Search PubMed⌕ Search

Biomedical subjects

R Kannagi

Publications and source records attributed to R Kannagi.

At least 181 records · Page 10Linked to original sources
PubMed1984

Characterization of a glycosphingolipid antigen defined by the monoclonal antibody MBr1 expressed in normal and neoplastic epithelial cells of human mammary gland.

The antigen defined by a monoclonal antibody, MBr1, was found to be expressed in normal human mammary gland epithelia and human mammary carcinoma cells (Ménard, S., Tagliabue, E., Canevari, S., Fossati, G., and Colnaghi, M. I. (1983) Cancer Res. 43, 1295-1300). The antigen has been isolated from breast cancer cell line MCF-7, which was used as immunogen, and its structure was determined by methylation analysis, NMR spectroscopy, direct probe mass spectrometry, and enzymatic degradation as identified below. Fuc alpha 1----2Gal beta 1----3GalNAc beta 1----3Gal alpha 1----4Gal beta 1----4Glc beta 1----1Cer The antibody cross-reacted weakly with fucosylasialo-GM1 (IV2FucGg4), which shares the same terminal sequence, Fuc alpha 1----2Gal beta 1----3GalNAc, with this antigen. However, various other structures, including lacto-series H structure (Fuc alpha 1----2 Gal beta 1----4/or 3GlcNAc beta 1----3Gal), did not show any reactivity with this antibody. Therefore, this antigen represents a blood group H antigen with a globo-series structure which is abundant in human teratocarcinoma (Kannagi, R., Levery, S. B., Ishigami, F., Hakomori, S., Shevinsky, L. H., Knowles, B. B., and Solter, D. (1983) J. Biol. Chem. 258, 8934-8942), although its presence must be limited in normal adult human tissue.

Antibodies, Monoclonal↗
PubMed1984

Blood group A glycolipid (Ax) with globo-series structure which is specific for blood group A1 erythrocytes: one of the chemical bases for A1 and A2 distinction.

A new blood group A-active glycolipid fraction, termed Ax, showing a chromatographic mobility between Aa and Ab was found in blood group A1 erythrocytes but not in A2 erythrocytes. Ax was identified by its conversion to "globo H" by alpha-N-acetylgalactosaminidase and by 1H-NMR spectroscopy as GalNAc alpha l----3[Fuc alpha l----2]Gal beta l----3GalNAc beta l----3Gal alpha l----4Gal beta l----4Glc beta l----lCer. Globo-H (Fuc alpha l----2Gal beta l----3GalNac beta l----3Gal alpha l----4Gal beta l----4Glc beta l----lCer) was found in blood group A, and O but not in A1 erythrocytes. Thus, one of the A1-specific determinants must be an A determinant carried by globo-series structure.

ABO Blood-Group System↗
PubMed1984

Blood group H antigen with globo-series structure. Isolation and characterization from human blood group O erythrocytes.

Blood group H antigen with globo-series structure, reacting with the monoclonal antibody MBrl, was isolated and characterized from human blood group O erythrocytes. The structure was identified by methylation analysis, direct probe mass spectrometry, and 1H-nuclear magnetic resonance spectroscopy as shown below: Fuc alpha 1----2Gal beta 1----3GalNAc beta 1----3Gal alpha 1----4Glc beta 1----1Cer.

ABO Blood-Group System↗
PubMed1984

Molecular diversity of calpastatin in mammalian organs.

The crude homogenates of various human and porcine organs were subjected to immunoelectrophoretic blot analysis using affinity-purified anti-calpastatin antibody which specifically reacts with human erythrocyte 70 kDa calpastatin. Multiple immuno-reactive bands were revealed which ranged from 100 to 50 kDa. The results indicated the diversity of monomeric calpastatin molecules. The band patterns were different from one organ to the other. Among them, lung, heart and skeletal muscle were characterized by the predominance of 90-100 kDa calpastatin, having a common antigenicity to erythrocyte 70 kDa calpastatin. Such molecular diversity of calpastatins was also substantiated by enzymatic and chromatographic analyses.

