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PubMed1993

Glycosphingolipids of human peripheral nervous system myelins isolated from cauda equina.

Compositions of neutral and sulfated glucuronyl glycosphingolipids purified from human motor and sensory nerves and myelins were studied. Higher neutral glycosphingolipids (fraction B), which were separated from GalCer (fraction A), were analyzed by TLC and TLC-immunostaining. Both nerve myelins contained paragloboside (nLc4Cer) and nLc6Cer dominantly as major higher glycosphingolipids and very little globoside (Gb4Cer), whereas both nerves contained Gb4Cer and nLc4Cer. Besides these major glycosphingolipids, a neutral glycolipid containing asialoGM1 (Gg4Cer) epitope and other minor components such as ceramide trihexoside and ceramide dihexoside were detected in both nerves and their myelins. Furthermore, sulfated glucuronyl nLc4Cer and nLc6Cer, which were monoclonal antibody HNK-1 reactive glycolipids, were detected in both nerves and myelins.

Cauda Equina↗
PubMed1981

Glycolipid receptors for uropathogenic Escherichia coli on human erythrocytes and uroepithelial cells.

A specific family of glycolipids, the globoseries, was shown to act as receptors on human uroepithelial cells and erythrocytes for the majority of uropathogenic Escherichia coli strains attaching to or hemagglutinating those cells. This was demonstrated in three different ways: (i) correlation between the natural presence of glycolipid in the target cell (erythrocytes of different species) and binding of bacteria; (ii) inhibition of attachment to human uroepithelial cells by preincubation of bacteria and glycolipid; and (iii) induction of binding to unreactive cells by coating of these cells with glycolipid. Strains reacting with the receptor agglutinated guinea pig erythrocytes in a mannose-resistant way after, but not before, coating of the cells with globotetraosylceramide. Unrelated glycolipids were not recognized. The reaction was made independent of simultaneous occurrence of mannose-sensitive adhesions on the strains by addition of D-mannose. The receptor-coated cells were used as a tool to screen for prevalence of receptor recognition in a collection of 453 E. coli strains isolated from patients with urinary tract infection or from the stools of healthy children. Of 150 strains attaching to human uroepithelial cells and agglutinating human erythrocytes, 121 bound to globotetraosylceramide (81%). Globoside recognition was especially frequent among pyelonephritis strains (74/81). The glycolipid composition of the urogenital epithelium and kidney tissue and the ability of uropathogenic E. coli to bind to these glycolipids may be a determinant in host-parasite interaction leading to urinary tract infection.

Animals↗
PubMed1986

The effects of glycolipids and carbohydrates on bilirubin cytotoxicity in vitro.

We have studied the inhibitory effects of various glycolipids including gangliosides and carbohydrates on bilirubin cytotoxicity towards human glia cells using a 51Cr release assay. All cytotoxic activity was abolished when 100 microM bilirubin was incubated with 40-80 microM gangliosides (GM1, galactosyl-N-acetylgalactosaminyl-(N-acetylneuraminyl) galactosylglucosylceramide; GM2, N-acetylgalactosaminyl-(N-acetylneuraminyl)galactosylglucosylceramide; GM3, N-acetylneuraminylgalactosylglucosylceramide, and GD1a, N-acetylneuraminylgalactosyl-N-acetylgalactosaminyl-(N-acetylneura minyl) galactosylglucosylceramide). On the other hand, inhibition by asialoglycolipids was less effective than that by gangliosides, but became more effective with increasing numbers of sugars per molecule, the efficacy of inhibition being as follows: globoside greater than trihexosyl ceramide greater than lactosyl ceramide greater than glucosyl ceramide. Glucosyl ceramide and the carbohydrates of neuramine lactose or N-acetylneuraminic acid (NANA) did not have inhibitory effects. These results suggest that the terminal acidic group of NANA and the numbers of sugar chains in glycolipids play important roles in the inhibition of bilirubin cytotoxicity.

Bilirubin↗
PubMed1985

Molecular basis of Escherichia coli colonization of the upper urinary tract in BALB/c mice. Gal-Gal pili immunization prevents Escherichia coli pyelonephritis in the BALB/c mouse model of human pyelonephritis.

