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Biomedical subjects

M Kuroki

Publications and source records attributed to M Kuroki.

At least 217 records · Page 12Linked to original sources
PubMed1986

Production of immunoreactive calcitonin and some other tumor markers by established human carcinoma cell lines.

Out of seven human carcinoma cell lines (M7609, CCK-81, FCC-1, RPMI#4788, QGP-1, HLC-1, and KNS-62), 4 cell lines were found to produce immunoreactive calcitonin (ICT), a potential tumor marker for various malignancies. During a 7-day culture, 1.4 X 10(5) QGP-1, RPMI#4788, HLC-1, and KNS-62 cells secreted 7,000 pg, 500 pg, 400 pg, and 400 pg of ICT in the medium, respectively. The production of ICT by QGP-1 cells was increased by addition of pentagastrin or calcium gluconate. Three different components of ICT (peak I, molecular weight greater than 40,000; peak II, 14,000-18,000; peak III, 3,400) were detected by gel filtration of the QGP-1 spent medium. In a competitive inhibition-type radioimmunoassay of serial dilutions of each ICT component, peak III component showed very similar immunoreactivity to synthetic calcitonin. However, the other two components gave clearly different immunoreactivities from the peak III component and showed very similar immunoreactivities to each other. All the cell lines were further screened for synthesis of 7 other tumor markers, carcinoembryonic antigen, nonspecific cross-reacting antigen, CA19-9, tissue polypeptide antigen, alpha-fetoprotein, beta 2-microglobulin and ferritin. Every cell line produced 2 to 6 markers concomitantly, and various combinations of positive markers were found among the cell lines.

Calcitonin↗
PubMed1985

Further comparative studies on chemical properties of carcinoembryonic antigen in tumor tissues and closely related antigens in adult feces and meconium.

The chemical structure of carcinoembryonic antigen (CEA) and two closely related antigens, normal fecal antigen-2 (NFA-2) in normal adult feces and nonspecific cross-reacting antigen-2 (NCA-2) in the meconium, were further analyzed comparatively. The NH2-terminal amino acid sequence of NCA-2 was newly determined to position 18 and found to be identical to that so far determined for CEA- and NFA-2. After proteolytic digestion with chymotrypsin or protease V8, the digests of these antigens showed two groups of fragments upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis. One consisted of the sharply banded fragments which were identical in all antigens and stained only with Coomassie brilliant blue (CBB) (five bands in the range 2500-10,000 daltons for chymotrypsin and 11 bands in the range 8000-35,000 daltons for protease V8, respectively), and the other consisted of the dispersed fragments which had variable mol. wts in the range 10,000-100,000 and were stainable with both CBB and periodic acid-Schiff reagent. Elution profiles of CEA, NFA-2, and NCA-2 from lectin columns, especially from concanavalin A-Sepharose columns, suggested some differences in oligosaccharide chains between them. These results indicate that the fundamental chemical structure of these antigens seems to be very similar to one another and is divided into two parts; an homologous portion(s) which is common to all three antigens and contains no sialylated sugar components, and a heterogeneous portion(s) which is variable among these antigens and contains sialylated sugar components.

Adult↗
PubMed1985

Effect of wall shear stress on intimal thickening of arterially transplanted autogenous veins in dogs.

To determine whether or not changes in wall shear stress play a determinant role in the induction of hyperplasia of intimal tissue of arterially transplanted vein grafts, we developed two models of canine femoral arteries. Wall shear stress was defined by variation of wall shear stress (tau-variation) in the cardiac cycle, with the use of a newly designed computational flow waveform analyzer. In the group I model autogenous vein grafts were implanted under flow conditions of 79.7 +/- 3.2 ml/min of the normally high flow rate with 33.1 +/- 1.9 dynes/cm2 of low tau-variation. In the group II model grafts were implanted under conditions of 2.9 +/- 1.8 ml/min of low flow rate with 178.8 +/- 11.0 dynes/cm2 of normally high value of tau-variation. The intimal thickness of 259 +/- 36 microns 4 weeks after implantation in group I was statistically significant compared with that of 31 +/- 14 microns in group II (p less than 0.005). Our study revealed that change in wall shear stress and not the rate of blood flow is the essential hemodynamic factor related to intimal hyperplasia.