Animals↗
PubMed1984

Hybrid type glycolipids (lacto-ganglio series) with a novel branched structure. Their presence in undifferentiated murine leukemia cells and their dependence on differentiation.

The structures of these glycolipids are hybrids of the lacto and ganglio series, which are characterized by the presence of GlcNAc beta 1----3 and GalNAc beta 1----4 linked to the Gal residue of Gal beta 1----4Glc beta 1----1Cer. This new hybrid series can be designated as "lacto-ganglio series." These glycolipids are present in undifferentiated murine leukemia cells. Their concentration declines with differentiation and they are virtually absent in differentiated M1+ cells, suggesting that lacto-ganglio structures could be markers of undifferentiated, malignant myeloid cells.

Animals↗
PubMed1984

Evidence for membrane-associated calpain I in human erythrocytes. Detection by an immunoelectrophoretic blotting method using monospecific antibody.

Low and high Ca2+-requiring forms of Ca2+-dependent cysteine proteinase are known as calpain I and calpain II, respectively. We have obtained, for the first time, monospecific antibodies for calpain I and for calpain II. Using these antibodies and an electrophoretic blotting method, we have found that a small, but reproducible, amount of calpain I was associated with human erythrocyte membranes while the bulk of the protease was contained in the cytosol. Most of membrane-associated calpain I was extractable with 1% Triton X-100, but not with 0.1% detergent. In the presence of 0.1 mM Ca2+ and 5 mM cysteine, membrane-associated calpain I degraded the membrane protein band 4.1 preferentially and band 3 protein only slowly. The Ca2+-induced autodigestion of the membrane preparation was inhibited by leupeptin but not by a cytosolic calpain inhibitor, calpastatin, added to the incubation medium. No calpain II was detected in either erythrocyte cytosol or membranes when anti-calpain II antibody was used under the same conditions as those for the detection of calpain I.

Antibodies, Monoclonal↗
PubMed1984

Novel fucolipids accumulating in human adenocarcinoma. I. Glycolipids with di- or trifucosylated type 2 chain.

Among a series of fucolipids accumulating in human colonic and liver adenocarcinoma, two neutral fucolipids have been isolated and their carbohydrate structures have been characterized by methylation analysis, mass spectrometry, enzymatic degradation, and proton NMR spectrometry as shown in Structures 1 and 2 below. These glycolipids are either absent or present in very small quantity in normal colonic mucosa and normal liver tissue. (formular; see text) Difucosyl neolactonorhexaosylceramide (Structure 1) gives a doublet or a triplet on high performance thin layer chromatography and can be separated into four to five fractions on high pressure high performance liquid chromatography. These components have the same carbohydrate structure, but have different fatty acid composition. Fractions are characterized by a predominance of either alpha-hydroxy C16 fatty acid or alpha-hydroxy C24 fatty acid. Trifucosyl neolactonoroctaosylceramide (structure 2) gives two discrete bands, which are designated Z3 and Z4, on high performance thin layer chromatography. Methylation analysis and nuclear magnetic resonance spectra show that the Z3 and Z4 have identical carbohydrate structures. All the cases of cancer tissue that accumulate di- or trifucosyl structure (1 or 2 above) also accumulated lactofucopentaose(III)ceramide. A possible enhancement in a specific synthetic pathway including type 2 chain elongation coupled with alpha 1----3 fucosylation in human cancer is discussed.

Adenocarcinoma↗
PubMed1984

Enzyme immunoassay of calpain I and calpastatin and its application to the analysis of human erythrocyte hemolysate.

A highly sensitive sandwich enzyme immunoassay for a Ca2+-dependent cysteine proteinase (calpain I) and its specific endogenous inhibitor protein (calpastatin) was developed. The calpain I and calpastatin used as immunogens were purified from human erythrocytes. Anti-calpastatin antisera having sufficiently high titer were obtained only when the immunogen was purified by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The assay method was principally based on the report by M. Imagawa et al. (1982, J. Appl. Biochem. 4, 41-57), using a specific antibody-coated polystyrene ball and horseradish peroxidase-conjugated Fab' fragment of the antibody. The sensitivity was 0.1 ng of calpain I or calpastatin per assay tube. Starting with 50 microliter of the hemolysate from human erythrocytes, the method permitted direct and simultaneous determination of calpain I and calpastatin, without prior separation of these two enzymatically counteracting components by chromatography. The present method as applied to the erythrocytes from 14 healthy adults gave 120-170 micrograms for calpain I and 164-211 micrograms for calpastatin per gram of hemoglobin, respectively.

