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Akemi Suzuki

Publications and source records attributed to Akemi Suzuki.

36 records · Page 2Linked to original sources
PubMed2004

Clinical, biochemical, and cytochemical studies on a Japanese Salla disease case associated with a renal disorder.

We report the first Japanese case of Salla disease. A 5-year-old male patient developed unique proteinuria with other clinical manifestations, including coarse facies, dysostosis multiplex, mild mitral valve regurgitation, umbilical and inguinal herniation, and mild developmental delay. Pathological analysis of biopsied kidney tissues showed marked vacuolation of podocytes, mesangial cells, capillary endothelial cells, and tubular cells. Biochemical studies involving thin-layer chromatography and mass spectrometry revealed increased excretion of free sialic acid (N-acetylneuraminic acid) into the patient's urine. Immuno- and lectin staining of the patient's cells demonstrated the accumulation of sialyl and asialyl glycoconjugates in lysosomes and late endosomes. A defect in sialyl glycoconjugate metabolism is thought to have occurred in the patient's cells, besides impairment of the lysosomal transport of free sialic acid residues. A renal disorder should be considered as an important manifestation, not only in infantile free sialic acid storage disease but also in Salla disease.

Biological Transport↗
PubMed2004

Identification of an essential sequence for dihydroceramide C-4 hydroxylase activity of mouse DES2.

Although the amino acid sequences of mouse DES1 (MDES1) and DES2 (MDES2) have 63% sequence identity, their enzymatic characteristics are quite different. MDES1 exhibits high dihydroceramide delta4-desaturase activity and very low C-4 hydroxylase activity, while MDES2 is similarly active as both a dihydroceramide delta4-desaturase and a C-4 hydroxylase. We constructed several chimeras of MDES1 and MDES2 and identified a region important for C-4 hydroxylase activity in MDES2. This region contains the sequence XAFGY (X=T or A or V; Y=T or N) and occurs on the C-terminal side of the first His-box of MDES2. We confirmed the conservation of this region in DES2 family members sequenced from humans, pigs, rats, chickens, zebrafish, and Xenopus.

Amino Acid Sequence↗
PubMed2004

SLI1 (YGR212W) is a major gene conferring resistance to the sphingolipid biosynthesis inhibitor ISP-1, and encodes an ISP-1 N-acetyltransferase in yeast.

ISP-1 (myriocin) is a potent inhibitor of serine palmitoyltransferase, the primary enzyme of sphingolipid biosynthesis, and is a useful tool for studying the biological functions of sphingolipids in both mammals and yeast (Saccharomyces cerevisiae). In a previous study, we cloned yeast multicopy suppressor genes for ISP-1, and one of these, YPK1/SLI2, was shown to encode a serine/threonine kinase which is a yeast homologue of mammalian SGK1 (serum/glucocorticoid-regulated kinase 1). In the present study, another gene, termed SLI1 (YGR212W; GenBank accession number CAA97239.1), was characterized. Sli1p has weak similarity to Atf1p and Atf2p, which are alcohol acetyltransferases. Although a sli1-null strain grew normally, the IC50 of ISP-1 for the growth of this strain was markedly decreased compared with that for the parental strain, indicating that Sli1p is a major contributor to ISP-1 resistance in yeast. On a sli1-null background, the increase in resistance to ISP-1 induced by YPK1 gene transfection was almost abolished. These data indicate that Sli1p co-operates with Ypk1p in mediating resistance to ISP-1 in yeast. Sli1p was found to convert ISP-1 into N-acetyl-ISP-1 in vitro. Furthermore, N-acetyl-ISP-1 did not share the ability of ISP-1 to inhibit the growth of yeast cells, and the serine palmitoyltransferase inhibitory activity of N-acetyl-ISP-1 was much lower than that of ISP-1. These data suggest that Sli1p inactivates ISP-1 due to its N-acetyltransferase activity towards ISP-1.

Acetylation↗
PubMed2004

Loss of disialyl Lewis(a), the ligand for lymphocyte inhibitory receptor sialic acid-binding immunoglobulin-like lectin-7 (Siglec-7) associated with increased sialyl Lewis(a) expression on human colon cancers.