Animals↗
PubMed1984

Identification and partial characterization of the unglycosylated peptide of carcinoembryonic antigen synthesized by human tumor cell lines in the presence of tunicamycin.

Unglycosylated peptide backbones of carcinoembryonic antigen (CEA) synthesized by human tumor cell lines in the presence of tunicamycin were identified and analyzed by SDS-polyacrylamide gel electrophoresis. Three tumor cell lines, QGP-1 (pancreas), FCC-1 (colon) and KNS-62 (lung) were found to produce CEA molecules of 180,000-190,000 mol. wts labeled with both [3H]leucine and [14C]glucosamine under conventional culture conditions. In contrast, in the presence of tunicamycin, the native CEA molecules disappeared, and a new component that was precipitated with anti-CEA antibodies and labeled only with [3H]leucine but not with [14C]glucosamine was identified in each cell line. Monoclonal antibodies each directed to different major antigenic determinants on the native CEA molecules also reacted with this unglycosylated peptide. The apparent mol. wts of the naked CEA peptides from QGP-1 and FCC-1 were equally about 78,000, whereas that from KNS-62 was somewhat larger than the other two, suggesting some differences in the peptide structure of the CEA molecules.

Antibodies, Monoclonal↗
PubMed1984

Monoclonal antibodies to carcinoembryonic antigen: a systematic analysis of antibody specificities by using related normal antigens and evidence for allotypic determinants on carcinoembryonic antigen.

Eleven hybridoma clones producing monoclonal antibodies to carcinoembryonic antigen (CEA) were prepared by fusions of the mouse myeloma cell line P3-X63-Ag8-U1 with splenocytes of BALB/c mice immunized with highly purified CEA. Their specificities were systematically analyzed by radioimmunoassay for reactivity with CEA and four related normal antigens: nonspecific cross-reacting antigen (NCA), nonspecific cross-reacting antigen-2 (NCA-2), normal fecal antigen-1 (NFA-1), and normal fecal antigen-2 (NFA-2). Antibody-antigen profiles revealed four groups of hybridomas. The Group I antibodies from two clones reacted with sites shared among CEA, NCA, NCA-2, and NFA-2. The Group II antibody from one clone recognized an epitope common to CEA, NCA-2, and NFA-2. The Group III antibodies from five clones reacted with sites commonly residing on CEA, NCA-2, NFA-1, and NFA-2, whereas antibodies of Group IV from three clones bound only CEA. Mutual inhibition assays between antibodies in the respective groups for CEA binding further revealed that at least eight different epitopes can be identified on the CEA molecule: two in Group I, one in Group II, three in Group III, and two in Group IV, respectively. Three monoclonal antibodies in Group IV appeared to be specific for CEA, but they did not react with any of five purified CEA preparations other than that used for immunization. In 13 sera from 50 various cancer patients with elevated CEA levels, however, we detected the CEA molecule reactive with these antibodies, indicating that the epitopes recognized by the Group IV antibodies may be of allotypic characters. These epitopes were unrelated to main blood group antigens and did not show any organ specificity. All of the eight epitopes identified in this study were found to be resistant to neuraminidase, mixed glycosidases, and Smith degradation (SI-stage) as determined by a competitive radioimmunoassay, suggesting that they reside on the protein moiety of the CEA molecule. Six epitopes were sensitive to reduction and alkylation, but two other epitopes (one in Group II and the other in Group III) were resistant to these treatments. This result indicates that the latter two epitopes are conformation-independent.

Adult↗
PubMed1984

A desktop computer to visualize the intraluminal velocity profile and its clinical application.