Calcium-Binding Proteins↗
PubMed1984

Glycolipid antigens with blood group I and i specificities from human adult and umbilical cord erythrocytes.

Neutral glycolipids and gangliosides of umbilical cord and adult human erythrocytes were separated by high performance liquid chromatography, and each fraction was analyzed by direct binding of anti-I (Ma) and anti-i (Den) on solid phase glycolipid-lecithin-cholesterol. The I- and i-active glycolipids were isolated and their structures were determined by methylation analysis and direct probe mass spectrometry. The major I antigen in adult erythrocytes, showing a remarkable binding activity with anti-I(Ma), was found in one neutral glycolipid fraction, designated fraction y4, which was identified as a mixture of two glycolipids of a new type, lactoisooctaosylceramide and monofucosyllactoisooctaosylceramide (for structures, see Table I). In addition, two gangliosides displaying direct binding activity with anti-I(Ma) were identified as monosialoganglioside G8, as previously described and disialosyllactoisooctaosylceramide, which showed the same level of I activity as the y4 glycolipid. The same ganglioside was recently isolated and characterized by Kundu and co-workers. The major i-active glycolipid antigen in umbilical cord erythrocytes, showing a strong binding activity with anti-i(Den), was a neutral glycolipid, x4a, which was identified as lactonorhexaosylceramide. This glycolipid without fucosyl or sialosyl substitution has not been isolated previously and was present as an obvious normal component of umbilical cord erythrocytes, but an extremely minor component of adult erythrocytes. Sialosyllactonorhexaosylceramide (G6) was isolated and characterized as a second i antigen of umbilical cord erythrocytes, but showed a very weak binding activity with the anti-i antibody. Although these sialosyl derivatives displayed only weak activity, the chemical quantity of the sialosyl derivatives is significantly large in fetal erythrocytes; therefore, Ii activity of human erythrocytes, in general, must be significantly dependent on sialosyl derivatives in addition to unsubstituted structures.

Adult↗
PubMed1983

Characterization of an epitope (determinant) structure in a developmentally regulated glycolipid antigen defined by a cold agglutinin Fl, recognition of alpha-sialosyl and alpha-L-fucosyl groups in a branched structure.

The antibody Fl shows preferential reactivity with adult erythrocytes over newborn erythrocytes, and its reactivity is abolished by sialidase treatment of the erythrocyte. The antibody was found to recognize binary determinants linked to the branched lacto-N-isooctaosylceramide (formula; see text) The presence of an N-acetylneuraminyl group at one end and L-fucosyl group at the other end is essential for the reactivity of the antibody. A substitution at the penultimate D-galactosyl residue of one of the chains with an alpha-D-(1 leads to 3)-linked 2-acetamido-2-deoxygalactosyl or galactosyl group did not inhibit the reactivity of the antibody. The new blood group A- and B-active, branched gangliosides are also isolated and characterized.

ABO Blood-Group System↗
PubMed1983

New globoseries glycosphingolipids in human teratocarcinoma reactive with the monoclonal antibody directed to a developmentally regulated antigen, stage-specific embryonic antigen 3.

