Serum antibodies to phosphatidylcholine in MS.

OBJECTIVE: To evaluate the value of serum immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies reactive with phosphatidylcholine (PC) and lactosylceramide (LC) as biomarkers in MS. METHODS: We developed an ultrasensitive ELISA technique to analyze serum IgG and IgM antibodies to LC and PC, which we used to analyze samples from 362 patients with MS, 10 patients with non-MS myelin diseases (Non-MSMYDs), 11 patients with nonmyelin neurologic diseases (Non-MYNDs), and 80 controls. MS serum samples included clinically isolated syndrome (CIS, n = 17), relapsing-remitting MS (RRMS, n = 62), secondary progressive MS (SPMS, n = 50), primary progressive MS (PPMS, n = 37), and benign MS (BENMS, n = 36). RESULTS: We detected higher levels of serum IgM antibodies to PC (IgM-PC) in MS than control samples; patients with CIS and RRMS showed higher IgM-PC levels than patients with SPMS, PPMS, and BENMS and controls. MS and control samples did not differ in serum levels of IgM antibodies reactive with LC, nor in IgG antibodies reactive with LC or PC. CONCLUSIONS: Serum IgM-PC antibodies are elevated in patients with MS, particularly during the CIS and RRMS phases of the disease. Thus, serum IgM-PC is a candidate biomarker for early inflammatory stages of MS. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that serum antibodies to PC are elevated in patients with MS. The study is rated Class III because of the case control design and the risk of spectrum bias: antibody levels in patients with MS were compared with healthy controls.

[Acute myelitis associated with anti-neutral glycolipid antibody].

A 48-year-old man with rapid onset of fever elevation developed acute myelitis over a period of a week. MRI of the spinal cord revealed a longitudinal T2-hyperintense intraspinal lesion extending from C6 to Th8 level. Clinical symptoms and findings resolved with immunotherapy. In serological analysis, no antibodies related to various collagen diseases, anti-aquaporin-4 (AQP4) antibody and anti-myelin oligodendrocyte glycoprotein (MOG) antibody were detected. Anti-lactosylceramide (LacCer) antibodies were detected in the acute phase of serum and cerebrospinal fluid, with titers showing decrements in the recovery phase. The present case supports the notion that acute myelitis can occur as an anti-neutral glycolipid antibody-related disorder. Anti-neutral glycolipid antibodies should be examined in future pertinent cases of myelitis.

[Lactosylceramide-enriched Lipid Raft-mediated Infection Immunity].

More than 100 years have passed since Elie Metchnikoff discovered phagocytes. As molecular biological techniques have been developed and improved, we have gained deeper knowledge about the molecular mechanisms of immunological responses to invasion. The innate immune system is the inborn defense mechanism and the first line of defense against all kinds of pathogenic organisms, including bacteria, fungi, viruses, etc. Innate immunity was originally considered to comprise non-specific reactions. However, we now know that innate immune systems develop molecular mechanisms specific to pathogenic microorganisms. In the 1970s, a neutral glycosphingolipid lactosylceramide (LacCer) was found to bind specifically to several kinds of microorganisms. LacCer is highly expressed in phagocytes and epithelial cells. LacCer forms lipid rafts on human neutrophils and is involved in neutrophil migration, phagocytosis, and superoxide generation. In contrast, mouse neutrophils express relatively little LacCer on their cell surfaces. Thus, it is difficult to observe LacCer-mediated innate immunological reactions in mice. Mycobacterium tuberculosis is a typical pathogen for humans but not mice in general. Interestingly, M. tuberculosis can escape killing by neutrophils through regulation of the LacCer-enriched lipid raft-mediated immunological reactions of these cells. These observations indicate that LacCer-enriched lipid rafts play an essential role in human innate immunity. This review describes LacCer-mediated innate immunity in humans.

[A case of acute disseminated encephalomyelitis concomitant with polyneuropathy associated with anti-lactosylceramide antibody].

