scholarly journals A novel M cell–specific carbohydrate-targeted mucosal vaccine effectively induces antigen-specific immune responses

2007 ◽  
Vol 204 (12) ◽  
pp. 2789-2796 ◽  
Author(s):  
Tomonori Nochi ◽  
Yoshikazu Yuki ◽  
Akiko Matsumura ◽  
Mio Mejima ◽  
Kazutaka Terahara ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Oral Vaccine ◽  
Goblet Cells ◽  
Carbohydrate Moiety ◽  
Mucosal Vaccine ◽  
M Cells ◽  
M Cell ◽  
Lethal Challenge ◽  
New Strategy ◽  
High Level

Mucosally ingested and inhaled antigens are taken up by membranous or microfold cells (M cells) in the follicle-associated epithelium of Peyer's patches or nasopharynx-associated lymphoid tissue. We established a novel M cell–specific monoclonal antibody (mAb NKM 16–2-4) as a carrier for M cell–targeted mucosal vaccine. mAb NKM 16–2-4 also reacted with the recently discovered villous M cells, but not with epithelial cells or goblet cells. Oral administration of tetanus toxoid (TT)– or botulinum toxoid (BT)–conjugated NKM 16–2-4, together with the mucosal adjuvant cholera toxin, induced high-level, antigen-specific serum immunoglobulin (Ig) G and mucosal IgA responses. In addition, an oral vaccine formulation of BT-conjugated NKM 16–2-4 induced protective immunity against lethal challenge with botulinum toxin. An epitope analysis of NKM 16–2-4 revealed specificity to an α(1,2)-fucose–containing carbohydrate moiety, and reactivity was enhanced under sialic acid–lacking conditions. This suggests that NKM 16–2-4 distinguishes α(1,2)-fucosylated M cells from goblet cells containing abundant sialic acids neighboring the α(1,2) fucose moiety and from non-α(1,2)-fucosylated epithelial cells. The use of NKM 16–2-4 to target vaccine antigens to the M cell–specific carbohydrate moiety is a new strategy for developing highly effective mucosal vaccines.

scholarly journals M CELLS ARE THE IMPORTANT POST IN THE INITIATION OF IMMUNE RESPONSE IN INTESTINE

2018 ◽  
Vol 8 (3) ◽  
pp. 263-272
Author(s):  
A. S. Bykov ◽  
A. V. Karaulov ◽  
D. A. Tsomartova ◽  
N. L. Kartashkina ◽  
V. L. Goriachkina ◽  
...  
Keyword(s):  
Immune System ◽  
Epithelial Cells ◽  
Immune Responses ◽  
Targeted Delivery ◽  
Oral Vaccine ◽  
Critical Event ◽  
M Cells ◽  
M Cell ◽  
Antigen Uptake ◽  
And Function

Microfold cells (M cells) are specialized intestinal epithelial cells that initiate mucosal immune responses. These unique phagocytic epithelial cells are specialized for the transfer of a broad range of particulate antigens and microorganisms across the follicle-associated epithelium (FAE) into the gut-associated lymphoid tissue (GALT) by a process termed transcytosis. The molecular basis of antigen uptake by M cells has been gradually identified in the last decade. Active sampling of intestinal antigen initiates regulated immune responses that ensure intestinal homeostasis. The delivery of luminal substances across the intestinal epithelium to the immune system is a critical event in immune surveillance resulting in tolerance to dietary antigens and immunity to pathogens (e.g., bacteria, viruses, and parasites) and their toxins. Several specialized mechanisms transport luminal antigen across the gut epithelium. Discovery of M cell-specific receptors are of great interest, which could act as molecular tags for targeted delivery oral vaccine to M cells. Recent studies demonstrated that M cells utilize several receptors to recognize and transport specific luminal antigens. Vaccination through the mucosal immune system can induce effective systemic immune responses simultaneously with mucosal immunity. How this process is regulated is largely unknown. This review aims to show a new understanding of the factors that influence the development and function of M cells; to show the molecules expressed on M cells which appear to be used as immunosurveillance receptors to sample pathogenic microorganisms in the gut; to note how certain pathogens appear to exploit M cells to inject the host; and, finally, how this knowledge is used to specifically "target" antigens to M cells to attempt to improve the efficacy of mucosal vaccines. Recently, substantial progress has been made in our understanding of the factors that influence the development and function of M cells.

scholarly journals Role of the glycocalyx in regulating access of microparticles to apical plasma membranes of intestinal epithelial cells: implications for microbial attachment and oral vaccine targeting.