Based on our previous study that the outcome of reconstructive surgery for lower limbs could well be predicted by flow waveforms and that this state was defined by variation of wall shear stress (tau) in a cardiac cycle, we set up a desktop computer that could calculate wall shear stress in a given patient. This work is based on a principle of a computational method for simulating pulsating blood flow. The flow data from the patient are sent to the waveform analyzer, where the intraluminal velocity profile as well as the shear stress variation is demonstrated. Our study revealed no late occlusion in patients with flow waveforms over 70 dynes/cm2 in tau-variation, early graft failure in those below 20 dynes/cm2, and late occlusion in considerable numbers of patients whose waveforms were between 70 and 20 dynes/cm2. Intraoperative use of the waveform analyzer disclosed the occult cause of obstruction, which would otherwise be overlooked, leading to a remarkable decrease in the rate of immediate graft failure. Follow-up of outpatients with this approach facilitated early detection of late failure, making it possible to repair the graft with minor surgery. Use of the waveform analyzer proved superior to ankle pressure index in predicting graft failure.

Aged↗
PubMed1984

A new human osteosarcoma strain (Os-Beppu) with a high rate of pulmonary metastasis in nude mice.

A new human osteosarcoma strain (Os-Beppu) transplantable in nude mice has been established. A high frequency of metastasis (87.2%) to the lung was observed throughout nine passages. Histologically the subcutaneous tumor and the metastatic lesions in nude mice were similar to the original human tumor, and osteoid and osteoclast formation were preserved. Osteoblasts were strongly positive for alkaline phosphatase, while osteoclasts were positive for acid phosphatase. Karyotype analysis revealed human chromosomes with a modal chromosome number of 41 (63.3%) and two marker chromosomes.

Animals↗
PubMed1983

Antigenic reactivities of purified preparations of carcinoembryonic antigen (CEA) and related normal antigens using four different radioimmunoassay systems for CEA.

Antigenic reactivities of 4 purified carcinoembryonic antigen (CEA) preparations including the WHO standard CEA and 4 CEA-related antigens--nonspecific cross-reacting antigen (NCA) from normal lungs, nonspecific cross-reacting antigen-2 (NCA-2) from meconium, normal fecal antigen-1 (NFA-1) and normal fecal antigen-2 (NFA-2) from normal adult feces--were investigated using 4 different radioimmunoassay (RIA) systems (Hoffman-La Roche, CEA-IRE-SORIN, Dainabot and our system). In general, all CEA preparations including the respective standard CEAs gave similar reactions with these assay systems. Differences in reaction intensity were, however, observed among the CEA preparations, and the order of reaction intensity among the preparations varied due to assay systems, e.g., a given CEA preparation which reacted most strongly with an assay system reacted very weakly with another assay system, and vice versa. Both NCA-2 and NFA-2, which possessed physiochemical properties very similar to those of CEA, revealed reactivities indistinguishable from those of the CEA preparations in all systems. NCA, which showed a partial cross-reactivity with CEA in the double immunodiffusion test, revealed a remarkably weak reactivity in all systems. NFA-1, however, which was also partially cross-reactive with CEA but antigenically unrelated to NCA, showed a fairly strong reactivity in all systems, but its dose-dependent slope was apparently different from that of the other antigen preparations.

Antigen-Antibody Reactions↗
PubMed1983

A new direct solid-phase radioimmunoassay for carcinoembryonic antigen without pretreatment of serum samples.

We examined some fundamental conditions for establishing a new simplified radioimmunoassay system for CEA. The system consists of antibody-coated plastic beads and radiolabeled tracer antibody. The reactivity of this system with several purified CEA preparations can vary due to differences in the nature of antibody preparations used either for coating beads or for tracer antibody. Among purified CEA preparations, we also found some differences in reactivity with a given immobilized antibody preparation. The assay system that resulted, referred to as bead-direct-RIA, needs neither pretreatment of serum samples nor centrifugation in any of its steps, and has a wide assay range, measuring CEA concentrations of 1-300 ng/ml using only 50 microliters of a serum sample. Its accuracy, reproducibility, dilution effect, and analytical recovery proved the reliability of this method. The mean value +/- S.D. for 48 normal adults was 2.5 +/- 1.0 ng/ml. It was found by comparative studies that the Z-gel method of Hoffmann-La Roche generally gave the highest values, the antibody-disc method of Dainabot gave the lowest, and the values by bead-direct-RIA fell between those of the 2 other methods and correlated well with either value. Follow-up studies on patients with malignancy indicated that the values by bead-direct-RIA fluctuated in parallel with those by Roche and Dainabot kits.