Expression of sialyl Lewis(a) is known to be increased in cancers of the digestive organs. The determinant serves as a ligand for E-selectin and mediates hematogenous metastasis of cancers. In contrast, disialyl Lewis(a), which has an extra sialic acid attached at the C6-position of penultimate GlcNAc in sialyl Lewis(a), is expressed preferentially on nonmalignant colonic epithelial cells, and its expression decreases significantly on malignant transformation. Introduction of the gene for an alpha2-->6 sialyl-transferase responsible for disialyl Lewis(a) synthesis to colon cancer cells resulted in a marked increase in disialyl Lewis(a) expression and corresponding decrease in sialyl Lewis(a) expression. This was accompanied by the complete loss of E-selectin binding activity of the cells. In contrast, the transfected cells acquired significant binding activity to sialic acid-binding immunoglobulin-like lectin-7 (Siglec-7)/p75/adhesion inhibitory receptor molecule-1, an inhibitory receptor expressed on lymphoid cells. These results indicate that the transition of carbohydrate determinants from disialyl Lewis(a)-dominant status to sialyl Lewis(a)-dominant status on malignant transformation has a dual functional consequence: the loss of normal cell-cell recognition between mucosal epithelial cells and lymphoid cells on one hand and the gain of E-selectin binding activity on the other. The transcription of a gene encoding the alpha2-->6 sialyltransferase was markedly down-regulated in cancer cells compared with nonmalignant epithelial cells, which is in line with the decreased expression of disialyl Lewis(a) and increased expression of sialyl Lewis(a) in cancers. Treatment of cancer cells with butyrate or 5-azacytidine induced strongly disialyl Lewis(a) expression, suggesting that histone deacetylation and/or DNA methylation may be involved in the silencing of the gene in cancers.

Antigens, Differentiation, Myelomonocytic↗
PubMed2004

Hypoxia induces adhesion molecules on cancer cells: A missing link between Warburg effect and induction of selectin-ligand carbohydrates.

Cancer cells undergo distinct metabolic changes to cope with their hypoxic environment. These changes are achieved at least partly by the action of transcriptional factors called hypoxia-inducible factors (HIFs). We investigated gene expression in cultured human colon cancer cells induced by hypoxic conditions with special reference to cell-adhesion molecules and carbohydrate determinants having cell-adhesive activity by using DNA-microarray and RT-PCR techniques. Hypoxic culture of colon cancer cells induced a marked increase in expression of selectin ligands, the sialyl Lewis x and sialyl Lewis a determinants at the cell surface, which led to a definite increase in cancer cell adhesion to endothelial E-selectin. The transcription of genes for fucosyltransferase VII (FUT7), sialyltransferase ST3Gal-I (ST3O), and UDP-galactose transporter-1 (UGT1), which are all known to be involved in the synthesis of the carbohydrate ligands for E-selectin, was significantly induced in cancer cells by hypoxic culture. In addition, a remarkable induction was detected in the genes for syndecan-4 (SDC4) and alpha5-integrin (ITGA5), the cell-adhesion molecules involved in the enhanced adhesion of cancer cells to fibronectin. The transcriptional induction by hypoxia was reproduced in the luciferase-reporter assays for these genes, which were significantly suppressed by the co-transfection of a dominant-negative form of HIF. These results indicate that the metabolic shifts of cancer cells partly mediated by HIFs significantly enhance their adhesion to vascular endothelial cells, through both selectin- and integrin-mediated pathways, and suggest that this enhancement further facilitates hematogenous metastasis of cancers and tumor angiogenesis.

Antigens, Tumor-Associated, Carbohydrate↗
PubMed2004

DES2 protein is responsible for phytoceramide biosynthesis in the mouse small intestine.

