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recombinant human secretory phospholipase a2: purification and characterization of the enzyme for active site studies.a secreted form of phospholipase a2 (pla2) is thought to play an important role in inflammatory diseases. to characterize this enzyme the cdna encoding a low molecular weight pla2 was cloned from a human placental cdna library. the cdna encoding the human pla2 was subcloned into an expression vector and subsequently transfected into chinese hamster ovary (cho) cells. a stable cho cell clone, secreting ca 1 mg/l of recombinant pla2 into the medium, was scaled up in culture to 180 l. the recombina ...19921339483
immediate cell membrane damage produced by entamoeba histolytica subcellular extracts.the alpha[14c]amino isobutyric acid is a synthetic amino acid with low molecular weight, not incorporated to protein. chinese hamster ovary (cho) cells were labeled with this radioactive compound and treated with the amebal subcellular fraction p30 at short times. we detected membrane damage in cho cells by specific radiolabel release from the first 30 sec of interaction between cell cultures and amebal fraction.19921340292
bile salts promote adherence-decreasing effect of colonic luminal hydrolases on entamoeba histolytica.e. histolytica trophozoites cultivated for > 7 h in the presence of glycosidases, produced by a subset of the colonic anaerobic bacteria of healthy humans, and pancreatic proteases show decreased adherence to chinese hamster ovary epithelial cells. since activities of the glycosidases are enhanced by bile salts we investigated whether bile salts would enhance the e. histolytica-cho cell adherence decreasing effects of the luminal hydrolases (glycosidases plus proteases). cho cell adherence of co ...19921340299
cloning and functional characterization of a family of human and mouse somatostatin receptors expressed in brain, gastrointestinal tract, and kidney.somatostatin is a tetradecapeptide that is widely distributed in the body. it acts on multiple organs including brain, pituitary, gut, exocrine and endocrine pancreas, adrenals, thyroid, and kidneys to inhibit release of many hormones and other secretory proteins. in addition, it functions as a neuropeptide affecting the electrical activity of neurons. somatostatin exerts its biological effects by binding to specific high-affinity receptors, which appear in many cases to be coupled to gtp-bindin ...19921346068
mapping of 262 dna markers into 24 intervals on human chromosome 11.we have extended our mapping effort on human chromosome 11 to encompass a total of 262 dna markers, which have been mapped into 24 intervals on chromosome 11; 123 of the markers reveal rflps. these clones are scattered throughout the chromosome, although some clustering occurs in r-positive bands (p15.1, p11.2, q13, and q23.3). fifty-two of the markers were found to contain dna sequences conserved in chinese hamster, and some of these 52 also cross-hybridized with dna from other mammals and/or c ...19921346079
sdz 280-446, a novel semi-synthetic cyclopeptolide: in vitro and in vivo circumvention of the p-glycoprotein-mediated tumour cell multidrug resistance.sdz 280-446 is a semi-synthetic derivative of a natural cyclic peptolide. its ability to sensitise in vitro tumour cells whose resistance is due to p-glycoprotein-mediated anticancer-drug efflux was shown using four different pairs of parental drug-sensitive (par-) and multidrug-resistant (mdr-) cell lines, from three different species (mouse, human, chinese hamster) representing four different cell lineages (monocytic leukaemia, nasopharyngeal epithelial carcinoma, colon epithelial carcinoma, o ...19921346365
antagonism by antimuscarinic and neuroleptic compounds at the five cloned human muscarinic cholinergic receptors expressed in chinese hamster ovary cells.we determined the affinity and selectivity of binding for 24 compounds: nine antimuscarinics (including some antiparkinson drugs) and 15 neuroleptics (including the atypical compounds clozapine, fluperlapine, melperone, rilapine, risperidone, tenilapine, tiosperone and zotepine) at the five human muscarinic receptor subtypes expressed in chinese hamster ovary cells. equilibrium dissociation constants (kd) were obtained from competitive radioligand binding studies with [3h]quinuclidinyl benzilate ...19921346637
peptide transport by the multidrug resistance pump.the membrane p-glycoprotein (p170) is an atp-hydrolyzing transmembrane pump, and elevated levels of p170, due to higher expression with or without amplification of the multidrug resistance gene (mdr1), result in resistance to a variety of chemotherapeutic agents in mammalian cells. the function of the p170 pump has been proposed as a protection against toxic substances present in animal diets. here we describe a chinese hamster ovary cell line that was selected for resistance to a synthetic trip ...19921348245
effect of p-glycoprotein expression on the accumulation and cytotoxicity of topotecan (sk&f 104864), a new camptothecin analogue.topotecan (tpt, 9-dimethylaminomethyl-10-hydroxycamptothecin) is the first topoisomerase i-directed cytotoxic agent to enter clinical trials in the united states in two decades. the effect of p-glycoprotein (pgp) overexpression on tpt cytotoxicity was examined in chrc5 (colchicine-resistant) and auxb1 (parental) chinese hamster ovary cells. examination of the ic50 values observed in colony-forming assays revealed that the chrc5 cells were 15-fold (sd, +/- 3; n = 3) resistant to tpt after a 1-h e ...19921348448
localization of the nucleotide excision repair gene ercc6 to human chromosome 10q11-q21.we have cloned the human dna excision repair gene ercc6 by virtue of its ability to correct the uv sensitivity of chinese hamster overy cell mutant uv61. this mutant is a member of complementation group 6 of the nucleotide excision repair-deficient rodent mutants. by means of in situ hybridization and southern blot analysis of mouse x human somatic cell hybrids, the gene was localized to human chromosome 10q11-q21. an rflp detected within the ercc6 locus can be helpful in linkage analysis.19921349298
ablation of stimulation of a camp-responsive promoter in cho cell lines defective in their camp-dependent protein kinase system.we have studied the requirement for an intact camp-dependent protein kinase (pka) system to regulate camp-mediated gene transcription in chinese hamster ovary (cho) cells. wild-type cho cells and mutant cho cell lines selected for their resistance to the growth inhibitory effect of 8-br-camp and defective in their pka system were transiently transfected with reporter plasmids containing 2.5 and 3.0 kb of the 5'-flanking sequence of the rat tyrosine aminotransferase (tat) gene promoter. this segm ...19921349445
perinatal changes in the coupling of beta 1- and beta 3 adrenergic receptors to brown fat adenylyl cyclase.the stimulation of adenylyl cyclase by catecholamines in neonatal brown adipose tissue (bat) is markedly biphasic, suggesting the existence of receptors that have both high and low affinities for catecholamines. the identities of these receptors were examined by comparing responses in neonatal bat membranes to those of chinese hamster ovary cells which had been transfected to express the cloned rat beta 1 and beta 3 receptors. the results from these experiments indicate that high-affinity stimul ...19921349643
sequence and expression of rat icam-1.we have isolated cdna clones-coding for rat intercellular adhesion molecule-1 (ricam-1) from a cdna library constructed from rat ax cells stimulated with il-1 beta using the mouse icam-1 cdna as a hybridization probe. the ricam-1 sequence shows 79.1% homology with mouse icam-1 and 55.6% homology with human icam-1 at the nucleic acid level. in order to examine the expression of ricam-1 on chinese hamster ovary (cho) cells, we constructed the vector, psv-ricam1-neo, containing the sv40 promoter. f ...19921349828