1996 ◽  
Vol 184 (3) ◽  
pp. 1045-1059 ◽  
Author(s):  
A Frey ◽  
K T Giannasca ◽  
R Weltzin ◽  
P J Giannasca ◽  
H Reggio ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Plasma Membranes ◽  
Intestinal Epithelial Cells ◽  
Oral Vaccine ◽  
M Cells ◽  
Intestinal Epithelial ◽  
M Cell ◽  
Antigen Uptake ◽  
Apical Membranes

Transepithelial transport of antigens and pathogens across the epithelial barrier by M cells may be a prerequisite for induction of mucosal immunity in the intestine. Efficient transport of antigens and pathogens requires adherence to M cell apical surfaces. Coupling of antigen-containing particles to the pentameric binding subunit of cholera toxin (CTB) has been proposed as a means for increasing antigen uptake because the CTB receptor, ganglioside GM1, is a glycolipid present in apical membranes of all intestinal epithelial cells. To test the accessibility of enterocyte and M cell membrane glycolipids to ligands in the size ranges of viruses, bacteria, and particulate mucosal vaccines, we analyzed binding of CTB probes of different sizes to rabbit Peyer's patch epithelium. Soluble CTB-fluorescein isothiocyanate (diameter 6.4 nm) bound to apical membranes of all epithelial cells. CTB coupled to 14 nm colloidal gold (final diameter, 28.8 nm) failed to adhere to enterocytes but did adhere to M cells. CTB-coated, fluorescent microparticles (final diameter, 1.13 microns) failed to adhere to enterocytes or M cells in vivo or to well-differentiated Caco-2 intestinal epithelial cells in vitro. However, these particles bound specifically to GM1 on BALB/c 3T3 fibroblasts in vitro and to undifferentiated Caco-2 cells that lacked brush borders and glycocalyx. Measurements of glycocalyx thickness by electron microscopy suggested that a relatively thin (20 nm) glycocalyx was sufficient to prevent access of 1-micron microparticles to glycolipid receptors. Thus, the barrier function of the intestinal epithelial cell glycocalyx may be important in limiting microbial adherence to membrane glycolipids, and in CTB-mediated targeting of vaccines to M cells and the mucosal immune system.

scholarly journals Comparative Analysis of EspF Variants in Inhibition of Escherichia coli Phagocytosis by Macrophages and Inhibition of E. coli Translocation through Human- and Bovine-Derived M Cells

2011 ◽  
Vol 79 (11) ◽  
pp. 4716-4729 ◽  
Author(s):  
Amin Tahoun ◽  
Gabriella Siszler ◽  
Kevin Spears ◽  
Sean McAteer ◽  
Jai Tree ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epithelial Cells ◽  
Cellular Localization ◽  
M Cells ◽  
Binding Assays ◽  
M Cell ◽  
Content Type ◽  
E Coli ◽  
Coculture System

ABSTRACTThe EspF protein is secreted by the type III secretion system of enteropathogenic and enterohemorrhagicEscherichia coli(EPEC and EHEC, respectively). EspF sequences differ between EHEC O157:H7, EHEC O26:H11, and EPEC O127:H6 in terms of the number of SH3-binding polyproline-rich repeats and specific residues in these regions, as well as residues in the amino domain involved in cellular localization. EspFO127is important for the inhibition of phagocytosis by EPEC and also limits EPEC translocation through antigen-sampling cells (M cells). EspFO127has been shown to have effects on cellular organelle function and interacts with several host proteins, including N-WASP and sorting nexin 9 (SNX9). In this study, we compared the capacities of differentespFalleles to inhibit (i) bacterial phagocytosis by macrophages, (ii) translocation through an M-cell coculture system, and (iii) uptake by and translocation through cultured bovine epithelial cells. TheespFgene fromE. coliserotype O157 (espFO157) allele was significantly less effective at inhibiting phagocytosis and also had reduced capacity to inhibitE. colitranslocation through a human-derivedin vitroM-cell coculture system in comparison toespFO127andespFO26. In contrast,espFO157was the most effective allele at restricting bacterial uptake into and translocation through primary epithelial cells cultured from the bovine terminal rectum, the predominant colonization site of EHEC O157 in cattle and a site containing M-like cells. Although LUMIER binding assays demonstrated differences in the interactions of the EspF variants with SNX9 and N-WASP, we propose that other, as-yet-uncharacterized interactions contribute to the host-based variation in EspF activity demonstrated here.