The C-4 hydroxylation of sphinganine and dihydroceramide is a rate-limiting reaction in the biosynthesis of phytosphingolipids. Mouse DES1 (MDES1) cDNA homologous to the Drosophila melanogaster degenerative spermatocyte gene-1 (des-1) cDNA leads to sphingosine Delta4-desaturase activity, and another mouse homologue, MDES2, has bifunctional activity, producing C-4 hydroxysphinganine and Delta4-sphingenine in yeast [Ternes, Franke, Zahringer, Sperling and Heinz (2002) J. Biol. Chem. 277, 25512-25518]. Here, we report the characterization of mouse DES2 (MDES2) using an in vitro assay with a homogenate of COS-7 cells transfected with MDES2 cDNA and N -octanoyl-sphinganine and sphinganine as substrates. MDES2 protein prefers dihydroceramide as a substrate to sphinganine, and exhibits dihydroceramide Delta4-desaturase and C-4 hydroxylase activities. MDES2 mRNA content was high in the small intestine and abundant in the kidney. In situ hybridization detected signals of MDES2 mRNA in the crypt cells. Immunohistochemistry using an anti-MDES2 peptide antibody stained the crypt cells and the adjacent epithelial cells. These results suggest that MDES2 is the dihydroceramide C-4 hydroxylase responsible for the biosynthesis of enriched phytosphingoglycolipids in the microvillous membranes of intestinal epithelial cells.

Animals↗
PubMed2004

Core 2 GlcNAc transferase and kidney tubular cell-specific expression.

The expression of glycan chains is precisely regulated in a time- and space-dependent manner. We summarize here our recent work on the kidney tubular cell-specific regulation of core 2 beta-1,6-GlcNAc transferase. Gsl5 gene was first identified by genetic analysis on the basis of polymorphic expression of kidney glycolipids among inbred strains of mice and turned out to be a regulatory gene controlling the level of mRNA of kidney-specific core 2 beta-1,6-GlcNAc transferase. This kidney-specific core 2 GlcNAc transferase takes glycolipids having Gal beta 1-3GalNAc at their termini, Gal beta 1-3GalNAc alpha 1- and beta 1-oligosaccharide derivatives, and glycoproteins having core 1 structure, as substrates. Immunohistochemistry with anti-core 2-Le( x ) monoclonal antibody demonstrated that vesicles located just below the microvillous membrane of proximal tubule cells were clearly stained in a Gsl5 -wild type mouse. Western blotting with the monoclonal antibody detected a major glycoprotein with a molecular mass of 500 kDa in the microsomal fraction of the wild type mouse kidney. In situ hybridization with anti-sense cDNA of kidney-specific core 2 GlcNAc transferase confirmed that Gsl5 gene controls the expression of the core 2 beta-1,6-GlcNAc transferase mRNA in a proximal tubular cell-specific manner. The 5' upstream sequences of the kidney-specific core 2 GlcNAc transferase gene in inbred and wild-derived strains of mice were analyzed, and the phylogenetic analysis of these sequences suggests that functional Gsl5 gene might be produced by the time of subspeciation of M. musculus, about one million years ago.

Animals↗
PubMed2004

Are N-glycolylneuraminic acid (Hanganutziu-Deicher) antigens important in pig-to-human xenotransplantation?