mapping of ornithine aminotransferase gene sequences to mouse chromosomes 7, x, and 3.ornithine aminotransferase (oat), a mitochondrial matrix enzyme, is deficient in patients with gyrate atrophy of the choroid and retina. in human, the oat structural gene maps to chromosome (chr) 10q26 and several oat-related sequences, some of which are known to be processed pseudogenes, which map to xp11.3-11.21. here, we report chromosomal localization in the mouse of the oat gene and related sequences. genomic dna blot analysis of a well-characterized panel of chinese hamster x mouse somatic ...19921349842
engagement of class i major histocompatibility complex molecules by cell surface cd8 delivers an activation signal.recent evidence has demonstrated that cross-linking class i major histocompatibility complex (mhc) molecules on human t cells with monoclonal antibodies (mab) triggers t cell activation. the only known natural ligand for mhc class i molecules is cd8. therefore, the possibility that cd8+ t cells might provide activation signals to other t cells by engaging mhc class i molecules was examined by culturing cd4+ peripheral blood t cells with chinese hamster ovary cells (cho) cells that had been trans ...19921350981
postnatal development of organic cation transport and mdr gene expression in mouse kidney.the apical surface of the proximal tubular epithelium is the site of both p-glycoprotein localization and postulated active secretion of organic cations in the mammalian kidney. p-glycoprotein has been shown to act as a pleiotropic drug efflux pump across the cell membrane of tumor cells expressing the multidrug resistance phenotype, whereas the renal organic anion and organic cation secretory systems serve the function of pleiotropic drug transport across the proximal tubule epithelium. because ...19921351097
identification of a processed protein related to the human chaperonins (hsp 60) protein in mammalian kidney.the chaperonin family of proteins, which includes groel protein of e. coli, yeast heat shock protein (hsp-60) and the ribulose-1-5-bisphosphate carboxylase (rubis co.) subunit binding protein of plant chloroplasts, shows strong sequence homology to the chinese hamster ovary (cho) mitochondrial p1 protein. we have identified a 60 kda protein from bovine kidney which by n-terminal sequencing gives the amino acid sequence akdvkfgadarallmlqgvdllada. bovine whole kidney membranes were delipidated, so ...19921351721
characterization of a new hybrid mink-mouse clone panel: chromosomal and regional assignments of the glo, acy, np, ckbb, adh2, and me1 loci in mink (mustela vison).to expand the mink map, we established a new panel consisting of 23 mink-mouse clones. on the basis of statistical criteria (wijnen et al. 1977; burgerhout 1978), we developed a computer program for choice of clones of the panel. assignments of the following mink genes were achieved with the use of the hybrid panel: glyoxalase (glo), chromosome (chr) 1; acetyl acylase (acy), chr 5; creatine phosphokinase b (ckbb), chr 10; alcohol dehydrogenase-2 (subunit b) (adh2), chr 8. using a series of clone ...19921352156
reduced mrna levels for the multidrug-resistance genes in camp-dependent protein kinase mutant cell lines.we have previously shown that in chinese hamster ovary (cho) cells, a mutant cell line with a defective regulatory subunit (ri) for the camp-dependent protein kinase (abraham et al: mol. cell. biol., 7:3098-3106, 1987), and a transfectant cell line expressing the same mutant kinase, showed increased sensitivity to a number of drugs that are known to be substrates for the multidrug transporter (p-glycoprotein). in the current study we have investigated the mechanism by which camp-dependent protei ...19921352302
pharmacological properties of two cloned somatostatin receptors.previous studies have shown that at least two subtypes of somatostatin (srif) receptors (srif1 and srif2) are expressed in mammalian cells. srif1 receptors have high affinity for mk 678, whereas srif2 receptors have no affinity for mk 678 but selectively bind peptides with structures similar to that of cgp 23996. recently, two srif receptor genes have been cloned from human and mouse genomic libraries. in the present study, the pharmacological properties of these two cloned srif receptors, expre ...19921353250
alternative polyadenylation of the amyloid protein precursor mrna regulates translation.the sequence of several cdnas encoding the amyloid protein precursor showed that two polyadenylation sites of the mrna are utilized; rna blot analysis with different riboprobes indicated that this explains the difference between the two major 3.2 and 3.4 kb mrnas found in the human brain. these two mrnas, which contain the whole sequence of the natural molecules, were synthesized by in vitro transcription and translated in xenopus oocytes. the long mrna using the second polyadenylation site prod ...19921353447
multifactorial resistance to antineoplastic agents in drug-resistant p388 murine leukemia, chinese hamster ovary, and human hela cells, with emphasis on the role of dna topoisomerase ii.the role of dna topoisomerase ii in multifactorial resistance to antineoplastic agents is reviewed. we have previously observed that in adriamycin (adr) resistant p388 murine leukemia cells, dna topoisomerase ii enzyme content and cleavage and catalytic activities were all reduced and correlated with drug sensitivity. a subsequent study provided evidence for an allelic mutation of the gene for dna topoisomerase ii as a possible molecular mechanism underlying the enzyme alterations. to ascertain ...19921353968
doxorubicin-loaded nanospheres bypass tumor cell multidrug resistance.we have demonstrated that in vitro resistance of tumor cells to doxorubicin (dox) can be fully circumvented by using doxorubicin-loaded nanospheres (dox-ns), consisting of biodegradable polyisohexylcyanoacrylate polymers of 300 nm diameter and containing 2.83 mg of dox per 31.5 mg of polymer. five different multidrug-resistant cell lines, characterized by mdr1 amplification, were used in this study: dox-r-mcf7, a human breast adenocarcinoma; skvbl1, a human ovarian adenocarcinoma; k562-r, a huma ...19921354963
a naturally occurring tyrosine to histidine replacement at residue 33 of human thymidylate synthase confers resistance to 5-fluoro-2'-deoxyuridine in mammalian and bacterial cells.structural changes in the macromolecular targets of pharmacological agents can result in alterations in the efficacy of these agents. in previous studies, we identified a variant structural form of thymidylate synthase (ts) that is associated with relative resistance to 5-fluoro-2'-deoxyuridine, in a human colonic tumor cell line. we now report on the use of dna transfer techniques to examine directly the effects of each ts form on drug response. ts cdna constructs, corresponding to the normal o ...19921355260
genetic aspects of multidrug resistance.mammalian cells exposed to a single cytotoxic natural product drug, such as vincristine or dactinomycin, can develop resistance to the selective agent and cross-resistance to a broad spectrum of structurally and functionally distinct antibiotics and alkaloids. this phenomenon, termed multidrug resistance (mdr), has been widely studied experimentally. the most consistent feature of cells with high-level mdr is amplification and overexpression of genes encoding an integral plasma membrane protein ...19921355404