Antibodies, Neoplasm↗
PubMed1983

[Antigenic structures of CEA and CEA-related normal antigens and problems related to the determination of blood CEA concentrations].

The antigenic structures of carcinoembryonic antigen (CEA) from tumor tissues, and of CEA-related antigens in normal adult feces and meconium were analyzed. Four CEA-related antigens were isolated from normal adult feces, three of them were inclusively called as normal fecal antigen (NFA) consisted of NFA-1, NFA-2 and NFCA (normal fecal cross-reacting antigen), the other was fecal NCA. NCA-2 was also isolated from meconium. CEA, NFA-2 and NCA-2 were very similar to each other physiochemically and antigenically, but CEA-distinctive and NFA-2-distinctive determinants were detected by using specifically prepared antisera. No distinctive determinant on NCA-2 was found so far tested with our anti-NCA-2 antisera. Both NFA-1 and NCA were partially cross-reactive with CEA, NFA-2 or NCA-2, but they were antigenically independent of each other. NFCA were partially cross-reactive with both NFA-2 and NCA. From these results, it was concluded that CEA and NFA-2 could be divided into four antigenic moieties, that is, NCA-common, NFCA-common, NFA-1-common determinant and CEA-distinctive or NFA-2-distinctive determinant. Among these, NFA-1-common and NFCA-common determinants seem to be dominant as judged from the reactivities with conventional anti-CEA antisera. Recently, we established a new solid-phase radioimmunoassay for CEA. It was found that the reactivity of the system with several purified CEA preparations could vary due to differences in nature, especially in the affinity to antigen, of antibody preparations used either for solid-phase or for tracer antibody. Among purified CEA preparations, some differences in reactivity with our assay system were also found. Reactivities of four CEA-assay systems, three commercially available and ours, with purified CEA and related antigen preparations were tested. It was found that the order of reaction intensity among antigen preparations varied due to assay systems, e.g. a given CEA preparation which reacted most strongly with an assay system reacted very weakly with another assay system, and vice versa. From these results, the problems concerned with variations in estimated CEA values due to assay methods were discussed.

Amino Acid Sequence↗
PubMed1983

Transplantable choriocarcinoma of rats induced by fetectomy and its biological activities.

Eight cases of choriocarcinoma were induced in WKA female rats by simple fetectomy without viral inoculation or administration of chemical carcinogens. Among them, one line has been successfully subcutaneously transplanted over 10 generations in syngeneic rats and this line was designated as RCHO. The histological appearance of RCHO consisted of trophoblastic giant cells and small basophilic tumor cells, which resembled normal rat placenta. RCHO possessed marked mammotropic and lactogenic activities in female recipients and induced atrophy of the testes in male recipients.

Animals↗
PubMed1982

Measurement of membrane potential in polymorphonuclear leukocytes and its changes during surface stimulation.

The membrane potential of guinea pig polymorphonuclear leukocytes has been assessed with two indirect probes, tetraphenylphosphonium (TPP+) and 3,3'-dipropylthiadicarbocyanine (disS-C3-(5)). The change in TPP+ concentration in the medium was measured with a TPP+-selective electrode. By monitoring differences in accumulation of TPP+ in media containing low and high potassium concentrations, a resting potential of -58.3 mV was calculated. This potential is composed of a diffusion potential due to the gradient of potassium, established by the Na+, K+ pump, and an electrogenic potential. The chemotactic peptide fMet-Leu-Phe elicits a rapid efflux of accumulated TPP+ (indicative of depolarization) followed by its reaccumulation (indicative of repolarization). In contrast, stimulation with concanavalin A results in a rapid and sustained depolarization without a subsequent repolarization. The results obtained with TPP+ and diS-C3-(5) were comparable. Such changes in membrane potential were observed in the absence of extracellular sodium, indicating that an inward movement of sodium is not responsible for the depolarization. Increasing potassium levels, which lead to membrane depolarization, had no effect on the oxidative metabolism in nonstimulated or in fMet-Leu-Phe-stimulated cells. Therefore, it seems unlikely that membrane depolarization per se is the immediate stimulus for the respiratory burst.