BACKGROUND: N-glycolylneuraminic acid (NeuGc) epitopes, so called Hanganutziu-Deicher (HD) antigens, which are widely expressed on endothelial cells of all mammals except humans, are considered to be potential targets for natural and elicited anti-nonGalalpha1-3 Gal (Gal) antibodies in humans. We previously reported that anti-NeuGc antibodies were not detected in healthy humans by enzyme-linked immunosorbent assay (ELISA) using NeuGc-GM3-coated plates, and no antibody production was observed in patients with a history of exposure to pig cells. However, a recent paper has revealed that (i) anti-NeuGc antibodies to porcine red blood cells (PRBC) are detectable in most healthy humans, and (ii) the majority of anti-nonGal antibodies are specific for NeuGc epitopes. The purpose of this study was to reassess whether NeuGc is important as an immunogenic nonGal epitope. METHODS: The binding of antibodies to PRBC and porcine aortic endothelial cells (PAEC) was compared. Cells were treated with (i) alpha-galactosidase, and then (ii) neuraminidase, which digests sialic acids, including NeuGc epitopes. Cells were incubated with human pooled sera, and applied to flow cytometric analysis. After enzyme digestion, almost complete reduction of Gal and NeuGc expression was confirmed by GS-IB4 and HU/Ch2-7 (a chicken monoclonal antibody against HD antigens), respectively. Trypsin, which removes membrane glycoproteins, and endoglycoceramidase which cleaves glycolipids, were used for differentiating between NeuGc-containing glycoproteins and glycolipids. RESULTS: Neuraminidase-treatment reduced the binding of immunoglobulin G (IgG) antibodies to PRBC; about half of the anti-nonGal IgG antibodies to PRBC were directed to NeuGc. In contrast, anti-nonGal antibodies to PAEC were not directed to NeuGc. Trypsin-treatment markedly reduced the expression of NeuGc only on PRBC. Endoglycoceramidase-treatment was followed by a greater reduction in NeuGc epitopes on PAEC than on PRBC. Most NeuGc on PRBC appeared to be linked to proteins, but most NeuGc on PAEC was expressed on glycolipids. CONCLUSIONS: Carbohydrate structures on PRBC are different from those on PAEC. Healthy human sera contain anti-nonGal IgG antibodies to NeuGc expressed on PRBC, but not on PAEC. We speculate that anti-nonGal IgG antibodies to PRBC can recognize both NeuGc and protein, and this may be the reason why such antibodies have not been detected by ELISA. A definite conclusion about the immunogenicity of NeuGc has not been obtained. More sera from patients (not from non-human primates) sensitized with porcine cells or organs need to be studied.

Animals↗
PubMed2004

Identifying rodent hantavirus reservoirs, Brazil.

We describe the genetic analysis of samples from hantavirus pulmonary syndrome (HPS) patients from southern and southeastern states of Brazil and rodents captured at the presumed site of infection of these patients. A total of 65 samples that were antibody-positive for Sin Nombre or Laguna Negra virus by enzyme-linked immunosorbent assay were processed by nested reverse transcription-polymerase chain reaction (RT-PCR) by using several primer combinations in the M and S genome segments. PCR products were amplified and sequenced from samples from 11 HPS patient and 7 rodent samples. Phylogenetic analysis of nucleotide sequence differences showed the cocirculation of Araraquara and Juquitiba-like viruses, previously characterized from humans. Our genetic data indicate that Araraquara virus is associated with Bolomys lasiurus (hairy-tailed Bolo mouse) and the Juquitiba-like virus is associated with Oligoryzomys nigripes (black-footed pigmy rice rat).

Adult↗
PubMed2002

Human epidermal glucosylceramides are major precursors of stratum corneum ceramides.

Ceramides are the major component of the stratum corneum, accounting for 30%-40% of stratum corneum lipids by weight, and are composed of at least seven molecular groups (designated ceramides 1-7). Stratum corneum ceramides, together with cholesterol and fatty acids, form extracellular lamellae that are responsible for the epidermal permeability barrier. Previous studies indicated that beta-glucocerebrosidase- and sphingomyelinase-dependent ceramide production from glucosylceramides and sphingomyelins, respectively, is important for epidermal permeability barrier homeostasis. A recent study indicated that sphingomyelins are precursors of two stratum corneum ceramide molecular groups (ceramides 2 and 5). In this study, we have examined the role of glucosylceramides in the generation of each of the seven stratum corneum ceramide molecular groups. First, the structures of various glucosylceramide species in human epidermis were determined by gas chromatography-mass spectrometry, fast atom bombardment-mass spectrometry, and nuclear magnetic resonance. The results indicate that total epidermal glucosylceramides are composed of six distinct molecular groups, glucosylceramides 1-6. Glucosylceramide 1 contains sphingenine and nonhydroxy fatty acids, glucosylceramide 2, phytosphingosine and nonhydroxy fatty acids, glucosylceramide 3, phytosphingosine with one double bond and nonhydroxy fatty acids, glucosylceramide 4, sphingenine and alpha-hydroxy fatty acids, glucosylceramide 5, phytosphingosine and alpha-hydroxy fatty acids, and glucosylceramide 6, phytosphingosine with one double bond and alpha-hydroxy fatty acids. The nonhydroxy fatty acids typically have 16-24-carbon-length chains, whereas alpha-hydroxy fatty acids are limited to 24-, 25-, and 26-carbon chains. The sphingosine bases are C18 or C20 chains. Next, acylglucosylceramides and glucosylceramides were treated with beta-glucocerebrosidase and the ceramides released were compared with stratum corneum ceramides. Ceramide moieties of acylglucosylceramides and glucosylceramides 1, 2, 4-6 correspond to stratum corneum ceramides 1-7. These results, together with those of our previous reports characterizing epidermal sphingomyelins, indicate that all ceramide species, including omega-hydroxy fatty-acid-containing ceramides, are derived from glucosylceramides, and fractions of ceramides 2 and 5 are from sphingomyelins. Furthermore, structural analysis of glucosylceramides revealed that human epidermal glycosphingolipids display a unique lipid profile that is rich in very long chain hydroxylated (alpha- and omega-hydroxy) fatty acids and phytosphingosine.