molecular cloning and chromosomal localization of a human gene encoding d-amino-acid oxidase.genomic clones covering the entire sequence of the gene encoding human d-amino-acid oxidase (dao) (ec 1.4.3.3), one of the principal and characteristic flavoenzymes of peroxisomes, were isolated from human placental genomic libraries with the aid of a previously cloned cdna for human dao as a probe. nucleotide sequence analysis revealed that the gene, present as a single copy in the human genome, comprises 11 exons and spans 20 kilobase pairs. the protein sequences containing the catalytically i ...19921356107
chinese hamster ovarian cell glycoproteins that mediate type 1 piliated gram-negative bacterial adherence.we used chinese hamster ovary (cho) cell lines to define the structures of glycoproteins responsible for type 1 piliated bacterial adherence. csh 50 escherichia coli, a type 1 piliated bacteria, adhered significantly better than an isogenic nonpiliated e. coli to all cho lines tested. csh 50 e. coli adhered least well to cho cells expressing intact complex type oligosaccharides on cell surface glycoproteins. csh 50 adherence increased when shorter oligosaccharides were present and was maximal wh ...19921356377
total body and hepatic clearance in rats of recombinant tissue-type plasminogen activator expressed in mouse c127 and chinese hamster ovary cells.the total body and hepatic clearance of two recombinant human tissue-type plasminogen activators (t-pa) produced in mouse c127 cells [t-pa(c127)] and in chinese hamster ovary cells [t-pa(cho)] were determined in rats. recombinant t-pa was administered as a constant iv or intraportal venous infusion for 60 min. the plasma t-pa level over 120 min was measured by an elisa. whole-body pharmacokinetics were characterized in terms of the total body clearance (clt), total volume of distribution, and me ...19921356732
in vitro study of a novel atypical beta-adrenoceptor agonist, sm-11044.sm-11044 (l-threo-3-(3,4-dihydroxyphenyl)-n-[3-(4-fluorophenyl) propyl] serine pyrrolidine amide hydrobromide) stimulated the relaxation of guinea pig ileum (ec50: 3.0 x 10(-8) m), trachea (ec50: 1.3 x 10(-7) m), lung parenchyma (ec50: 2.1 x 10(-6) m) and the increase in right atrial rate (ec50: 6.9 x 10(-6) m). it also induced lipolysis of rat white adipocytes (ec50: 1.2 x 10(-6) m). both the relaxant response of ileum and the lipolytic response of adipocytes to sm-11044 were resistant to inhib ...19921356795
intrinsic activity determinations at the dopamine d2 guanine nucleotide-binding protein-coupled receptor: utilization of receptor state binding affinities.guanine nucleotide-binding protein-coupled receptors have been shown to exist in both a high affinity agonist (hiag) and a low affinity agonist (lowag) state. the formation of the hiag state is promoted by agonists, and the formation of this state of the receptor appears to be a critical factor in the generation of the effector-activating complex g alpha.gtp.mg2+ and in the production of a stimulus. the magnitude of the difference in the affinity a compound has for the hiag versus the lowag stat ...19921357542
homology of a vesicular amine transporter to a gene conferring resistance to 1-methyl-4-phenylpyridinium.the vesicular amine transporter (vat) catalyzes transport and storage of catechol and indolamines into subcellular organelles in a wide variety of cells. it plays a central role in neurotransmission and is the primary target for several pharmacological agents. one of the drugs, reserpine, binds very tightly to the transporter and remains bound even after solubilization, a finding that has proven useful for purification of the transporter from bovine adrenal medulla in a fully functional state. t ...19921357668
disposition of metabolically labeled recombinant soluble cd4 (st4) in male sprague-dawley rats following intravenous and subcutaneous administration.soluble cd4 (st4) has been metabolically labeled with [3h]leucine in chinese hamster ovary cells and purified by s sepharose chromatography. over 250 microci of high specific radioactivity [3h]st4 (42 ci/mmol) was prepared. the radiolabeled molecule was chemically and biologically representative of the unlabeled molecule and thus appropriate for in vivo metabolic investigations. to explore the biotransformation and disposition of a recombinant protein, this uniformly labeled [3h]st4 was administ ...19921358574
involvement of sequences near both amino and carboxyl termini in the rapid intracellular degradation of tyrosine aminotransferase.the degradation of rat liver tyrosine aminotransferase has been studied after transfection of suitable expression vectors into mammalian cells in culture. a normal rapid rate of degradation (half-life about 6 h) was observed in cells under stable transfection conditions. however, the higher enzyme levels produced during transient transfections or after amplification with methotrexate caused the apparent half-life of degradation to increase substantially. analysis of expression in chinese hamster ...19921358885
chinese hamster ovary mrna-dependent, na(+)-independent l-leucine transport in xenopus laevis oocytes.in freshly prepared uninjected folliculated oocytes, na(+)-independent leucine uptake is mediated predominantly by a system l-like transport system. removal of follicular cells, however, results in an irreversible loss of this transport activity. when total poly(a)+ mrna derived from chinese hamster ovary (cho) cells was injected into prophase-arrested stage v or vi xenopus laevis oocytes, enhanced expression of na(+)-independent leucine transport was observed. the injected mrnas associated with ...19921360143
liposome-mediated modulation of multidrug resistance in human hl-60 leukemia cells.multidrug resistance (mdr) is a major obstacle in cancer treatment. resistance of cultured tumor cells to major classes of cytotoxic drugs is frequently due to expression of a plasma membrane p-glycoprotein encoded by mdr genes. we have demonstrated that liposome-encapsulated doxorubicin is more toxic than the free drug and that it modulates mdr in chinese hamster lz cells and human colon cancer cells.19921361008
thyroid stimulatory autoantibodies in different patients with autoimmune thyroid disease do not all recognize the same components of the human thyrotropin receptor: selective role of receptor amino acids ser25-glu30.to study the interaction between tsh, autoantibodies and the amino-terminal half of the tsh receptor extracellular region (amino acids 1-260; domains abc), we constructed 20 lh/cg chimeric receptor cdnas. the prototypic receptor for modification contained domains abc of the tsh receptor and domains de of the lh/cg receptor. segments (6-13 amino acids) within the abc domains were replaced with the corresponding amino acids of the rat lh/cg receptor. fifteen of the 20 chimeric receptors could be e ...19921361193
binding assay for thyrotropin receptor autoantibodies using the recombinant receptor protein.we have characterized a transfected chinese hamster ovary cell line, jp09, which expresses high levels of the human tsh receptor (tsh-r). based on a theoretical biological activity for tsh of 40 iu/mg, jp09 has approximately 90,000 receptors per cell, having a dissociation constant of 1.64 x 10(3) mu/l or 1.47 x 10(-9) mol/l. we have used jp09 to prepare solubilized tsh-rs which have formed the basis of a binding assay for thyroid-binding inhibiting immunoglobulins in unfractionated sera. we hav ...19921361194
factors influencing the dna content of radiation-induced micronuclei.the distribution of the dna content of radiation-induced micronuclei was analysed in several cell lines (chinese hamster, syrian hamster and mouse nih-3t3 cells) by flow cytometry. frequency and dna content of micronuclei were measured simultaneously using fluorescence and forward scatter signals of micronuclei and nuclei in suspension stained with ethidium bromide. computerized random breakage of chromosomes and random combination of fragments was performed to compare the measured micronucleus ...19921361516