Most human pyelonephritis Escherichia coli isolates express both mannose (MS)- and globoside (Gal-Gal)-binding pili. An ascending E. coli urinary tract infection model was established in the 16-wk-old female BALB/c mouse to compare the pathogenic significance of MS and Gal-Gal pili and their efficacy as vaccines for the prevention of pyelonephritis. The distribution and density of pilus receptor compounds in urogenital tissues and as soluble compounds in urine were determined with antibodies to the synthetic receptor analogues, alpha D-Gal(1----4) beta D-Gal and alpha D-Man(1----2) alpha D-Man. Both carbohydrates were detected in vagina, bladder, ureter, and renal pelvis epithelium and in collecting duct and tubular cells. A pilus receptor compound also was detected in urine. It competitively inhibited the binding capacity of MS pili and was found to be physically, chemically, and immunologically related to Tamm-Horsfall uromucoid. Infectivity and invasiveness were quantitatively and histologically characterized for four E. coli strains: J96, a human pyelonephritis strain that expresses both MS and Gal-Gal pili; two recombinant strains prepared from J96 chromosomal DNA encoding MS pili or Gal-Gal pili; and the nonpiliated K12 recipient. Intravesicular administration of J96 (10(6) colony-forming units [CFU]) resulted in renal colonization and invasion in each of nine mice. The Gal-Gal clone (10(6) CFU) colonized the kidneys in each of 10 mice but did not invade. In contrast, the MS clone (10(6) CFU) did not colonize renal epithelium or invade. This effect was superceded when larger doses (greater than or equal to 10(10) CFU) of the MS clone were administered in volumes that cause acute vesicoureteric reflux. The efficacy was determined of vaccines composed of pure MS or Gal-Gal pili or the lipopolysaccharide containing O somatic antigen of the challenge strain, J96. The Gal-Gal pilus vaccine blocked renal colonization in 19 of 22 mice and renal invasion in 10 of 11 mice. Gal-Gal pili may be useful immunogens for the prevention of pyelonephritis in anatomically normal urinary tracts.

Animals↗
PubMed2000

Inverse relationship of expression between GM3 and globo-series ganglioside in human renal cell carcinoma.

Renal cell carcinoma (RCC) is highly metastatic. We previously showed that expression of globo-series ganglioside is associated with the metastatic potential of RCC. However, the mechanism of metastasis remains largely unknown, and there is no effective therapy for metastasis. It was recently shown that induction of differentiation of colon cancer cells by brefeldin A was accompanied by an increase of GM3 with a concomitant decrease of neolacto-series gangliosides. To get a clue to a new method of therapy for RCC, we investigated whether the similar changes occur in RCC cells expressing globo-series ganglioside. Growth suppression and an increase of GM3 simultaneous with a decrease of monosialosyl galactosyl globoside, a member of globo-series gangliosides, were observed in human RCC cell line ACHN following brefeldin A treatment. The resultant change of the ganglioside profile is inversely related to the ganglioside pattern associated with the malignant potential of RCC and almost coincided with that representative of RCC cases showing favorable prognoses. It is suggested that the inverse relationship of expression between GM3 and globo-series ganglioside is reflected on the degree of malignancy of RCC, and may be useful as one of the indicators for exploiting treatment methods of RCC.

Brefeldin A↗
PubMed1979

[Study on glycolipids in human lung carcinoma of histologically different types (author's transl)].

Human lung carcinoma tissues with histological types of adenocarcinoma, squamous cell and small cell undifferentiated carcinomas were investigated for glycolipids. Carcinoma tissues, as well as normal adult and embryonic lungs contained ceramide mono-, di- and trihexosides, globoside and hematoside as major glycolipids. In addition to them, sulfatide which was identified as ceramide 3-sulfate-galactoside, was isolated in much lesser amount. The content of sulfatide was markedly increased in adenocarcinoma than that in other carcinomas and normal lung. Adenocarcinoma was also characterized by significantly lower level of total glycolipids which was largely due to the diminished contents of ceramide mono- and dihexosides, and hematoside, as compared to those in other two carcinomas. Squamous cell carcinoma had a characteristic pattern with an increment of hematoside. In small cell undifferentiated carcinoma, glycolipid contents were similar with those in squamous cell carcinoma but the relative composition of major glycolipids was markedly differed from that in other types. All the types of carcinoma examined showed marked increase of ceramide mono- and dihexosides (except for ceramide dihexoside in adenocarcinoma) compared to those in normal adult lung. Overall feature of glycolipids in embryonic lung appeared to be an intermediate between carcinomas and normal adult lung.

Adenocarcinoma↗
PubMed1986

Monoclonal antibodies detecting different epitopes on the Forssman glycolipid hapten.