scholarly journals Identifying human and murine M cells in vitro

2019 ◽  
Vol 244 (7) ◽  
pp. 554-564 ◽  
Author(s):  
Ana Klisuric ◽  
Benjamin Thierry ◽  
Ludivine Delon ◽  
Clive A Prestidge ◽  
Rachel J Gibson
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
In Vitro Models ◽  
Oral Drug ◽  
M Cells ◽  
Cell Marker ◽  
M Cell ◽  
Follicle Associated Epithelium

M cells are an epithelial cell population found in the follicle-associated epithelium overlying gut-associated lymphoid tissues. They are specialized in the transcytosis of luminal antigens. Their transcytotic capacity and location in an immunocompetent environment has prompted the study of these cells as possible targets for oral drug delivery systems. Currently, the models most commonly used to study M cells are restricted to in vivo experiments conducted in mice, and in vitro studies conducted in models comprised either of primary epithelial cells or established cell lines of murine or human origin. In vitro models of the follicle-associated epithelium can be constructed in several ways. Small intestinal Lgr5+ stem cells can be cultured into a 3D organoid structure where M cells are induced with RANKL administration. Additionally, in vitro models containing an “M cell-like” population can be obtained through co-culturing intestinal epithelial cells with cells of lymphocytic origin to induce the M cell phenotype. The evaluation of the efficiency of the variations of these models and their relevance to the in vivo human system is hampered by the lack of a universal M cell marker. This issue has also hindered the advancement of M cell-specific targeting approaches aimed at improving the bioavailability of orally administered compounds. This critical review discusses the different approaches utilized in the literature to identify M cells, their efficiency, reliability and relevance, in the context of commonly used models of the follicle-associated epithelium. The outcome of this review is a clearly defined and universally recognized criteria for the assessment of the relevance of models of the follicle-associated models currently used. Impact statement The study of M cells, a specialized epithelial cell type found in the follicle-associated epithelium, is hampered by the lack of a universal M cell marker. As such, many studies lack reliable and universally recognized methods to identify M cells in their proposed models. As a result of this it is difficult to ascertain whether the effects observed are due to the presence of M cells or an unaccounted variable. The outcome of this review is the thorough evaluation of the many M cell markers that have been used in the literature thus far and a proposed criterion for the identification of M cells for future publications. This will hopefully lead to an improvement in the quality of future publications in this field.

scholarly journals Inoculum Composition and SalmonellaPathogenicity Island 1 Regulate M-Cell Invasion and Epithelial Destruction by Salmonella typhimurium

1998 ◽  
Vol 66 (2) ◽  
pp. 724-731 ◽  
Author(s):  
M. Ann Clark ◽  
Barry H. Hirst ◽  
Mark A. Jepson
Keyword(s):  
Salmonella Typhimurium ◽  
Cell Invasion ◽  
Cultured Cells ◽  
Oral Vaccine ◽  
Luria Bertani ◽  
M Cells ◽  
Wild Type ◽  
M Cell ◽  
Experimental Conditions ◽  
Phosphate Buffered Saline

ABSTRACT In the mouse model of Salmonella typhimurium infection, the specialized antigen-sampling intestinal M cells are the primary route of Salmonella invasion during the early stages of infection. Under certain experimental conditions, M-cell invasion is accompanied by M-cell destruction and loss of adjacent regions of the follicle-associated epithelium (FAE), although the conditions responsible for expression of the cytotoxic phenotype in a proportion of previous studies have not been defined. In the present study, we have demonstrated that the cytotoxic effect exerted by wild-typeS. typhimurium on mouse Peyer’s patch FAE is dependent on the inoculum composition. We have also demonstrated that the extent of FAE destruction correlates with the extent of M-cell invasion. Bacteria inoculated in Luria-Bertani (LB) broth induce extensive FAE loss and exhibit efficient M-cell invasion, whereas bacteria inoculated in phosphate-buffered saline fail to induce significant FAE disruption and invade M cells at significantly lower levels. Similarly, inoculation in LB significantly enhances invasion of Madin-Darby canine kidney cells by wild-type S. typhimurium. Mutants defective for expression of invA, a component of Salmonellapathogenicity island 1 which is vital for efficient invasion of cultured cells, fail to induce FAE destruction and, when inoculated in LB, are attenuated for M-cell invasion. Variation in invgene expression is, therefore, one possible mechanism by which inoculate composition may regulate the virulence of wild-type S. typhimurium. Our findings suggest that the composition of the gut luminal contents may be critical in determining the outcome of naturally acquired Salmonella infections and that both vaccine formulation and dietary status of vaccine recipients may significantly affect the efficacy and safety of liveSalmonella oral vaccine delivery systems.

scholarly journals M-cells in the rabbit tonsil exhibit distinctive glycoconjugates in their apical membranes.