growth behavior of chinese hamster ovary cells in a compact loop bioreactor: 1. effects of physical and chemical environments.chinese hamster ovary (cho) cells were cultivated in a compact loop bioreactor using mem-alpha medium supplemented with 10% fetal calf serum. effects of physical and chemical environments, i.e., ph in the medium, stirring speed of impellers, temperature and partial pressure of oxygen (po2) upon growth of suspended cells in the bioreactor were determined in batch cultures. growth behavior was characterized by specific rates of growth (mu), glucose consumption (qg) and lactate production (ql), and ...19901366683
effect of free fatty acids and phospholipids on growth of and product formation by recombinant baby hamster kidney (rbhk) and chinese hamster ovary (rcho) cells in culture.recombinant bhk and cho cells producing human antithrombin iii (rh atiii) were used to investigate the utilization of phospholipids and free fatty acids from low-serum (0.1% fbs) culture medium. both cell lines show distinctly different patterns of fatty acid utilization. for rbhk atiii cells it is shown that under low serum conditions several different combinations of free fatty acids (bound to bovine albumin) elicit an identical growth stimulatory effect although individual consumption and pro ...19911366984
cultivation of anchorage-dependent animal cells in microsphere-induced aggregate culture.diethylaminoethyl-derivatized dextran microspheres were used to cultivate chinese hamster ovary, 293, vero and swine testicular cells. cells became attached to the microspheres but did not spread out. instead, they grew in a more spherical shape and eventually formed multiple-cell-layer aggregates. viability in these aggregates remained high after the cultures reached high cell concentrations. this cultivation method allows a high cell density to be achieved with a low microsphere concentration.19911367199
high level expression of the humanized monoclonal antibody campath-1h in chinese hamster ovary cells.we have cloned the light and heavy chain cdnas for the humanized monoclonal antibody campath-1h and expressed them in chinese hamster ovary (cho) cells using a dihydrofolate reductase (dhfr) amplification procedure. each cdna was positioned under control of the strong human beta-actin promoter/polyadenylation signals and used to evaluate alternative vector design and amplification procedures. by employing a dual selection co-transfection strategy, initial transformants accumulated antibody level ...19911367214
high density culture of anchorage-dependent animal cells by polyurethane foam packed-bed culture systems.monkey kidney cells (vero) and chinese hamster ovary cells (cho-k1) attached to the internal surface of polyurethane foam (puf) and grew to a high cell density (1.1 x 10(8) cells/cm3 puf and 4.2 x 10(7) cells/cm3 puf, respectively) in a puf-plates packed-bed culture system. this density of vero cells was twice that obtained previously with a puf-particles packed-bed culture system. a maximum cell density of 6.7 x 10(7) cells/cm3 culture vessel volume was obtained in a puf-disc packed-bed culture ...19911367366
suppression of the degradation of recombinant human apolipoprotein e by a protease inhibitor obtained from fetal bovine serum in serum-free culture.the recombinant human apolipoprotein e (apo-e) produced by chinese hamster ovary cells (cho-322 cells) in serum free culture was degraded to 24k and 23k fragments that contained n-terminal amino acid. the degradation site of apo-e to 24k fragment was between arg180 and leu181 and the c-terminal amino acid of 23k fragment was gly169. in fetal bovine serum (fbs)-containing culture, the degradation was inhibited. however, in calf serum (cs) the inhibitory activity was not detected. thus, we attempt ...19911367397
spin filter perfusion system for high density cell culture: production of recombinant urinary type plasminogen activator in cho cells.we have used a 20 liter stirred tank fermentor, equipped with a 127 mesh ethylene-tetrafluoroethylene rotating screen for cell recycle, for the continuous production of recombinant single chain urokinase-type plasminogen activator (rscu-pa) from chinese hamster ovary (cho) cells. viable cell densities between 60 and 74 million per ml were maintained at medium perfusion rates of 3.0 to 4.0 fermentor volumes per day. cells were retained by the 120 micron nominal opening filter through the formatio ...19901367426
anchorage-dependent mammalian cell culture using polyurethane foam as a new substratum for cell attachment.anchorage-dependent mammalian cells were cultivated at high cell density in a novel culture system using polyurethane foam (puf) as a substratum for cell attachment. puf has a macroporous structure giving a high surface area to volume ratio. monkey kidney cells (vero) and chinese hamster ovary cells (cho-k1) attached to the internal surface of puf and grew to a high cell density (1.04 x 10(8) cells/cm3 puf and 3.5 x 10(7) cells/cm3 puf, respectively) in puf stationary cultures. in addition, we h ...19901367457
chinese hamster ovary cell growth and interferon production kinetics in stirred batch culture.recombinant human interferon-gamma production by chinese hamster ovary cells was restricted to the growth phase of batch cultures in serum-free medium. the specific interferon production rate was highest during the initial period of exponential growth but declined subsequently in parallel with specific growth rate. this decline in specific growth rate and interferon productivity was associated with a decline in specific metabolic activity as determined by the rate of glucose uptake and the rates ...19911367524
"differentiation induction" culture of human leukemic myeloid cells stimulates high production of macrophage differentiation inducing factor.a suitable procedure for the production of human monokines was defined as 'differentiation-induction' culture. human monocytic leukemia thp-1 cells were well-differentiated from nonfunctional promonocytes into macrophage-like cells by the induction with a combination of mezerein, retinoic acid, and a mycoplasma fermentans extract. the differentiated thp-1 cells secreted a high amount of macrophage differentiation-inducing factor (dif) activity and concomitantly produced other known monokines, su ...19911367527
a cho strain producing high-level human il-6 with the 3' deletion construct.a chinese hamster ovary (cho) strain producing high-level human interleukin 6 (il-6) was obtained, by introduction of the il-6 cdna from which the 3' au-rich region was deleted. the il-6 mrna of this strain was stable. the productivity was more than 15 times higher than the previously obtained clone, which has intact il-6 cdna in the same expression vector. this strain produced il-6 at a high level of 30 micrograms ml-1 in batch culture, or a continuous 20 micrograms per 10(6) cells per day in m ...19921367984
production of human antithrombin-iii in a serum-free culture of cho cells.a simple method was developed to establish serum-independent chinese hamster ovary (cho) cells that grew and secreted high level of human antithrombin-iii (at-iii). first, human at-iii and mouse dihydrofolate reductase (dhfr) cdnas were transfected into dhfr-deficient cho cells. transfected cells were treated with increasing concentrations of methotrexate (mtx) and clones secreting high levels of at-iii (10-20 micrograms/ml/3 day) in a serum-containing medium were obtained. serum-independent clo ...19921368211
effect of glycosylation on properties of soluble interferon gamma receptors produced in prokaryotic and eukaryotic expression systems.we investigated the influence of glycosylation on solubility, chromatographic behavior and resistance to heat- and chaotrope-dependent denaturation and proteolytic digestion of three recombinant human interferon gamma receptors produced in escherichia coli, spodoptera frugiperda and chinese hamster ovary cells. the proteins produced in the eukaryotic expression systems were glycosylated, carrying different, heterogeneous carbohydrate moieties. they were assayed fully glycosylated and after remov ...19921368793