Glycolipids in a cultured human teratocarcinoma cell line (2102Ep) were investigated. The major glycolipids in these cells are globoseries glycolipids having the following structures: (formula; see text) Synthesis of these structures by serial addition of galactose, fucose, and N-acetylneuraminic acid to globoside (Gb4) in this teratocarcinoma is obvious, although further elongation of Gb4 in human cells and tissues has not been previously found with the exception of the presence of a small quantity of Forssman glycolipid in some tissues in the human population (Fs+ group) and in some human cancers. The latter four glycolipids (b-e), with the common internal structure R leads to 3GalNAc beta 1 leads to 3Gal alpha 1 leads to 4R', were all reactive to a monoclonal antibody directed to the 4- to 8-cell stage of murine embryos, known as the stage-specific embryonic antigen 3 (SSEA-3 (Shevinsky, L. H., Knowles, B. B., Damjanov, I., and Solter, D. (1982) Cell 30, 697-705]; structure (c) showed the strongest reactivity. These findings, together with the demonstration of the glycolipid nature of SSEA-1 antigen (Kannagi, R., Nudelman, E., Levery, S. B., and Hakomori, S. (1982) J. Biol. Chem. 257, 14865-14874), indicate that cell surface glycolipids play significant roles as differentiation antigens during the course of embryogenesis.

Antibodies, Monoclonal↗
PubMed1983

A glycolipid antigen associated with Burkitt lymphoma defined by a monoclonal antibody.

The antigen defined by a rat monoclonal antibody directed to a Burkitt lymphoma cell line was identified as globotriaosylceramide [Gal alpha (1 leads to 4)-Gal beta (1 leads to 4)-Glc beta (1 leads to 1)-ceramide]. The antibody demonstrated a strict steric specificity since it did not react with globoisotriaosylceramide [Gal alpha (1 leads to 3)-Gal beta (1 leads to 4)-Glc beta (1 leads to 1)-ceramide], the positional isomer of the antigen associated with the Burkitt lymphoma. Chemical analysis of various Burkitt lymphoma cell lines revealed that the Burkitt lymphoma cells contained more than 100 times as much of the glycolipid antigen as was found in other human lymphoma and leukemia cell lines.

Animals↗
PubMed1983

Stage-specific embryonic antigens (SSEA-3 and -4) are epitopes of a unique globo-series ganglioside isolated from human teratocarcinoma cells.

Two monoclonal antibodies (MC631 and MC813-70) raised against 4- to 8-cell stage mouse embryos and a human teratocarcinoma cell line, respectively, detect the stage-specific embryonic antigens, the previously defined SSEA-3 and SSEA-4, described herein. These antibodies were both reactive with a unique globo-series ganglioside with the structure shown below: (formula; see text) The antibodies were found to recognize sequential regions of this ganglioside, i.e., MC813-70 recognizes the terminal 'a' structure whereas antibody MC631 recognizes the internal 'b' structure. Thus, a set of two antibodies defines this unique embryonic antigen. During differentiation of human teratocarcinoma 2102Ep cells, the globo-series glycolipids defined by these antibodies decrease and the lacto-series glycolipids, reacting with the SSEA-1 antibody appear. This antigenic conversion suggests that a shift of glycolipid synthesis from globo-series to lacto-series glycolipids occurs during differentiation of human teratocarcinoma and perhaps of pre-implantation mouse embryos.

Animals↗
PubMed1983

Sequential change of carbohydrate antigen associated with differentiation of murine leukemia cells: i-I antigenic conversion and shifting of glycolipid synthesis.

Cell surface carbohydrate antigens and their metabolism were investigated during the course of differentiation of murine cultured leukemia cells (M1) into macrophage-like cells. The major glycolipids in undifferentiated M1 cells were of the ganglio series, with a small amount of lacto-series glycolipids. A novel branched structure was found as a tetraosylceramide of M1- cells. Upon differentiation, synthesis of lacto-series glycolipids was significantly enhanced and synthesis of globo-series glycolipids was newly induced but the ganglio-series synthesis was much reduced. Undifferentiated cells expressed only i antigen (i+I-Pk-); differentiated macrophage-like cells became I-antigen dominant and Pk-antigen positive (i+/-I+Pk+). The changes proceeded in two sequential steps: (i) an enhancement of lacto-series glycolipid synthesis associated with the conversion of i antigen to I antigen, and (ii) subsequent induction of globo-series glycolipid synthesis accompanied by the appearance of Pk antigen. The experimental system offers a clue for studies on the process of branching (i-to-I conversion) as well as the biological significance of three major glycolipids (globo-, lacto-, and ganglio-series) as markers of cell differentiation.