Two hybridomas, derived by fusing mouse myeloma cells with spleen cells from a rat immunized with mouse mammary tumors, have been shown to produce antibodies that recognize cell surface antigens on mesenchymal cells in a variety of tissues. Evidence presented in this report suggests that these antibodies detect overlapping epitopes on the Forssman glycolipid hapten (GalNAc alpha 1-3GalNAc beta 1-3Gal alpha 1-4Gal beta 1-4Glc beta 1-1Cer). One antibody (33B12) reacts with the terminal sugar sequence GalNAc alpha 1-3GalNAc and is specific for Forssman. The other antibody (117C9) recognizes the internal sugar sequence GalNAc beta 1-3Gal. The terminal sugar sequence GalNAc beta 1-3Gal in globoside, as well as the internal sugar sequence GalNAc beta 1-4Gal in asialo-GM1, is not recognized as an antigenic determinant by 117C9. Nevertheless, the 117C9 antibody does not react exclusively with the Forssman antigen. In a lipid extract fractionated by Folch partition of mouse mammary tumors, the antibody also detects other glycolipids.

Animals↗
PubMed1987

Anti-human tumor antibodies induced in mice and rabbits by "internal image" anti-idiotypic monoclonal immunoglobulins.

MBr1 is a murine monoclonal antibody, defining a saccharidic epitope [CaMBr1] of a human tissue-specific, tumor-associated globoside, present on the mammary carcinoma cell line MCF-7. The same epitope is shared by glycoproteins present on normal and neoplastic mammary epithelial cells, and by mucins from some ovarian cyst fluids. We have used MBr1 as the monoclonal antitumor antibody in an idiotypic sequence of immunizations in order to obtain and characterize "internal images" of the original epitope to be used as substitutes of the nominal antigen in serologic immunoassays. Two monoclonal anti-idiotypic antibodies (beta-1 and beta-2), which reacted with paratope-related idiotopes on MBr1, were obtained. The analysis of the antigenic and immunogenic properties of these molecules by both "antigen" and "antibody" competition assays provided evidence that both beta-1 and beta-2 bear "internal images" of the MBr1-defined epitope. Moreover, when injected in mice and rabbits both beta-1 and beta-2 induced anti anti-idiotypic antibodies, which mimicked MBr1 in binding MCF-7 as well as normal and neoplastic mammary gland epithelial cells. These data are discussed in terms of their possible application to the production of tumor-associated antigen substitutes and their use in serologic immunoassays.

Animals↗
PubMed1989

Globotetraosylceramide is recognized by the pig edema disease toxin.

The pig edema disease toxin has been shown by a tlc glycolipid binding assay to bind specifically to globotetraosylceramide (Gb4, GalNAc beta 1-3Gal alpha 1-4Gal beta 1-4GlcCer.). Binding was reduced for globotriosylceramide (Gb3, Gal alpha 1-4Gal beta 1-4GlcCer) and more markedly for the Forssman antigen (GalNAc alpha 1-3GalNAc beta 1-3Gal alpha 1-4Gal beta 1-4GlcCer). Paragloboside, blood group A glycolipids, glycolipids terminating in Gal NAc beta 1-4Gal-, and glycolipids in which globoside was present as an internal sequence did not bind the toxin. Isogloboside (GalNAc beta 1-3Gal alpha 1-3Gal beta 1-4GlcCer) was efficiently recognized. This toxin is genetically related to the verotoxin (or Shiga-like) family of toxins for which Gb3 has been shown to be the receptor. The difference in susceptibility of cell lines to the cytotoxicity of the pig edema disease toxin and the Shiga and Shiga-like toxins is consistent with the difference in receptor glycolipid binding.

Animals↗
PubMed1987

Forssman glycolipid, an antigenic marker for a major subpopulation of macrophages from murine spleen and peripheral lymph nodes.

Three hybridoma clones were isolated after hybridization of a mouse myeloma line with splenocytes from rats immunized with Forssman glycosphingolipid (Fo). Two of these clones produced Fo-specific monoclonal antibodies (MAB) of the IgM class, one MAB of the IgG2c class. In complement-dependent depletion experiments and immunofluorescence studies on the nature of Fo-positive leukocytes in CBA/J mice the following results were obtained: whereas blood monocytes, polymorphonuclear leukocytes, and lymphocytes were Fo negative, 5 to 10% of suspended spleen cells were positive. The majority of these were macrophage-like, glass- and nylon-adherent, nonspecific esterase-positive phagocytizing cells carrying Ia and globoside markers. These cells participated as accessory cells in the mixed lymphocyte culture reaction. In cell suspensions from axillary and inguinal lymph nodes, 2% were Fo positive. They were enriched up to 70% in the glass-adherent, esterase-positive population from this source. In contrast, no Fo-positive cells were detected in mesenteric lymph nodes, and less than 0.1% of the resident peritoneal macrophages bore this marker. The percentage of Fo-positive cells increased to 1% in thioglycollate-elicited peritoneal cells. Immunostaining of cryosections of lung and liver tissue showed alveolar macrophages and Kupffer cells, respectively, to be Fo negative.

