3H]bestatin.

Adjuvants, Immunologic↗
PubMed1983

Nuclear-envelope nucleoside triphosphatase: stimulation by poly(A) (+)mRNA and modulation by microtubule protein.

Nuclear envelopes contain a nucleoside triphosphatase which is thought to be involved in the supply of energy for nucleo-cytoplasmic RNA transport. This enzyme is stimulated most efficiently by poly(A) and to a lesser extent by poly(G) and poly(dT). Half-maximal stimulation of the enzyme from rat liver nuclei, which was associated with the poly(A)-specific Endoribonuclease IV and was free from poly(A) polymerase and Endoribonuclease V activity, was determined to occur at a concentration of 1.1 X 10(6) poly(A) molecules/nuclear ghost. Double-reciprocal plot analysis revealed a 2.8-fold stimulation of the enzyme by poly(A). Poly(A) in the hybrid form had no influence on the activity of the nucleoside triphosphatase. Stimulation by oligo(A) required a minimal chain length of 18 nucleotide units. Naturally occurring RNA species enhanced the nucleoside triphosphatase activity, provided they contained a poly(A) segment. Using poly(A)(+)mRNA, half-maximal stimulation was determined to proceed at 0.5 X 10(6) molecules/nuclear ghost. Removal of the poly(A) segment from mRNA mRNA abolished the stimulatory effect on the enzyme. Microtubule protein was found to inhibit the nucleoside triphosphatase efficiently. At a concentration of 2.0 mg/ml, polymerized microtubule protein reduced the enzyme activity by 96%. Dimeric tubulin was less inhibitory, while actin was without any significant effect. From these findings it is suggested that a possible nucleoside triphosphatase-mediated transport of poly(A)(+) mRNA through nuclear envelope is controlled firstly, by the poly(A) segment of this RNA species and secondly, by cytoplasmic microtubules.

Animals↗
PubMed1982

Interaction of polyribosomal components and polyribonucleotides with microtubule proteins.

To demonstrate the affinity of RNA-containing polyribosomal components (isolated from L5178y cells) to microtubules, microtubule protein was attached to an insoluble matrix. In contrast to ribosomes, poly(A)(+)mRNA and poly(A)-RNP were found to bind to the matrix. Using synthetic polyribonucleotides, no significant differences in the binding properties of single- and double stranded polymers of different base composition to microtubule protein were observed. However, binding is dependent on the size of the polymer; a minimal chain length of 12 nucleotide units is required.

Animals↗
PubMed1982

The effect of benzo[a]pyrene on sponges as model organisms in marine pollution.

The majority of the investigations were performed with the marine sponge Tethya lyncurium at concentrations of 2 X 10(-8) to 1 X 10(-11) g/ml of benzo[a]pyrene (BaP). Sea-pollution was characterized as BaP equivalent activity in the Ames test. Increased activity of ornithine decarboxylase (ODC) was observed when sponges were artificially exposed at polluted marine areas for 3 weeks. In contrast to the situation in higher animals no ODC induction of the fast type was observed. Mixed function oxygenases (MFO) were not detected in sponges nor could they be induced as in vertebrates. BaP was absorbed by Tethya and concentrated 30--60-fold. In live, but not dead, artificially perfused sponges [3H]- and [14C]BaP-radiolabeled became firmly associated with DNA, RNA and protein of the sponges. The association persisted in isolated fractions, in nucleotides, in nucleosides and in protein hydrolysates. The BaP binding ratio to DNA was found to be strongly correlated to the concentration of BaP. Light modifies BaP and thus enables binding. In the dark only very low association, if any, is observed. The possible consequences of these findings are discussed.

Animals↗
PubMed1982

Role of actin and tubulin in the regulation of poly(A) polymerase-endoribonuclease IV complex from calf thymus.

