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Biomedical subjects

A Fridland

Publications and source records attributed to A Fridland.

67 records · Page 4Linked to original sources
PubMed1975

DNA replication in methotrexate-treated human lymphoblasts.

The rate of DNA synthesis in cultures of human lymphoblasts decreased more than 80% within 30 min after the cells were exposed to methotrexate, a potent inhibitor of dihydrofolate reductase. Despite this rapid initial inhibition, DNA continued to be synthesized for at least an additional 6 h. The mode of this subsequent replication appeared to be semiconservative, as indicated by the buoyant density of 5-bromodeoxyuridine-substituted DNA in alkaline CsCl gradients. The growth rates of DNA chains in cells exposed to methotrexate were determined by sedimentation rate analysis in alkaline sucrose gradients. DNA synthesized during 2-min or 10-min pulses with labeled deoxycitidine in the presence of methotrexate had about the same sedimentation coefficient, 35 S, as controls. When methotrexate-treated cultures were pulse-labeled for 10 min and then chased for various times, DNA fragments of about 80 S accumulated. DNA synthesized in the presence of methotrexate was stable and elongated to bulk-size DNA after methotrexate inhibition of growth was removed by addition of thymidine and deoxycytidine. The data suggest that methotrexate reduces the rate of DNA replication by inhibiting chain initiation independently of chain elongation.

Cell Line↗
PubMed1967

Studies on glycogen synthesis in pigeon liver homogenates. Incorporation of hexose into glycogen.

Liver homogenates of avian species, but not of mammals, form glycogen from glucose, mannose, fructose and galactose. Incorporation of labelled glucose, fructose and mannose, but not of labelled galactose, into glycogen is diluted isotopically by unlabelled glucose. Except for fructose, glycogen formation from other substrates by pigeon liver homogenates compares favourably with that from the same substrates in pigeon liver slices. Optimum conditions for glycogen synthesis from glucose by pigeon liver homogenate are: medium of incubation, 0.175m-sucrose-45mm-potassium chloride-15mm-glycylglycine buffer, pH7.5; concentration of substrate, 15mm; concentration of tissue, less than 120mg./ml.; temperature of incubation, 37-43 degrees ; atmosphere, oxygen. Uncouplers of oxidative phosphorylation, Ca(2+), EDTA, PP(i), 2-deoxyglucose 6-phosphate and microsomal fraction of rat liver are inhibitory to glycogen synthesis from glucose. Starvation of pigeons for 24 and 48hr. leads to a slight stimulation of glycogen synthesis in their liver homogenates as compared with fed controls. Pigeon liver homogenates can be separated into subcellular fractions that on reconstitution can synthesize glycogen. All the enzymes of the glycogen pathway except soluble high-K(m) glucokinase are present in pigeon liver.

Animals↗
PubMedDate not supplied

Cellular phosphorylation of 2',3'-dideoxyadenosine-5'-monophosphate, a key intermediate in the activation of the antiviral agent DDI, in human peripheral blood mononuclear cells.

2',3'-dideoxyadenosine 5-monophosphate (ddAMP), is a key intermediate in the metabolism of the antiviral agent 2',3'-dideoxyinosine (ddI) to its active triphosphate derivative, 2',3'-dideoxyadenosine-5'-triphosphate (ddATP). The potential role of adenylate kinase in the phosphorylation of ddAMP was studied in human peripheral blood mononuclear cells (PBMC) and a human T cell line, CEMss. Subcellular distribution, sulfhydryl inhibitor, and substrate specificity studies support the hypothesis that the mitochondrial adenylate kinase (AK2) is a major route of cellular activation of these compounds in human lymphocytes.

Adenylate Kinase↗
PubMedDate not supplied

Rhodamine 123 and flow cytometry to monitor the cytotoxic actions of nucleoside analogues in nondividing human lymphocytes.

We used human lymphocytes from the peripheral blood of normal individuals to compare the cytolytic actions of 2-chloro-2'-deoxyadenosine (2-Cl-dAdo) and 9-beta-D-arabinofuranosylguanine (ara-G). Membrane integrity was ascertained by flow cytometric quantification of the cells' uptake of the fluorescent mitochondria-specific probe rhodamine 123. Addition of 10 microM ara-G to lymphocyte cultures led to a progressive decline in the number of cells that could assimilate rhodamine 123, and after 2 days exposure to drug less than 50% of the cells showed normal staining compared to untreated cells. 2-Cl-dAdo had similar cytotoxic effects at a 0.1 microM concentration. Under the same conditions trypan blue revealed only a 10-20% toxic effect by these drugs. Using rhodamine 123 in combination with the vital stain propidium iodide, we found that 90% of the cells with abnormal rhodamine 123 uptake also stained with propidium iodide. The changes in membrane permeability to fluorescent dyes correlated with the loss of lymphocyte ability to respond to the plant mitogen phytohemagglutinin. Our data indicate that rhodamine 123 is a more sensitive probe than trypan blue for ascertaining early loss of the membrane integrity of dying lymphocytes. Combined with flow cytometry, rhodamine 123 uptake represents a simple and rapid method for identifying the important biochemical events associated with the cytocidal and cytostatic effect of drugs against normal and leukemic cells.

Arabinonucleosides↗
PubMedDate not supplied

Anomalous accumulation and decay of 2',3'-dideoxyadenosine-5'-triphosphate in human T-cell cultures exposed to the anti-HIV drug 2',3'-dideoxyinosine.

The rates of accumulation and decay of 2',3'-dideoxyadenosine-5'-triphosphate (ddATP) have been examined after incubation with the anti-human immunodeficiency virus (HIV) agents 2',3'-dideoxyinosine (ddIno) and 2',3'-dideoxyadenosine (ddAdo) in human T-cell systems frequently used for assay of anti-HIV agents (MOLT-4 and CEM). Formation of ddATP from ddIno or ddAdo was rapid and concentration-dependent, with no saturation of phosphorylation being observed up to extremely high levels (1 mM) of drug. Rates of removal of ddATP from MOLT-4 cells were slow (t1/2 = 25-40 hr) and appeared to be monophasic. These unusually long half-times for ddATP utilization are not a general property of purine dideoxypurine nucleosides: when the corresponding guanine analog (2',3'-dideoxyguanosine) was examined under the same conditions, the t1/2 of ddGTP removal was only 3-5 hr. Similar results were observed with the human T-cell line CCRF-CEM. Coadministration with ddIno of inosine monophosphate dehydrogenase inhibitors, such as ribavirin and tiazofurin, yielded higher levels of ddATP in MOLT-4 and CEM cells, but did not influence the slow removal of ddATP from T-cells. The long half-time for disappearance of ddATP from cells may permit the maintenance of pharmacologically effective levels of ddATP within cells with relatively infrequent administration of the parent drug (ddIno or ddAdo).

Antiviral Agents↗