Animals↗
PubMed1985

Identification of the neutral glycosphingolipids of murine mast cells: expression of Forssman glycolipid by the serosal but not the bone marrow-derived subclass.

The expression of neutral glycosphingolipids by mouse T cell-dependent, bone marrow-derived mast cells (BMMC) obtained in vitro was determined by chromatographic and immunochemical criteria. Neutral glycosphingolipids were isolated from BMMC by extraction of 3 to 5 X 10(8) cells in chloroform/methanol (1/1, v/v) and chromatography on DEAE-Sephadex, and were analyzed by thin layer chromatography with orcinol staining. The predominant neutral glycosphingolipids of BMMC were glucosylceramide (CMH), lactosylceramide (CDH), globotriosylceramide (CTH), globotetraosylceramide (globoside), and a molecule migrating slightly faster than gangliotetraosylceramide (asialo GM1) and slower than globopentaosylceramide (Forssman glycolipid). The profiles on thin layer chromatograms of the neutral glycosphingolipids were the same for BMMC derived from BALB/c, C57BL/6, WBBF1-W/Wv, and WBBF1-+/+ mice, and for cells differentiated in either WEHI-3 conditioned medium or concanavalin A-splenocyte conditioned medium. High performance liquid chromatography of benzoylated neutral glycosphingolipids of BMMC on a Zipax column confirmed the identity of the four neutral glycosphingolipids identified by thin layer chromatography. The fifth major glycosphingolipid had an elution time greater than that of globotetraosylceramide and did not co-elute with any of the standards tested. Direct biochemical analyses of the neutral glycosphingolipids of mouse serosal mast cells (SMC) were not feasible because only 2 X 10(6) SMC could be isolated per 100 mice. However, mouse SMC bound a rat monoclonal anti-globopentaosylceramide antibody (M1/87.27.7) and rat monoclonal B1.1 antibody, as assessed by indirect immunofluorescence and flow cytometry, whereas mouse BMMC did not. The binding of B1.1 antibody to SMC could be blocked by the anti-globopentaosylceramide antibody, and the specificity of B1.1 antibody for globopentaosylceramide was confirmed immunochemically with the use of a solid phase radioimmunoassay. As estimated immunochemically, the amount of globopentaosylceramide in mouse SMC was 62 ng/10(6) cells, whereas BMMC contained less than 8 ng/10(6) cells. Thus, the expression of globopentaosylceramide is a characteristic of the mouse SMC that is lacking in the T cell-dependent BMMC.

Animals↗
PubMed1980

Isolation, characterization, and biosynthesis of Forssman antigen in human lung and lung carcinoma.

A glycolipid was isolated from normal lung and lung carcinoma tissues. This glycolipid was identified as the Forssman antigen of pentaglycosyl ceramide by means of chemical and immunological methods. The presence of this antigenic glycolipid was observed in all the tissues examined of adult and embryo lungs, and of lung tumors irrespective of histological type. The extracts of human lung and lung tumors were capable of catalyzing the synthesis of Forssman antigen from globoside.

Adenocarcinoma↗
PubMed1981

Expression of Forssman antigen synthesis and degradation in human lung cancer.

The activities of the two enzymes (UDP-acetylgalactosamine-globoside alpha-N-acetylgalactosaminyltransferase and alpha-N-acetyl-D-galactosaminidase) involved in the synthesis and degradation of Forssman antigen were studied in uninvolved and neoplastic human lungs. The Forssman synthetic enzyme activities of 17 of 18 squamous cell carcinomas were higher than those of the uninvolved lung tissues of the subjects studied, and the degradation enzyme activities of 16 of 18 squamous cell carcinomas were higher than those of the uninvolved portions. No consistent abnormalities in both enzyme activities were seen in 28 adenocarcinomas, whereas the mean activities of the two enzymes were elevated in these neoplasms. These differences in enzyme activities between those samples may indicate that the synthesis and degradation of Forssman antigen in adenocarcinoma of the lung are expressed or repressed according to the individual, whereas in squamous cell carcinoma, these activities are expressed unrelated to the individual.