two-dimensional electrophoresis as a tool for control of quality and consistency in production systems using animal cells. two-dimensional electrophoresis in animal cell culture technology.a nonrecombinant human melanoma cell line and recombinant chinese hamster ovary (cho) cells were used as examples for long-term in vitro cultivation in protein-free media. the method used to monitor the consistency of protein release by these mammalian cells was two-dimensional electrophoresis with immobilized ph gradient. secreted proteins from a melanoma cell line cultivated in a continuous fermentation system over a period of 22 months were monitored. two-dimensional patterns of secreted prot ...19921368812
a novel protease obtained from fbs-containing culture supernatant, that processes single chain form hepatocyte growth factor to two chain form in serum-free culture.the recombinant human hepatocyte growth factor (r-hhgf) produced by chinese hamster ovary cells transfected with hhgf cdna (cho bd-24 cells) was the two chain form in fetal bovine serum (fbs) containing culture. however, in serum-free culture the non-processed r-hhgf, single chain form, was detected with two chain form r-hhgf. we purified the protease that proteolytically processed single chain r-hhgf to two chain form r-hhgf. a protease was purified to give a single peak from the culture supern ...19921368819
gene transfer to tumor-infiltrating lymphocytes and other mammalian somatic cells by microprojectile bombardment.we demonstrate the application of particle delivery to the transformation of mammalian somatic cells. two mouse t-lymphocyte cell lines and chinese hamster ovary (cho) cells were transformed transiently with the rsv-adh (alcohol dehydrogenase) gene. stable gene transfer was also demonstrated in cho cells at a frequency of 6 x 10(-4) and in t-cells at a frequency of 1 x 10(-4), using beta-gal/neomycin-resistance gene constructs. the helium gas acceleration mechanism (pds 1000/he) and particle del ...19921369001
application of a statistical design to the optimization of culture medium for recombinant interferon-gamma production by chinese hamster ovary cells.the importance of serum-free medium components on the growth of chinese hamster ovary (cho) cells and production of recombinant human interferon(ifn)-gamma was investigated. the complexity of the medium led to the adoption of a statistical optimization approach based on a plackett-burman design. from this analysis a set of nutritional components was identified as important for cell growth and recombinant protein production. glycine was identified as an important determinant of specific growth ra ...19921369013
effect of amplification of dhfr and lac z genes on growth and beta-galactosidase expression in suspension cultures of recombinant cho cells.studies were conducted to characterize the effect of gene amplification and foreign gene expression on recombinant cho cell growth. chinese hamster ovary (cho) cells were transfected with an expression vector containing the gene for dihydrofolate reductase (dhfr) and the gene for human beta-interferon (beta-ifn) or the lac z gene which codes for beta-galactosidase (beta-gal). the recombinant genes in these cho cells were amplified stepwise by growth in 0, 10(-7), and 10(-6) m methotrexate (mtx), ...19921369176
construction of a toxic insulin molecule: selection and partial characterization of cells resistant to its killing effects.we have constructed an insulin-diphtheria hormono-toxin which migrates as a single 29 kd band on 10% sds polyacrylamide gel electrophoresis. this corresponds to a one to one molar ratio of the diphtheria a-chain (23 kda) and insulin (6 kda) molecules. the diphtheria a-chain: insulin (dtai) hormono-toxin demonstrates cytotoxicity in v-79 chinese hamster cells exhibiting an ld50 of 1.1 x 10(-8) m, which is 22 x more potent than whole diphtheria toxin. also, dtai can competitively displace [125i]-i ...19921369208
production of analytical quantities of recombinant proteins in chinese hamster ovary cells using sodium butyrate to elevate gene expression.sodium butyrate was used to enhance expression levels and thereby facilitate the generation of analytical quantities of nine different tissue plasminogen activator (tpa) analogues expressed under the control of the cytomegalovirus immediate early (cmv ie) promoter by the chinese hamster ovary (cho) mammalian expression system. production involved growth in roller bottles, using serum free or low serum media formulations, together with repetitive, sodium butyrate inductions. average inductions in ...19911369310
constitutive high-level production of human lymphotoxin by cho-k1 cells transformed with the human lymphotoxin gene controlled by a human beta-actin promoter.to achieve high-level production of human lymphotoxin (hlt), a plasmid (p beta lt-ldhfr) containing the hlt genomic dna, a mouse dihydrofolate reductase (dhfr) cdna, and a bacterial ecogpt gene was cotransfected with a plasmid (p beta ltml) encoding only the hlt genomic dna into chinese hamster ovary (cho-k1) cells at a 1:7 molar ratio. subsequently one of the ecogpt-positive clones (clone a31) was grown in stepwise increasing concentrations of methotrexate (mtx). a large amount of the hlt was s ...19911369325
comparison of human lymphotoxin gene expression in cho cells directed by genomic dna or cdna sequences.four recombinant plasmids coding for human lymphotoxin (lt) were constructed with genomic dna (gdna) or cdna sequences. the simian virus 40 (sv40) early region, which contains the early promoter, an intron of the small-t-antigen-encoding gene, and polyadenylation signal sequences, was used for transcriptional and post-transcriptional regulatory elements in the construction of these plasmids. two of them contained gdna and the other two contained cdna. one of the gdna plasmids and one of the cdna ...19901370034
tunicamycin treatment of cho cells abrogates multiple blocks to retrovirus infection, one of which is due to a secreted inhibitor.chinese hamster ovary (cho) cells are resistant to infection by all of the major classes of murine retroviruses and are partially resistant to infection by gibbon ape leukemia virus. treatment of cho cells with the glycosylation inhibitor tunicamycin rendered these cells susceptible to infection by retroviral vectors with ecotropic, xenotropic, and amphotropic host ranges and increased the titer of gibbon ape leukemia virus pseudotyped vectors 10-fold. vectors having a polytropic host range did ...19921370096
clastogenicity of lead chromate particles in hamster and human cells.several insoluble compounds of chromium, such as lead chromate, are respiratory carcinogens in experimental animals and suspected to be so in humans. lead chromate induces neoplastic transformation in cultured cells but the mechanism of genotoxicity is unknown. we examined the effect of lead chromate on the integrity of chromosomes of chinese hamster ovary (cho) and human foreskin fibroblasts (hff) after a 24-h exposure. at 0.4 microgram/cm2, 0.8 microgram/cm2, 2 microgram/cm2 and 8 microgram/cm ...19921370121
stable expression of monkey cytochrome p-450ia1 cdna in chinese hamster chl cells and its application for detection of mutagenicity of aflatoxin b1.a monkey cytochrome p-450ia1 cdna (mkah1) was transfected into chinese hamster chl cells using a vector containing the sr alpha promoter, and sublines stably expressing p-450ia1 were established. the cells showed 25-fold higher sensitivity to the cytotoxic effect of aflatoxin b1 than the parental chl cells. this hypersensitivity was almost completely suppressed by alpha-naphthoflavone, which is a known specific inhibitor of p-450ia. the cells expressing p-450ia1, but not chl cells, showed a posi ...19921370240