Animals↗
PubMed1983

Differentiation-associated changes of glycolipid composition and metabolism in mouse myeloid leukemia cells. Induction of globotriaosylceramide and a galactosyltransferase.

This study was to find out whether induction of special glycolipids or glycosyltransferases for glycolipid synthesis which might be involved in the cell functions occurred during the differentiation. Mouse myeloid leukemia cell line (M1-), the differentiated cells (M1+), and a subcloned cell line (Mm1) were used for this purpose. Gangliotriaosylceramide (GA2) was the major glycolipid component in M1- cells. As a result of differentiation of M1- into M1+ cells, globotriaosylceramide (CTH) was newly induced as the main glycolipid, while GA2 decreased to a minor component. GA2 was found to be the main glycolipid in Mm1 cells but no CTH was recognized. All precursor glycolipids and glycosyltransferases required to complete the biosynthetic pathway glucosylceramide (CMH) leads to lactosylceramide (CDH) leads to GA2 leads to gangliotetraosylceramide (GA1) leads to sialosylgangliotetraosylceramide (GM1b) were found in M1- and also in Mm1 cells. A galactosyltransferase activity for CTH synthesis from CDH increased 10 fold during the differentiation. The induction of CTH in M1+ cells could be attributed to the increase of the galactosyltransferase activity. Both CTH as a surface marker and the galactosyltransferase as an enzyme marker are proposed as valuable markers of differentiation in M1- cells. Besides the galactosyltransferase, N-acetylglucosaminyltransferease involved in the formation of lactotriaosylceramide (amino-CTH) increased up to 3 fold during the differentiation. The increase of the enzyme activity seemed to be responsible for the biosynthesis of lactoneotetraosylceramide (paragloboside) which appeared in M1+ cells.

Animals↗
PubMed1983

Human erythroleukemia cell line (HEL) undergoes a drastic macrophage-like shift with TPA.

We investigated the effect of 12-O-tetradecanoyl-phorbol-13-acetate (TPA) on the human erythroleukemia cell line, HEL, and found that TPA addition (10(-6)-10(-8) M) to HEL cell cultures induces morphological, functional, and biochemical changes in HEL cells that are characteristic for macrophage-like cells. Apart from the drastic changes in morphology, the cells greatly enhance their phagocytic ability and acquire receptors for binding and degradation of chemically modified lipoproteins. At the biochemical level, a newly synthesized 85K glycoprotein is observed, and the cells are unresponsive to inducers of globin synthesis. Comparative observations with K562 cells indicate that TPA inhibits, as in HEL cells, spontaneous and induced globin synthesis, but induces minimal macrophage-like properties in these cells. The results with HEL cells are interpreted to indicate that TPA uncovers a latent monocyte-like phenotype in these cells.

Acid Phosphatase↗
PubMed1983

Carbohydrate antigen profiles of human erythroleukemia cell lines HEL and K562.

The expression of major carbohydrate antigens carried by polylactosaminyl chains in human erythroleukemia cell lines, K562 and HEL, was investigated by applying monoclonal antibodies recognizing specific carbohydrate determinants. The two cell lines showed common differences in their glycolipid compositions: (1) the presence of significant amounts of ganglio-series glycolipids, which are absent in normal erythrocytes; and (2) a remarkable reduction in the amount of globo-series glycolipids, which are the major glycolipids in normal human erythrocytes. A variety of differences were also detected in the carbohydrate antigens carried by lacto-series glycolipids and glycoproteins having related carbohydrate chains. K562 cells were i+H-X+, with a minor population of I+ cells. HEL cells were I-i+H+X-. The presence of the I+ population in K562 cells is particularly noteworthy, since I-antigen is characteristic of adult mature erythrocytes and is absent in most human leukemic cell lines. Several clones showing I+, I+/-i+/-, or I-i+ specificities were isolated from K562 cells by cloning in either methylcellulose media or limiting dilution, and I+ and I- cells were sorted by FACS fluorometer. HEL cells and these K562 clones provide a useful experimental model for studying the biologic significance and enzymatic control in expression of cell surface polylactosamines.

ABO Blood-Group System↗