Adolescent↗
PubMed2002

Lymph node macrophages, but not spleen macrophages, express high levels of unmasked sialoadhesin: implication for the adhesive properties of macrophages in vivo.

Sialoadhesin is a macrophage-restricted adhesion molecule that recognizes N-acetylneuraminylalpha2-3galactose structure. We prepared a multivalent neoglycoprotein probe carrying this oligosaccharide and characterized the binding activity of sialoadhesin on native rat macrophages. Macrophages from mesenteric and axillar lymph nodes exhibited 36-fold higher activity than those from the spleen. The K(d) values of the probe binding to macrophages of the two organs were indistinguishable (1-2 nM), whereas the B(max) value of lymph node macrophages was markedly higher than that of splenic macrophages. Western blot analysis revealed that the quantity of sialoadhesin present in lymph node macrophages was 25-fold higher than in splenic macrophages. High cell surface expression of sialoadhesin on lymph node macrophages was also shown by flow cytometry. To examine the "masking" of sialoadhesin by endogenous sialoglycoconjugates, we treated macrophages with sialidase before measuring the probe binding. After sialidase treatment, the binding activity of splenic macrophages increased fourfold, whereas that of lymph node macrophages did not increase. In conclusion, we have identified macrophages expressing high levels of unmasked sialoadhesin in lymph nodes. The unmasked forms on these macrophages are available for sialoadhesin-dependent adhesive functions, unlike the masked forms on the majority of splenic macrophages.

Animals↗
PubMed2002

Phylogenetic development of a regulatory gene for the core 2 GlcNAc transferase in Mus musculus.

In our previous study, we identified a mouse gene, Gsl5, that controls the expression of a glycolipid, GL-Y [Galbeta1-4(Fucalpha1-3)GlcNAcbeta1-6(Galbeta1-3)GalNAcbeta1-3Galalpha1-4Galbeta1-4Glcbeta1-Cer], and the core 2 structure of O-linked glycans of glycoproteins, GlcNAcbeta1-6(Galbeta1-3)GalNAcalpha-Ser/Thr, in a kidney tubular cell-specific manner through the regulation of UDP-GlcNAc beta-1,6-GlcNAc transferase (GNT). Regulation by the Gsl5 gene occurs at the level of GNT mRNA and the recessive allele of Gsl5 is rare and carried by DBA/2 and its related strains. Here, we report a sequence comparison of the 5' flanking region of the GNT gene among 5 laboratory strains and 10 wild-derived strains, demonstrating that the DBA/2 allele sequence is similar to the sequence carried by Asian Mus m. musculus and differs substantially from the East European M. m. musculus. These results suggest that the DBA/2 allele of Gsl5 was introduced into laboratory mouse strains by Asian wild-derived mice. Phylogenetic comparison of the 5' flanking region sequences between the recessive and dominant Gsl5 alleles indicates that mutations to create a functional Gsl5 gene occurred approximately one million years ago during the subspeciation of M. musculus, and provides a case for studies on the creation of functional genes involved in tissue-specific transcriptional regulation.

Animals↗