cd58 and cd59 molecules exhibit potentializing effects in t cell adhesion and activation.we have generated stable chinese hamster ovary (cho) cell transfectants expressing either cd58 or cd59 or both molecules to compare their respective parts played in t cell adhesion and activation. using a rosetting assay, we have shown the following: 1) the cd59 molecule was directly responsible for adhesive interaction between human t cells and cd59+ cho transfectants. cd59-mediated adhesion induced 12 +/- 2% (mean +/- sem, n = 25) of rosettes. 2) the cd58 molecule expressed on cd58+ cho transf ...19921370512
plant extracts induce chromosome aberrations and sister-chromatid exchanges in chinese hamster ovary cells and human lymphocytes.effects of extracts from vicia faba were compared with those of zea mays for the induction of sister-chromatid exchanges (sces) and of chromosome aberrations (cas) in chinese hamster ovary (cho) cells. ca induction by the maize extract was also tested in human lymphocytes. the extracts from roots and leaves of vicia faba induced cas and sces in cho cells. the extracts from maize leaves also induced sces and cas in cho cells, and cas in human lymphocytes. maize extracts were more potent in induci ...19921371591
antibodies against the cystic fibrosis transmembrane regulator.rabbit polyclonal antibodies have been raised against high-performance liquid chromatography purified synthetic peptides corresponding to two discrete regions of the cystic fibrosis transmembrane regulator (cftr) protein: the r-domain (residues 785-796) and the extreme cooh-terminus (residues 1467-1480). antibodies (ab) to each of these peptides were affinity purified either by passage over a peptide-derivatized agarose matrix (ab 785) to produce monospecific polyclonal antibodies or by protein ...19921371641
cloning and expression of a human atp-citrate lyase cdna.a full-length cdna clone of 4.3 kb encoding the human atp-citrate lyase enzyme has been isolated by screening a human cdna library with the recently isolated rat atp-citrate lyase cdna clone [elshourbagy et al. (1990) j. biol. chem. 265, 1430]. nucleic-acid sequence data indicate that the cdna contains the complete coding region for the enzyme, which is 1105 amino acids in length with a calculated molecular mass of 121,419 da. comparison of the human and rat atp-citrate lyase cdna sequences reve ...19921371749
biphenyl and fluorinated derivatives: liver enzyme-mediated mutagenicity detected in salmonella typhimurium and chinese hamster v79 cells.hepatocarcinogenic polychlorinated and polybrominated biphenyls usually show negative results in in vitro mutagenicity assays. problems in their testing result from their low water solubility and their slow rate of metabolism. we therefore investigated better soluble model compounds, namely biphenyl and its 3 possible monofluorinated derivatives. in the direct test, these compounds proved to be nonmutagenic in salmonella typhimurium ta98 and ta100 (reversion to histidine prototrophy) and in chin ...19921371835
differential effect of the amino acid cystine in cultured mammalian and bacterial cells exposed to oxidative stress.the effect of cystine in the cytotoxic response of cultured chinese hamster ovary and escherichia coli cells to challenge with hydrogen peroxide has been investigated. it was found that this amino acid could either protect or sensitize cells, depending on the cellular system. in fact, although a reduction in the growth-inhibitory effect of hydrogen peroxide was observed in mammalian cells, a marked increase in the susceptibility to oxidative stress was induced by cystine in bacteria. none of the ...19921371836
cultivation of mammalian cells on macroporous microcarriers.adherent cells can be cultivated in a stirred-tank bioreactor by attaching to microcarriers. macroporous microcarriers, with their intraparticle space and surface area for cell growth, can potentially support a higher cell concentration than conventional microcarriers, which support cell growth only on the external surface. chinese hamster ovary (cho) cells and green monkey kidney (vero) cells were cultivated on macroporous microcarriers, cultispher-g. cells attached to the microcarriers at a sl ...19921372509
functional expression and growth factor activation of an epitope-tagged p44 mitogen-activated protein kinase, p44mapk.mitogen-activated protein kinases (mapks) or extracellular signal-regulated kinases (erks) are serine/threonine kinases of apparent mr 42-44 kda that are rapidly activated by a variety of extracellular signals in many cell types. this activation coincides with their phosphorylation on tyrosine and threonine residues, and these covalent modifications are required for full activity of the enzymes. they are thought to play a pivotal role in integrating and transmitting transmembrane signals for gro ...19921372523
cloning and expression of a cdna for mouse prostaglandin e receptor ep3 subtype.a functional cdna clone for mouse ep3 subtype of prostaglandin (pg) e receptor was isolated from a mouse cdna library using polymerase chain reaction based on the sequence of the human thromboxane a2 receptor and cross-hybridization screening. the mouse ep3 receptor consists of 365 amino acid residues with putative seven-transmembrane domains. the sequence revealed significant homology to the human thromboxane a2 receptor. ligand binding studies using membranes of cos cells transfected with the ...19921372606
antibody and b7/bb1-mediated ligation of the cd28 receptor induces tyrosine phosphorylation in human t cells.cd28 is an adhesion receptor expressed as a 44-kd dimer on the surface of a major subset of human t cells. the cd28 receptor regulates the production of multiple lymphokines, including interleukin 2 (il-2), by activation of a signal transduction pathway that is poorly understood. here we show that ligation of cd28 by a monoclonal antibody (mab) or by a natural ligand, b7/bb1, induces protein tyrosine phosphorylation that is distinct from t cell receptor (tcr)-induced tyrosine phosphorylation. cd ...19921372649
characterization of the coexisting multiple mechanisms of methotrexate resistance in mouse 3t6 r50 fibroblasts.we have studied the discrepancy in the degree of methotrexate (mtx) resistance that exists between two clonal cell lines, mouse 3t6 r50 cells and chinese hamster ovary b11 0.5 cells that overexpress comparable levels of dihydrofolate reductase, yet exhibit a 100-fold difference in mtx resistance while maintaining similar sensitivity to the lipophilic antifolates trimetrexate and piritrexim. these data suggested that r50 cells may possess additional mechanism(s) of antifolate resistance, such as ...19921372892
effects of benzyladenine on prokaryotic and eukaryotic cells.comparative studies of the effect of benzyladenine (ba) on the yeast saccharomyces cerevisiae, the bacterium salmonella typhimurium, the shallot allium ascalonicum and chinese hamster fibroblast cells were performed. the tested substance had no mutagenic activity on yeast, bacteria and cultured fibroblast cells. changes in mitotic activity and cell division abnormalities were observed after ba treatment in shallot root-tip cells.19921373222
contribution of the n-linked carbohydrate of erythrocyte antigen cd59 to its complement-inhibitory activity.the contribution of n-linked carbohydrate to the complement-inhibitory function of the human erythrocyte membrane glycoprotein, cd59, was investigated. amino acid sequence analysis of tryptic peptides labeled with [3h]borohydride revealed an n-linked carbohydrate moiety at the asn18 residue. no o-linked carbohydrate was detected, as judged by the failure of asialo-cd59 to bind peanut agglutinin and by its resistance to digestion by o-glycanase. the apparent molecular mass of cd59 was reduced fro ...19921373727