We report a case of acute disseminated encephalomyelitis (ADEM) concomitant with polyneuropathy associated with anti-lactosylceramide antibody. A 68-year-old man was admitted to our hospital with ophthalmoparesis, bulbar palsy, tetraplegia after suffering from upper respiratory infection and headache. Subsequently, he developed respiratory failure requiring mechanical ventilation. Fluid-attenuated inversion recovery (FLAIR) MRI showed high intensities in the pons and medulla, and a nerve conduction study revealed motor-dominant axonal polyneuropathy. Although the laboratory tests revealed the presence of anti-lactosylceramide antibody in his serum, he was diagnosed with acute disseminated encephalomyelitis concomitant with polyneuropathy. Whereas the intensive treatment with corticosteroids, plasmapharesis, and high-dose intravenous immunoglobulin (IVIg) brought a moderate improvement, his tetraparesis continued to exist.

Lipoarabinomannan binding to lactosylceramide in lipid rafts is essential for the phagocytosis of mycobacteria by human neutrophils.

Pathogenic mycobacteria use virulence factors, including mannose-capped lipoarabinomannan (ManLAM), to survive in host phagocytic cells, such as neutrophils. We assessed the roles of lactosylceramide (LacCer, CDw17)-enriched lipid rafts in the phagocytosis of mycobacteria by human neutrophils and in the intracellular fate of phagocytosed mycobacteria. We showed that the association of the Src family kinase (SFK) Lyn with C24 fatty acid chain-containing LacCer was essential for the phagocytosis of mycobacteria by neutrophils. Assays with LacCer-containing liposomes, LacCer-coated plastic plates, and LAM-coated beads demonstrated that the phagocytosis of mycobacteria was mediated through the binding of LacCer to LAM. Both ManLAM from pathogenic species and phosphoinositol-capped LAM (PILAM) from nonpathogenic Mycobacterium smegmatis bound equivalently to LacCer to stimulate phagocytosis. However, PILAM from an M. smegmatis α1,2-mannosyltransferase deletion mutant (ΔMSMEG_4247), lacking the α1,2-monomannose side branches of the LAM mannan core, did not bind to LacCer or induce phagocytosis. An anti-LacCer antibody immunoprecipitated the SFK Hck from the phagosomes of neutrophils that internalized nonpathogenic mycobacteria but not from those that internalized pathogenic mycobacteria. Furthermore, knockdown of Hck by short inhibitory RNA abolished the fusion of lysosomes with phagosomes containing nonpathogenic mycobacteria. Further analysis showed that ManLAM, but not PILAM, inhibited the association of Hck with LacCer-enriched lipid rafts in phagosomal membranes, effectively blocking phagolysosome formation. Together, these findings suggest that pathogenic mycobacteria use ManLAM not only for binding to LacCer-enriched lipid rafts and entering neutrophils but also for disrupting signaling through Hck-coupled, LacCer-enriched lipid rafts and preventing phagolysosome formation.

Properties and functions of lactosylceramide from mouse neutrophils.

Lactosylceramide (LacCer), which is essential for many cellular processes, is highly expressed on the plasma membranes of human neutrophils and mediates innate immune functions. Less is known, however, about the properties and biological functions of LacCer in mouse neutrophils. This study therefore analyzed the properties of mouse neutrophil LacCer. LacCer was observed on the surface of these cells, with flow cytometry indicating that mouse neutrophil LacCer could be detected by the anti-LacCer mAb T5A7, but not by the anti-LacCer antibodies Huly-m13 and MEM-74. The molecular species of LacCer were nearly identical in mouse and human neutrophils, including C24:0 and C24:1 fatty acid chain-containing species, although the LacCer content in plasma membranes was ∼ 20-fold lower in mouse than in human neutrophils. Surface plasmon resonance analysis revealed that T5A7 bound to a lipid monolayer composed of LacCer, DOPC, cholesterol and sphingomyelin (molar ratio 0.1 : 10 : 10 : 1), whereas Huly-m13 did not. T5A7 induced neutrophil migration, which was abolished by inhibitors of Src-family kinases, PI-3 kinases, and trimeric G (o/i) proteins. T5A7 also inhibited phagocytosis of non-opsonized zymosans by neutrophils. Taken together, these findings suggest that in mouse neutrophils, (i) LacCer is expressed as LacCer-enriched microdomains in cell surface plasma membranes, (ii) these microdomains are recognized by T5A7 but not by other known anti-LacCer antibodies and (iii) LacCer is involved in cell migration and phagocytosis.

Fine specificity and cross-reactions of monoclonal antibodies to group B streptococcal capsular polysaccharide type III.