Animals↗
PubMed1982

Immunological characterization and structural studies of normal fecal antigen-1 related to carcinoembryonic antigen.

Normal fecal antigen-1 (NFA-1), which is a carcinoembryonic antigen (CEA)-related glycoprotein with a mol. wt of 20,000-30,000, was purified from normal adult feces by immuno-adsorption, gel filtration and ion exchange chromatography. Highly purified NFA-1 was partially cross-reactive with CEA but antigenically unrelated to nonspecific cross-reacting antigen (NCA) which was also cross-reactive with CEA. NFA-1 also had a unique determinant not present in CEA or other related antigens including NCA. In a solid-phase RIA system, the reactivity of NFA-1 with a specific anti-CEA antiserum was much stronger than that of NCA. Although digestion with Pronase E did not affect the antigenicity of NFA-1, reduction and alkylation destroyed its antigenic reactivity. The total amount of carbohydrate in NFA-1 was 13.3%, compared to 52.4% in CEA and 21.6% in NCA. The amino acid composition of NFA-1 was similar to that of CEA. The sequence of the first 10 NH2-terminal amino acids in NFA-1 was Ala-Glu-Pro-Pro-Lys-Pro-Phe-Ile-(Thr)-Ser. This was totally different from that of the first NH2-terminal amino acids of CEA isolated from tumor tissue.

Amino Acid Sequence↗
PubMed1982

Immunochemical differences among carcinoembryonic antigen in tumor tissues and related antigens in meconium and adult feces.

We have isolated carcinoembryonic antigen (CEA)-related antigens from meconium and compared them with those in adult feces. Two CEA-related antigens were detected in meconium [nonspecific cross-reacting antigen 2 (NCA-2) and meconium nonspecific cross-reacting antigen] while four CEA-related antigens were found in adult feces [normal fecal antigen 1, normal fecal antigen 2 (NFA-2), normal fecal cross-reacting antigen, and fecal nonspecific cross-reacting antigen, respectively]. By conventional anti-CEA antisera, NCA-2 in meconium, NFA-2 in adult feces, and CEA in tumor tissues were indistinguishable from each other, but they could be distinguished by specific antibody preparations against a determinant unique to CEA (CEA-distinctive determinant) or to NFA-2 (NFA-2-distinctive determinant). Neither the CEA-distinctive determinant nor the NFA-2-distinctive determinant was detected on the NCA-2 molecule. No antigenic determinants unique to NCA-2 have been detected with the anti-NCA-2 antisera which we have prepared thus far. The molecular weight of purified NCA-2 was estimated to be 150,000 to 170,000 as compared to 160,000 to 170,000 for NFA-2 and 170,000 to 180,000 for CEA. NCA-2 had amino acid and carbohydrate compositions similar to those of CEA and NFA-2. All NFA-2 preparations and about one-half of the CEA preparations were sensitive to Pronase E digestion, which released two antigen fragments from these molecules, but NCA-2-preparations were resistant to such digestion.

Amino Acids↗
PubMed1982

Coproduction of carcinoembryonic antigen and nonspecific cross-reacting antigen by a continuous cell line from a human pancreatic tumor.

A simultaneous production of nonspecific cross-reacting antigen (NCA) and carcinoembryonic antigen (CEA) by the same individual cells of an established human pancreatic cell line (QGP-1) was demonstrated by the immunoperoxidase method. Kinetics of cell proliferation and production of CEA and NCA were analyzed, and active synthesis of both antigens was found to be accompanied with the active proliferation of cultured cells. Both antigens in culture medium were purified by immunoadsorption and gel filtration. Immunochemical studies confirmed that CEA and NCA produced by the QGP-1 cells had properties identical to those of authentic CEA derived from metastatic colorectal carcinoma and to those of NCA from normal lungs, respectively.

Antigens, Neoplasm↗