chromosomal changes in chinese hamster aa8 cells caused by podophyllin, a common treatment for genital warts.podophyllin, a plant derivative of variable composition, is used widely within new zealand as a treatment for genital warts. one local source of podophyllin has been tested for ability to cause mutagenic effects in salmonella typhimurium as well as for effects on chromosomes of chinese hamster aa8 cells. although only weakly mutagenic in one strain of salmonella, podophyllin caused structural aberrations as well as changes in chromosome number in the chinese hamster cells. the range of aberratio ...19921373833
predictive assay for rodent carcinogenicity using in vivo biochemical parameters: operational characteristics and complementarity.111 chemicals of known rodent carcinogenicity (49 carcinogens, 62 noncarcinogens), including many promoters of carcinogenesis, nongenotoxic carcinogens, hepatocarcinogens, and halogenated hydrocarbons, were selected for study. the chemicals were administered by gavage in two dose levels to female sprague-dawley rats. the effects of these 111 chemicals on 4 biochemical assays (hepatic dna damage by alkaline elution (dd), hepatic ornithine decarboxylase activity (odc), serum alanine aminotransfera ...19921373835
the shape of dna elution dose-response curves under non-denaturing conditions: the contribution of the degree of chromatin condensation.we have re-examined the effect of detergent type, ph and temperature of lysis on the shape of the dna elution dose response under non-denaturing conditions using plateau-phase cho cells. results practically identical to those previously published (okayasu and iliakis, 1989) were obtained, with a 1 h incubation at 60 degrees c during lysis with sodium-n-laurylsarcosine (nls) resulting in almost linear dose-response curves. we also examined chromatin decondensation as a contributing factor in the ...19921374110
purification and characterization of soluble forms of human and rat stem cell factor recombinantly expressed by escherichia coli and by chinese hamster ovary cells.stem cell factor (scf) is a novel, early-acting hematopoietic factor. it was isolated from the medium of a rat cell line in a soluble, processed form (zsebo et al., 1990, cell 63, 195). the cloned human and rat genes encode the soluble form plus additional c-terminal amino acids including a hydrophobic transmembrane domain (martin et al., 1990, cell 63, 203). we have recombinantly expressed forms of human and rat scf corresponding to the soluble, processed form in escherichia coli and in chinese ...19921374224
beta 1,4-galactosyltransferase: a short nh2-terminal fragment that includes the cytoplasmic and transmembrane domain is sufficient for golgi retention.beta 1,4-galactosyltransferase (beta 1,4-gt) is a golgi-resident, type ii membrane-bound glycoprotein that functions in the coordinate biosynthesis of complex oligosaccharides. additionally, beta 1,4-gt has been localized to the cell surface of a variety of cell types and tissues where it is proposed to function in intercellular recognition and/or adhesion. thus beta 1,4-gt is an appropriate molecule to be used in analyzing the molecular basis for retention of a membrane-bound enzyme in the golg ...19921374389
cyclic amp-dependent protein kinase activation of cystic fibrosis transmembrane conductance regulator chloride channels in planar lipid bilayers.membrane vesicles, prepared from mouse nih-3t3 fibroblasts and chinese hamster ovary cells expressing high levels of cystic fibrosis transmembrane conductance regulator (cftr), were fused with mueller-rudin planar lipid bilayers. upon addition of the catalytic subunit of camp-dependent protein kinase and atp, low conductance cl(-)-selective ion channels were observed in 10 of 16 experiments. the channels had a linear current-voltage relationship and a unitary conductance of approximately 6.5 ps. ...19921374403
gene amplification in the murine sewa system.considerable work with dna amplification has been carried out in the murine sewa ascites tumor cell system. in sewa cells there is 'spontaneous' amplification of the c-myc oncogene, and transitions between different cytogenetic expressions of gene amplification such as dm (double minutes), cm (c-bandless chromosomes) and hsr (homogeneously staining regions) of the amplified dna have been recorded during serial in vivo transplantations. in sewa cells it has also been shown that the c-myc-containi ...19921374520
the mechanism of carcinogen-induced dna amplification: in vivo and in vitro studies.exposure to chemical carcinogens provides a means for the enhancement of the frequency of gene amplification and for the facilitation of research into its mechanism(s). using carcinogen-induced sv40 amplification as a model system it was shown that amplification of the viral sequences occurs via a replication-dependent mode. this process involves overactivation of the origin region and the generation of inverted repeats. carcinogen-induced enhancement of gene amplification is triggered by cellul ...19921374526
sensitivity and single-strand dna break repair in chinese hamster mutants exposed to the carcinogen aflatoxin b1 epoxide and its dichloride model.aflatoxin b1 (afb1) is a potent carcinogen and mutagen. it requires metabolic activation to be converted to the dna-binding product aflatoxin b1 epoxide (afb1-epoxide). a model of this epoxide is aflatoxin b1 dichloride (afb1cl2). both react at the n7 position of guanine to form large adducts. the major adduct formed can either be rapidly removed to leave an apurinic site or can undergo ring opening of the imidazole ring to form a chemically stable adduct. a number of chinese hamster dna repair- ...19921374845
immunoreactivity of recombinant carcinoembryonic antigen proteins expressed in escherichia coli.immunoreactivities of recombinant carcinoembryonic antigen (cea) proteins expressed in escherichia coli (e. coli) were analyzed in relation to the cea domain structure [domains n, i (a1-b1), ii (a2-b2), iii (a3-b3) and m]. we reconstructed in a prokaryotic expression vector, pucpl-ci, the cdnas fro cea-n, cea-i, cea-ii, and cea-iii-m. the latter three were expressed as fusion products with bacterial beta-galactosidase. the recombinant proteins were solubilized by sonication in 1% sodium dodecyl ...19921375188
nuclease modification in chinese hamster cells hypersensitive to dna cross-linking agents--a model for fanconi anemia.fanconi anemia is a human inherited disease that is characterized by cellular hypersensitivity to dna cross-linking agents. a number of potential experimental models for that disorder have been developed by selecting mutants that are hypersensitive to bifunctional mutagens. the six mutants of that class in drosophila, all of which map to the mus308 locus, express an alteration in a mitochondrial nuclease. a recent extension of that observation to cell lines from complementation group a of fancon ...19921375328
cytotoxicity and genotoxicity testing of sodium fluoride on chinese hamster v79 cells and human eue cells.the cytotoxic effects of sodium fluoride (naf) on hamster v79 cells and human eue cells were studied by measuring the cloning efficiency and dna, rna and protein synthesis in cells cultured in the presence of naf. potential mutagenicity of naf was followed on the basis of induced 6-thioguanine-resistant mutants in treated chinese hamster v79 cells. the results showed that the addition of 10-150 micrograms of naf per ml of culture medium induced 10-75% cytotoxic effect on hamster v79 cells but ha ...19921375335