Adenocarcinoma↗
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↗
PubMed1980

Identification of ganglio-N-tetraosylceramide as a new cell surface marker for murine natural killer (NK) cells.

The BCG-induced NK cell activity of murine peritoneal exudate cells was abolished by preincubation of effector cells with anti-ganglio-N-tetraosylceramide (anti-asialo GM1) and C but not with other anti-glycolipid antibodies, anti-ganglioside GM1, anti-globoside, and anti-ganglio-N-triosylceramide (anti-asialo GM2). In contrast, the cytotoxic activity of alloimmune T cells was not affected by treatment with anti-asialo GM1 antisera. These findings suggest that asialo GM1 display may be characteristic of NK cell populations and aid in the isolation of this population of cytotoxic cells.

Animals↗
PubMed1980

Gangliosides and neutral glycolipids of human adrenal medulla.

Glycolipids were isolated from human adrenal medulla by DEAE-Sephadex A-25 and Iatrobeads column chromatography. The lipid-bound sialic acid was about 234 microgram/g fresh tissue. The glanglioside fraction contained two major gangliosides which accounted for 93% of the total lipid-bound sialic acid. They were identified as GM3, N-acetylneuraminylgalactosylglucosylceramide and GD3, N-acetylneuraminyl N-acetylneuraminylgalactosylglucosylceramide on the basis of cochromatography with authentic standards, sugar composition analysis, and neuraminidase digestion. GM3, N-acetylneuraminylgalactosylglucosylceramide and GD3, N-acetylneuraminyl N-acetylneuraminylgalactosylglucosylceramide occurred in a ratio of approximately 3 : 2, and the ratio seemed to be rather constant irrelevant of age and sex differences. The neutral glycolipid fraction consisted of GL1a, glucosylceramide (18%), GL1b, galactosylceramide (23%), GL2a, lactosylceramide (27%), GL3, digalactosylglucosylceramide (20%), and GL4, globoside (12%). The major fatty acids of all these glycolipids were 16 : 0, 18 : 0, 22 : 0, 24 : 0 and 24 :1.

Adrenal Medulla↗
PubMed1980

Sandhoff's disease (GM2 gangliosidosis type 2). Histopathology and ultrastructure of the eye.

Sandhoff's disease (GM2 gangliosidosis type 2) was diagnosed in an infant in whom a progressive neurologic disorder and cherry-red foveal spots developed. At autopsy, ultrastructural examination of the retina and optic nerve disclosed abundant pleomorphic storage cytosomes in all neurons of the retina, including the inner segments of the photoreceptor cells, and in glial cells of the optic nerve. Electron microscopy of the cornea showed, within the keratocytes, distended clear lysosomes that contained some fibrillogranular material and an occasional collection of lamellae. We discuss the pathogenesis of the clinical and pathologic ocular findings with regard to the inherited absence of the enzymes hexosaminidase A and B and an accumulation of the substrates, GM2 ganglioside and asialo GM2, in the nervous system (including retina and optic nerve) and globoside and other hexosamine-containing substances in the viscera (including cornea).

Adolescent↗
PubMed2004

Impaired cell adhesion and apoptosis in a novel CLN9 Batten disease variant.

We describe the ninth variant of neuronal ceroid lipofuscinosis (NCL) or Batten disease, due to defects in a putative new gene, CLN9. We therefore refer to the new variant as CLN9-deficient. Two Serbian sisters and two German brothers are described. Their clinical history is characteristic for juvenile NCL. They show similar gene expression patterns. The existence of this variant is supported by the presence of curvilinear inclusions, fingerprint profiles, and granular osmiophilic deposits in neurons, lymphocytes, and conjunctival cells. Enzyme screening and sequencing of the coding regions of other NCL genes was negative. CLN9-deficient cells have a distinctive phenotype. They have rounded cell bodies, have prominent nucleoli, attach poorly to the culture dish, and are sensitive to apoptosis but have increased growth rates. Gene expression of proteins involved in cell adhesion and apoptosis is altered in these cells. Sphingolipid metabolism is also perturbed. They have decreased levels of ceramide, sphingomyelin, lactosylceramide, ceramide trihexoside, and globoside and increased activity of serine palmitoyl transferase.

Adolescent↗