The poly(A) polymerase from calf thymus (Mr = 62,000) is associated with the poly(A)-specific endoribonuclease IV (Mr = 45,000); this complex is dissociable at pH 8.2. The activities of the two enzymes in the complex are strongly inhibited by the G-actin and tubulin (both in the subunit-dissociated and in the polymeric form); F-actin was less inhibitory, while myosin and actomyosin were without any effect. Gel filtration experiments revealed a complex formation between poly(A) polymerase/endoribonuclease IV and G-actin or tubulin at an approximately 1:1 molar ratio. The homogeneous poly(A) polymerase was determined to be maximally inhibited by a 5-fold excess of G-actin and by less than a 2-fold excess of tubulin dimer, if calculated on the molecular basis. The inhibitory potency of G-actin on poly(A) polymerase was reduced after co-incubation with DNase I or phalloidin and that of tubulin by colchicine. It is suggested that actin and/or tubulin is involved also in regulation of poly(A) metabolism in intact cells.

Actins↗
PubMed1982

Fibronectin is apparently not involved in species-specific reaggregation of cells from the marine sponge geodia cydonium.

Experiments were carried out to test the hypothesis that fibronectin is involved in reaggregation of dissociated sponge cells. Cells from the siliceous sponge Geodia cydonium were extracted with urea to solubilize fibronectin from cells of higher multicellular organisms. The crude extract was further fractionated by DNA, heparin, and collagen affinity chromatography; they were termed Geodia fibronectin like fractions. The fibronectin like fractions contained a series of proteins with molecular weights different from that of the genuine fibronectin. The Geodia fibronectin like fractions did not react with antiserum, produced against human fibronectin, under formation of a precipitin line. Using this antiserum the sponge cells could not be specifically labeled with FITC-anti-IgG antiserum. Radioimmunoprecipitation experiments revealed that the Geodia fractions contain--if at all--0.1% fibronectin or fibronectin like protein at the most. In the crucial experiments it was shown that the Geodia fibronectinlike fractions, human fibronectin, and antifibronectin antiserum exerted no influence on adhesion of Geodia cells either in the absence or in the presence of the soluble aggregation factor. Based on these findings, we conclude that fibronectin is apparently not present on Geodia cells and does not play a role in aggregation of this biological system.

Animals↗
PubMed1982

The effects of halothane on the DNase I activity in an isolated enzyme preparation and in the DNase I-G actin complex.

The effects of halothane on the DNase I activity in an isolated enzyme preparation and in a DNase I-globular (G) actin complex was investigated. DNase I, DNase I-G actin complexes and G actin were exposed to various (0.2-4.0 vol./%) halothane concentrations for 3 h. Thereafter, DNase I was mixed with a DNA solution and the extinction of the acid soluble supernatant of the DNase I assay was determined as a measure of DNase I activity. After 10 min of halothane exposure the DNase I activity is inhibited in direct proportion to halothane concentrations between 0.6 and 4.0 vol/%. After 10 min halothane activates inactive DNase I by inhibiting G actin, an inhibitor of DNase I. G actin, exposed to halothane, does not inhibit the activity of DNase I. The results suggest a mechanism by which halothane may contribute to chromosomal defects and disturbances of DNA metabolism in cells.

Actins↗
PubMed1982

Electron microscopical characterization of sponge aggregation factors.

Aggregation factors, purified from 14 sponges which belong to the classes Tetraxonida and Cornacuspongia, Were visualized electron microscopically. Two types of basic structural forms were detected; first, circular structures from the species Ancorina cerebrum, Mycale massa, Hemimycale columella, Crella rosea, Clathria coralloides, Axinella cannabina, Pellina semitubulosa, Ircinia muscarum, Hippospongia communis, Verongia aerophoba and second, rod-like structures from Tethya lyncurium, Tedania anhelans, Hymeniacidon sanguinea, Dysidea tupha. In most of the cases the structures carry side chains.

Animals↗
PubMed1982

Age-dependent gene induction in quail oviduct. XV. Alterations of the poly(A)-associated protein pattern and of the poly(A) chain length of mRNA.