Group B streptococcus (GBS) is a major cause of neonatal sepsis and meningitis. Despite aggressive campaigns using antenatal prophylactic antibiotic therapy, infections continue. Developing an effective maternal vaccine is a public health priority. Antibody (Ab) to the capsular polysaccharide (CPS) is considered the dominant "protective" immune mediator. Here we study the fine specificity and potential host reactivity of a panel of well-characterized murine monoclonal Abs against the type III CPS by examining the binding of the Abs to intact and neuraminidase-digested GBS, purified CPS, synthetic carbohydrate structures, and cells. The results showed marked differences in the fine specificity among these mAbs to a single carbohydrate structure. Cross-reactions with synthetic GD3 and GT3 carbohydrates, representing structures found on surfaces of neural and developing cells, were demonstrated using carbohydrate array technology. The anti-CPS(III) mAbs did not react with cells expressing GD3 and GT3, nor did mAbs specific for the host carbohydrates cross-react with GBS, raising questions about the physiological relevance of this cross-reaction. But in the process of these investigations, we serendipitously demonstrated cross-reactions of some anti-CPS(III) mAbs with antigens, likely carbohydrates, found on human leukocytes. These studies suggest caution in the development of a maternal vaccine to prevent infection by this important human pathogen.

Production and characterization of monoclonal antibodies specific to lactotriaosylceramide.

We have established hybridoma cell lines producing monoclonal antibodies (mAbs) directed to N-acetylglucosaminylß1-3galactose (GlcNAcß1-3Gal) residue by immunizing BALB/c mice with lactotriaosylceramide (Lc(3)Cer). These obtained hybridoma cells, specific to Lc(3)Cer, were dual immunoglobulin (Ig)-producing cells which secreted both IgM and IgG molecules as antibodies. The established mAbs are able to react with not only Lc(3)Cer but also GlcNAcß1-3-terminal glycosphingolipids (GSLs) despite branching or lactosamine chain lengths and human transferrin with terminal GlcNAc residues. Comparison of the variable regions of the cloned IgM and IgG by reversed transcription-polymerase chain reaction analysis confirmed that the variable regions determine the specificity, the other amino acids are conserved, and these mAbs are encoded by J558 and Vκ-21family genes. Furthermore, we have analyzed the expression of GSLs with GlcNAcß1-3 epitope in acute leukemia cell lines and mouse fetal tissues using these mAbs, in which antigens were distributed comparatively. These mAbs are useful for studying the precise distribution of GlcNAcß1-3Gal-terminating GSL expression in tissues as well as for detecting GSLs carrying terminal GlcNAcß1-3Gal carbohydrate structure.

Epithelial GM-CSF induction by Candida glabrata.

The main cytokine induced by the interaction of oral epithelial cells with C. glabrata is granulocyte monocyte colony-stimulating factor (GM-CSF); however, the mechanisms regulating this response are unknown. Based on previously published information on the interactions of C. albicans with oral epithelial cells, we hypothesized that interaction with viable C. glabrata triggers GM-CSF synthesis via NF-kappaB activation. We found that C. glabrata-induced GM-CSF synthesis was adhesion-dependent, enhanced by endocytosis, and required fungal viability. NF-kappaB activation was noted during interaction of epithelial cells with C. glabrata, and pre-treatment with an NF-kappaB inhibitor partly inhibited GM-CSF synthesis. Blocking TLR4 with anti-TLR4 antibody did not inhibit GM-CSF production. In contrast, an anti-CDw17 antibody triggered significant inhibition of NF-kappaB activation and GM-CSF synthesis. beta-glucans did not stimulate GM-CSF synthesis, suggesting that the CDw17/NF-kappaB/GM-CSF pathway may be beta-glucan-independent. This study provides new insights into the mechanism of GM-CSF induction by C. glabrata.

Different mechanisms are involved in apoptosis induced by melanoma gangliosides on human monocyte-derived dendritic cells.

Tumor escape is linked to multiple mechanisms, notably the liberation, by tumor cells, of soluble factors that inhibit the function of dendritic cells (DC). We have shown that melanoma gangliosides impair DC differentiation and induce their apoptosis. The present study was aimed to give insight into the mechanisms involved. DC apoptosis was independent of the catabolism of gangliosides since lactosylceramide did not induce cell death. Apoptosis induced by GM3 and GD3 gangliosides was not blocked by inhibitors of de novo ceramide biosynthesis, whereas the acid sphingomyelinase inhibitor desipramine only prevented apoptosis induced by GM3. Furthermore, our results suggest that DC apoptosis was triggered via caspase activation, and it was ROS dependent with GD3 ganglioside, suggesting that GM3 and GD3 induced apoptosis through different mechanisms.