protective effect of two sunscreens against lethal and genotoxic effects of uvb in v79 chinese hamster cells and saccharomyces cerevisiae strains xv185-14c and d5.effects of p-aminobenzoic acid (paba) and of 4-[(2-oxo-3-bornylidene)methyl]-phenyl trimethylammonium methylsulfate (omm), two components used in sunscreen formulations, on the mutagenicity of uvb irradiation are compared in three genetic assay systems. a haploid strain of saccharomyces cerevisiae xv185-14c was used to measure reverse mutations at three loci. the diploid strain d5 of saccharomyces cerevisiae was used to screen for reciprocal mitotic recombination. the induction of forward mutati ...19921375337
sulfated polymers inhibit the interaction of human cytomegalovirus with cell surface heparan sulfate.several sulfated polysaccharides (dextran sulfate, pentosan polysulfate, heparin) and copolymers of acrylic acid with vinylalcohol sulfate have proved to be potent inhibitors of human cytomegalovirus (cmv) infectivity in vitro. sulfated alpha-cyclodextrins are only weak inhibitors of cmv. a close correlation was found between the 50% inhibitory concentrations of the sulfated polymers for cmv cytopathogenicity, virus-cell binding, and expression of immediate early antigens (iea) in human embryoni ...19921376540
dna double labelling with idurd and cldurd for spatial and temporal analysis of cell proliferation and dna replication.a procedure was developed that very effectively distinguishes between idurd and cldurd incorporated in the dna of cell nuclei and chromosomes. for double staining we used the rat anti-brdurd monoclonal antibody from sera-lab that binds specifically to cldurd and brdurd but not to idurd, in combination with the mouse anti-brdurd monoclonal antibody from becton dickinson. this antibody binds to all three halogenated deoxyuridines, but when the nuclei are washed in tris buffer with a high salt conc ...19921376726
expression and chromosome localization of the murine cystic fibrosis transmembrane conductance regulator.a 13.5-kb genomic fragment of the mouse cystic fibrosis transmembrane conductance regulator (cftr) gene was isolated from a c57bl/6j liver dna library, using a human cftr exon 10 probe. this region of the human gene includes the most common cystic fibrosis mutation (deletion of the phe508 residue) in the first nucleotide binding domain of cftr. sequence analysis demonstrated 87% identity between the predicted mouse and the normal human cftr exon 10 sequences, including conservation of the phe508 ...19921377165
determination of the amino acid residues in substance p conferring selectivity and specificity for the rat neurokinin receptors.we have measured the affinity of various analogs and fragments of the tachykinin substance p for the cloned rat nk1, nk2, and nk3 receptors heterologously expressed in chinese hamster ovary cells. the hydrophobic carboxyl-terminal pentapeptide sequence substance p-(7-11) binds with similar affinity (2-20 microm) to all three receptors. our data suggest that addition of one to three amino-terminal residues to this sequence results in the optimization of its interaction within the binding pocket o ...19921377326
mutation induction and dna adduct formation in chinese hamster ovary cells treated with 6-nitrochrysene, 6-aminochrysene and their metabolites.6-nitrochrysene, 6-aminochrysene and several of their metabolites were assayed for mutagenic activity at the hypoxanthine-guanine phosphoribosyl transferase (hprt) locus in dna-repair-proficient chinese hamster ovary (cho-k1) cells and excision-repair-deficient cho-uv5 cells. mutagen-dna adducts were analyzed by 32p-postlabeling in cells treated under the conditions of the mutagenicity assay and compared with the adduct patterns produced from the in vitro reaction of metabolites of 6-nitrochryse ...19921377330
evaluation of genotoxicity of tert.-butylhydroquinone in an hepatocyte-mediated assay with v79 chinese hamster lung cells and in strain d7 of saccharomyces cerevisiae.tert.-butylhydroquinone (tbhq) has been reported to be genotoxic in some short-term assays but non-genotoxic in others. we have examined cytotoxicity and genotoxicity of tbhq, a principal metabolite of the phenolic antioxidant 2(3)-tert.-butyl-4-hydroxyanisole (bha), in an hepatocyte-mediated assay with v79 chinese hamster lung cells including both sister-chromatid exchange (sce) and thioguanine-resistance (tgr) endpoints. the ability of bha and of tbhq to elicit a genotoxic response in saccharo ...19921377341
simultaneous analysis of chromosome damage and aneuploidy in cytokinesis-blocked v79 chinese hamster lung cells using an antikinetochore antibody.a modified antikinetochore antibody technique was established in the v79 chinese hamster lung cells to simultaneously analyze chromosome damage and aneuploidy induced by various agents. the method involved sequential treatment of slides with crest serum, fluoresceinated goat-antihuman and swine-antigoat antibodies, and propidium iodide. in this method, cytoplasm (green), nuclei or micronuclei (red), and kinetochores (yellow), are identified using the same filter setting under blue excitation (44 ...19921377355
genotoxicity of n-acryloyl-n'-phenylpiperazine, a redox activator for acrylic resin polymerization.n-acryloyl-n'-phenylpiperazine is a promoter of redox reactions synthesized recently, and proposed as an activator for the polymerization of acrylic resins for biomedical use. the chemical was analyzed for different genotoxicity endpoints, to obtain both information on its possible mutagenic/carcinogenic potential and a model analysis of a tertiary arylamine, which belongs to a class of chemicals commonly used as polymerization accelerators in the biomaterial field. the genotoxicity endpoints co ...19921377358
epitopes predominantly retained on the carcinoembryonic antigen molecules in plasma of patients with malignant tumors but not on those in plasma of normal individuals.we have previously reported that a group of monoclonal antibodies (mabs) to carcinoembryonic antigen (cea), designated group f mabs, are able to discriminate cea in tumor tissues from normal fecal antigen-2, a soluble form cea-counterpart in normal adult feces, and that the protein epitopes recognized by them are present on the domain a3-b3 of the cea molecule. in this study, we further investigated the molecular localization of the epitopes recognized by the group f mabs using three new recombi ...19921377668
spectrum of sialylated and nonsialylated fuco-oligosaccharides bound by the endothelial-leukocyte adhesion molecule e-selectin. dependence of the carbohydrate binding activity on e-selectin density.carbohydrate recognition by the human endothelial-leukocyte adhesion molecule, e-selectin, has been investigated by binding studies using 3h-labeled chinese hamster ovary cells expressing different levels of the transfected full-length adhesion molecule and a series of structurally defined oligosaccharides linked to the lipid phosphatidylethanolamine dipalmitoate (neoglycolipids) and synthetic glycolipids chromatographed on silica gel plates or immobilized on plastic wells. evidence is presented ...19921377689
the hinge region of the cd8 alpha chain: structure, antigenicity, and utility in expression of immunoglobulin superfamily domains.the lymphocyte surface cd8 antigen is a heterodimer with each chain containing a single ig-related domain, a hinge-like sequence, a transmembrane segment, and a short cytoplasmic sequence. a soluble form of the rat cd8 alpha chain was produced by introducing a stop codon into the cdna at the end of the region encoding the extracellular sequence and expressed in chinese hamster ovary cells. scd8 alpha was produced at 20 mg/l, and consisted of monomers, dimers, and higher aggregates. the latter co ...19921377946
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