The effect of ageing on polyadenylate [poly(A)] metabolism of mRNA was studied in two age groups of female quails: mature (250-320 days' old) and senescent animals (3-3.5 years' old). In introductory experiments it was shown that poly(A)-associated proteins can not be recovered from cytosol by affinity chromatography. We isolated the poly(A)-associated proteins from polyribosomal poly(A)-ribonucleoprotein complex [poly(A)-RNP] and radioactively labeled them with dansyl chloride. Three main protein species were identified with molecular masses of 48000 (P48), 35000 (P35) and 24000 (P24). During ageing the percentage portion of P48 in poly(A)-RNP from liver (mitotic tissue) and from oviduct or heart (post-mitotic tissue) is reduced at the expense of P35 and P24. Quantitative analyses revealed that the amount of poly(A)-RNP in the different organs decreases significantly with age if the values are based on DNA. The protein content in poly(A)-RNP was found to be reduced especially in post-mitotic tissue. From this finding we assume that the number of poly(A)-associated protein molecules per poly(A) stretch drops from approximately 4.7 molecules (mature oviduct) to 1.9 molecules (senescent oviduct). Control experiments revealed that free, non-polyribosomal poly(A)-RNP accounts only for 10% of total poly(A)-RNP. The size of the poly(A) segment of mRNA decreases with age. After labeling with [3H] dimethylsulfate, the poly(A) stretch from mature oviduct was found to consist mainly of 120-180 AMP units, and those from mature liver and mature heart of 110 and 100, respectively. In organs from senescent animals the percentage of shorter poly(A) stretches is enlarged; on the average, poly(A)-70 chains were detected. These results support the assumption that age-dependent changes occur also on the post-transcriptional level during the maturation steps of poly(A)(-) hnRNA to poly(A)-(+) mRNA.

Aging↗
PubMed1982

Age-dependence of polyadenylate stimulation of nuclear-envelope nucleoside triphosphatase.

Nuclear envelopes of mammalian cells contain a nucleoside triphosphatase which is probably involved in mRNA transport through the nuclear membrane. The activity of the enzyme, studied in RNA-depleted nuclear ghosts, can be stimulated by poly(A) or by poly(A) (+)mRNA. Using nuclear ghost preparations from mature (8-10 months' old) and old (40-42 months' old) Wistar rats, it was shown that in "old" preparations the basal activity of the enzyme is significantly reduced (by 15%). In addition, the enzyme from old animals responds only very little to poly(A) or poly(A) (+)mRNA, compared to preparations from mature animals. Using a concentration of 6.8 X 10(11) poly(A) (+) mRNA molecules per microgram of enzyme preparation, the nucleoside triphosphatase from mature animals is stimulated by 77% and the enzyme from old animals by only 26%. Binding studies of poly(A) to pore laminae revealed that the number of binding sites in unphosphorylated preparations from old animals is significantly reduced (by 24%) compared to "mature" preparations. As a consequence of in vitro phosphorylation, no difference is observable in the number of binding sites between the two age groups. The values for half-maximal saturation binding constants for poly(A) are identical in unphosphorylated and phosphorylated pore-laminae preparations, irrespective of the age group studied. The results presented indicate that in old animals the pathway from the phosphorylated to the dephosphorylated nuclear-envelope protein which is controlled by poly(A) is impaired in the proposed cycle for mRNA efflux from nuclei.

Aging↗
PubMed1982

Properties and specificity of binding sites for the immunomodulator bestatin on the surface of mammalian cells.

The binding of the immunomodulator bestatin, an inhibitor of cell surface bound leucine aminopeptidase and aminopeptidase B, to mammalian cells of varying origin has been studied. The specific binding of [3H] bestatin was a rapid and saturable process exhibiting one affinity, characterized by an association constant of 0.8 x 10(5) M-1, as determined in the L5178y mouse lymphoma system. Optimal binding was observed at 37 degrees C. L-leucine and L-leucine-beta-naphthylamide prevented the binding, suggesting that the complex was formed between leucine aminopeptidase and bestatin. The protein nature of the bestatin-"receptor" was suggested by its susceptibility to trypsin. Under the conditions used here intracellular translocation of bestatin appeared to be negligible. A maximum of about 2.2 x 10(6) bestatin molecules could bind to L5178y mouse lymphoma cells. Under identical conditions by far the highest amount of bestatin was bound to macrophages from mice. Lower levels were measured with T-lymphocytes; very low binding capacity was observed with B-lymphocytes. Experiments with synchronized L5178y cells revealed a cell cycle dependent change of binding capacity for bestatin; the highest level was observed during the transition from S-to G2 phase and the lowest during G1- and early S phase. These data lend further support to the assumption that the immuno-potentiating activity of bestatin is due to a stimulation of T-lymphocyte proliferation probably mediated through the activation of macrophages.

Adjuvants, Immunologic↗