Developmental expression of the type I diabetes related antigen sulfatide and sulfated lactosylceramide in mammalian pancreas.

Previous studies have shown that sulfatide is present and functionally involved in beta cells, and that anti-sulfatide antibodies (ASA) exist during development of type I diabetes mellitus. To further explore the possible role of sulfatide in type I diabetes, developmental expression was examined in human pancreas and in pancreas of the type I diabetes models BB rat and NOD mouse compared to Lewis rat and BALB/c mouse, respectively. Sulfatide was not only expressed in adult pancreas, but also in human fetal and rodent neonatal pancreas, i.e., during the growing period of the immunological self. Sulfatide had a different expression pattern in human beings and rodents, concerning both the amounts of sulfatide and expression during development. There was no change in the sulfatide fatty acid isoform expression during development. The pancreatic expression of another sulfated glycosphingolipid, sulfated lactosylceramide, indicated that this molecule is a potential fetal/neonatal marker, which was further expressed in the type I diabetic models. In conclusion, these findings give further support to the possibility that sulfatide is a relevant autoantigen in type I diabetes and that sulfated lactosylceramide might function as a potential risk factor for disease development, at least in the animal models.

Lactosylceramide-enriched glycosphingolipid signaling domain mediates superoxide generation from human neutrophils.

This study is focused on the functional significance of neutrophil lactosylceramide (LacCer)-enriched microdomains, which are involved in the initiation of a signal transduction pathway leading to superoxide generation. Treatment of neutrophils with anti-LacCer antibody, T5A7 or Huly-m13, induced superoxide generation from the cells, which was blocked by PP1, a Src kinase inhibitor; wortmannin, a phosphatidylinositol-3 kinase inhibitor; SB203580, a p38 mitogen-activated protein kinase (MAPK) inhibitor; and H7, an inhibitor for protein kinase C. When promyelocytic leukemia HL-60 cells were differentiated into neutrophilic lineage by dimethyl sulfoxide (DMSO) treatment, they acquired superoxide-generating activity but did not respond to anti-LacCer antibodies. Density gradient centrifugation revealed that LacCer and Lyn were recovered in detergent-insoluble membrane (DIM) of neutrophils and DMSO-treated HL-60 cells. However, immunoprecipitation experiments indicated that LacCer was associated with Lyn in neutrophils but not in DMSO-treated HL-60 cells. Interestingly, T5A7 induced the phosphorylation of Lyn in neutrophils but not in DMSO-treated HL-60 cells. Moreover, T5A7 induced the phosphorylation of p38 MAPK in neutrophils. T5A7-induced Lyn phosphorylation in neutrophil DIM fraction was significantly enhanced by cholesterol depletion or sequestration with methyl-beta-cyclodextrin or nystatin. Collectively, these data suggest that neutrophils are characterized by the presence of cell surface LacCer-enriched glycosphingolipid signaling domain coupled with Lyn and that the ligand binding to LacCer induces the activation of Lyn, which may be suppressibly regulated by cholesterol, leading to superoxide generation through the phosphatidylinositol-3 kinase-, p38 MAPK-, and protein kinase C-dependent signal transduction pathway.

Antibodies to GD3, GT3, and O-acetylated species in Guillain-Barré and Fisher's syndromes: their association with cranial nerve dysfunction.

We examined serum antibodies to the four fetal antigens GD3, O-acetyl GD3, GT3, and O-acetyl GT3 ganglioside in patients with Guillain-Barré syndrome (GBS) or its variant Fisher's syndrome (FS). The patients with FS more often had significant IgG antibodies against GD3, GT3, and O-acetyl GT3 than did the healthy controls. Furthermore, anti-GD3 and anti-GT3 IgG antibodies were more often significantly present in the patients with FS than in those with GBS. IgG antibody to GD3, GT3, and O-acetyl GT3 had a significant association with the presence of ophthalmoparesis. These antibodies, however, cross-reacted with GQ1b and we detected no antibodies which specifically reacted with fetal gangliosides. In addition, oculomotor involvement was more closely related to IgG antibodies to GQ1b than to those to fetal gangliosides. No evidence was obtained that the serum antibodies to these fetal gangliosides are associated with specific neurologic signs of cranial nerves.

Developmentally regulated O-acetylated sialoglycans in the central nervous system revealed by a new monoclonal antibody 493D4 recognizing a wide range of O-acetylated glycoconjugates.

We have previously detected an alkali-labile and developmentally regulated antigen in rat embryonic cerebral cortex, which may be 9-O-acetylsialylated GT3 ganglioside (Hirabayashi Y, Hirota M, Suzuki Y, Matsumoto M, Obata K, Ando S (1989) Neurosci Lett 106:193-98). In this study we established a mouse monoclonal antibody, 493D4, that recognizes 9-O-acetyl GT3 ganglioside, but not non-O-acetyl gangliosides. This antibody also reacted with 9-O-acetyl GD3 to a much lesser extent. By using this antibody, we found that O-acetyl GT3 as well as O-acetyl GD3 were expressed strongly in fetal murine cerebral cortex and decreased to an undetectable level after birth. With the assistance of TLC-immunostaining using 493D4 together with Q-Sepharose column chromatography, O-acetyl gangliosides of bovine brain were purified and the structural analysis showed the presence of O-acetyl GD3, O-acetyl LD1, O-acetyl GD2 and O-acetyl GD1b in the adult brain as extremely minor components. Interestingly, the antibody 493D4 could detect O-acetyl sialoglycoproteins in rat brain tissues. One of the major immunoreactive proteins was shown to be synaptophysin, an integral membrane protein specifically present in synaptic vesicles. This monoclonal antibody was therefore useful for sensitive detection of both O-acetylated gangliosides and glycoproteins with O-acetylated sialic acids.

Compensatory occurrence of IV2Fucalpha,II3NeuAcalpha-Gg4Cer and human fetal antigen Lc4Cer in small cell carcinomas of the human lung.

By means of a thin-layer chromatography immunostaining procedure involving a human monoclonal anti-Lc4Cer antibody, which was established by hybridizing murine myeloma cells and human lymphocytes from a cancer patient, Lc4Cer was proven to be a fetal antigen of human lung and to be a cancer-related antigen in small cell carcinomas of human lung, but not of other lung cancers, i.e., large cell carcinomas, adenocarcinomas, and squamous carcinomas. With the simultaneous detection of IV2Fuc alpha,II3NeuAc alpha-Gg4Cer with rabbit anti-IV2Fuc alpha,II3NeuAc alpha-Gg4Cer antiserum, the expression of Lc4Cer and IV2Fuc alpha,II3NeuAc alpha-Gg4Cer was found to be compensatory and, consequently, small cell lung carcinomas could be classified into Lc4Cer- and IV2Fuc alpha,II3NeuAc alpha-Gg4Cer-expressing types, L-SCLC and F-SCLC, respectively, which were detected in four and 27 of 31 patients' tissues and in one and three of four nude mouse-transplanted small cell lung carcinoma tissues, respectively. The compensatory expression of Lc4Cer and IV2Fuc alpha,II3NeuAc alpha-Gg4Cer in small cell carcinomas indicated that different metabolic pathways for glycosphingolipids were activated to give the distinct glycosphingolipid compositions in the two types of small cell lung carcinomas.

Galactose to ceramide linkage is essential for the binding of a polyclonal antibody to galactosyl ceramide.

Characterization of a polyclonal antibody to galactosyl ceramide (Gal-Cer) which inhibits the internalization and infection of HIV-1 in neural cell lines was carried out. Polyclonal antibody to Gal-Cer was produced by injecting rabbits with Gal-Cer liposomes. The specificity of anti-Gal-Cer binding was studied by high performance thin layer chromatography (HPTLC)-based immunoassay. Using natural and semisynthetic lipids, the specificity of anti-Gal-Cer interaction was studied. The antibody bound to Gal-Cer and its derivatives. The antibody did not bind to glucosyl ceramide or lactosyl ceramide. Glucosyl ceramide differs from Gal-Cer by a hydroxyl group at the fourth carbon and in lactosyl ceramide galactose is linked to ceramide by an intervening glucose molecule. This indicates that D-galactose linked to ceramide is essential for binding. Removal of fatty acid from Gal-Cer, as seen with N-palmitoyl- and N-oleoyl Gal-Cer, had no effect on the binding. It appears that the third carbon of Gal-Cer is not involved in the binding. This is supported by the binding of anti-Gal-Cer to sulfatide or GM4 in which sulfate or sialic acid are added at the third carbon of Gal-Cer, respectively.

Monoclonal antibody A2B5, which detects cell surface antigens, binds to ganglioside GT3 (II3 (NeuAc)3LacCer) and to its 9-O-acetylated derivative.

The monoclonal antibody A2B5 recognizes antigens at the surface of neuronal and glial cells but also at the surface of thymus epithelia and pancreatic islet cells. Although these antigens have been characterized as polysialogangliosides, A2B5 also reacts with other unidentified gangliosides. In order to characterize further the epitope of A2B5, two new ganglioside antigens isolated from chicken brain are identified in this study. One is the ganglioside NeuAc alpha 2-8NeuAc alpha 2-8NeuAc alpha 2-3Gal beta 1-4Glc beta 1-1ceramide (GT3) and the other is a 9-O-acetylated derivative of GT3). This derivative was purified from 10-day embryonic chicken brain. Acetyl groups substituted on sialic acid were removed either by alkali treatment or by incubation with influenza virus C, which contains receptor-destroying enzyme (a neuraminidate 9-O-acetyl esterase). The product of alkali treatment or viral action was detected by the antibody 18B8 which is specific for GT3. The deacetylated product still reacts with A2B5. These data and the results of mild oxidation of the antigen with sodium periodate suggest that the epitope recognized by antibody A2B5 contains the trisialyl structure found in GT3 but does not include the polyalcohol chain of the terminal sialic acid which can be oxidized by periodate or acetylated without modifying the affinity for the antibody. The epitope recognized by A2B5 is different from the epitope recognized by the antibody 18B8 in that 18B8 requires the three sialic acids with an intact and unsubstituted polyalcohol chain. Antibody 18B8 does not bind to 9-O-acetylated GT3 or GT3 oxidized by sodium periodate.

Human monoclonal antibody (HMST-1) against lacto-series type 1 chain and expression of the chain in uterine endometrial cancers.

A human monoclonal antibody termed HMST-1 was produced by fusing lymphocytes from segments of human pelvic lymph nodes from an endometrial cancer patient with murine myeloma cells. The epitope recognized by HMST-1 was determined to be lacto-series type 1 chain-containing glycosphingolipid (Gal beta 1-3GlcNAc beta 1-3Gal beta 1-4Glc beta 1-1Cer) by isolating the antigen from endometrial cancer cell line SNG-II and analyzing with fast atom bombardment mass spectrometry, permethylation analysis, and exoglycosidase treatment. By the immunohistochemical avidin-biotin-peroxidase complex method, no normal endometrium and benign endometrial hyperplasia were stained with HMST-1, but HMST-1 reacted with about 35% of endometrial cancer cases. These facts indicate that the rate of expression of the antigen increases along with the course of malignancy in the endometrium. By sialidase treatment of the section, the positive rate increased to 57% in endometrial cancers and to 13% in normal endometrium, indicating that the antigen was masked with sialic acid and exposed by neuraminidase treatment. Immunohistochemistry also revealed that the antibody reacted with human fetal alimentary tract epithelium and mesothelium, indicating the oncodevelopmental nature of Gal beta 1-3GlcNAc beta 1-3Gal beta 1-4Glc beta 1-1Cer.

Campylobacter pylori colonization factor shows specificity for lactosylceramide sulfate and GM3 ganglioside.

The specificity of Campylobacter pylori cell surface lectin, a presumptive colonization factor, was investigated using various sulfated and sialic acid containing glycolipids. C. pylori cells, cultured from human antral mucosal biopsies, were incubated with intact and modified glycolipid preparations and examined for agglutination inhibition of human erythrocytes. Titration data revealed that the inhibitory activity was highest with lactosylceramide sulfate and GM3 ganglioside, while galactosylceramide sulfate GM1, GD1a and GD1b gangliosides were less effective. A strong inhibitory activity towards C. pylori hemagglutin was also observed with an antiulcer agent, sucralfate. The inhibitory effect of both types of glycolipids was abolished by the removal of sialic acid and sulfate ester groups, thus indicating that sulfated and sialic acid containing glycolipids with terminal lactosyl moieties serve as mucosal receptors for colonization of gastric epithelium by C. pylori.