1996 ◽  
Vol 44 (9) ◽  
pp. 1033-1042 ◽  
Author(s):  
A Gebert
Keyword(s):  
Epithelial Cells ◽  
Immune Responses ◽  
Cell Types ◽  
Lectin Histochemistry ◽  
Lymphoid Cells ◽  
M Cells ◽  
M Cell ◽  
Double Labeling ◽  
Specific Composition ◽  
Apical Membranes

The tonsil crypt epithelium contains membranous (M)-cells that transport antigens from the lumen to underlying lymphoid cells, thereby initiating specific immune responses. Mechanisms mediating the adhesion of antigens to the M-cell surface are important for effective and selective uptake of potential pathogens but are still poorly understood. Therefore, the carbohydrates present on crypt epithelial cells of the rabbit palatine tonsil were studied by lectin histochemistry. Ultrathin LR White sections were incubated with a panel of eight lectins conjugated to colloidal gold or biotin. The glycocalyx of the apical membrane of M-cells was selectively labeled by UEA-I, LTA, HPA, and VVA, whereas that of the remaining squamous epithelial cells preferentially bound RCA-I and PNA. WGA and ConA showed only little binding, with no discernible preference for any of the cell types. Double labeling of UEA-1 together with anti-vimentin antibodies revealed that UEA-I-positive epithelial cells also contained the rabbit M-cell marker vimentin, and vice versa. The results show that a specific composition of glycoconjugates, which differs from that on squamous epithelial cells, is found on M-cells of the rabbit tonsil. The M-cell-specific glycoproteins and glycolipids could be selectively targeted by microorganisms that adhere to M-cells and enter the host along this pathway.

scholarly journals Mycobacterium avium subsp. paratuberculosis Fibronectin Attachment Protein Facilitates M-Cell Targeting and Invasion through a Fibronectin Bridge with Host Integrins

2004 ◽  
Vol 72 (7) ◽  
pp. 3724-3732 ◽  
Author(s):  
T. E. Secott ◽  
T. L. Lin ◽  
C. C. Wu
Keyword(s):  
Epithelial Cells ◽  
Mycobacterium Avium ◽  
Cell Invasion ◽  
M Cells ◽  
Intestinal Epithelial ◽  
M Cell ◽  
Attachment Protein ◽  
Β3 Integrin ◽  
Mycobacterium Avium Subsp Paratuberculosis

ABSTRACT Efficient attachment and ingestion of Mycobacterium avium subsp. paratuberculosis by cultured epithelial cells requires the expression of a fibronectin (FN) attachment protein homologue (FAP-P) which mediates FN binding by M. avium subsp. paratuberculosis. Invasion of Peyer's patches by M. avium subsp. paratuberculosis occurs through M cells, which, unlike other intestinal epithelial cells, express integrins on their luminal faces. We sought to determine if the interaction between FAP-P of M. avium subsp. paratuberculosis and soluble FN enabled targeting and invasion of M cells by M. avium subsp. paratuberculosis in vivo via these surface integrins. Wild-type and antisense FAP-P mutant M. avium subsp. paratuberculosis strains were injected alone or coinjected with blocking peptides or antibodies into murine gut loops, and immunofluorescence microscopy was performed to assess targeting and invasion of M cells by M. avium subsp. paratuberculosis. Nonopsonized M. avium subsp. paratuberculosis preferentially invaded M cells in murine gut loops. M-cell invasion was enhanced 2.6-fold when M. avium subsp. paratuberculosis was pretreated with FN. Invasion of M cells by the antisense FAP-P mutant of M. avium subsp. paratuberculosis was reduced by 77 to 90% relative to that observed for the control strains. Peptides corresponding to the RGD and synergy site integrin recognition regions of FN blocked M. avium subsp. paratuberculosis invasion of M cells by 75 and 45%, respectively, whereas the connecting segment 1 peptide was noninhibitory. Antibodies against the α5, αV, β1, and β3 integrin subunits inhibited M-cell invasion by 52 to 73%. The results indicate that targeting and invasion of M cells by M. avium subsp. paratuberculosis in vivo is mediated primarily by the formation of an FN bridge formed between FAP-P of M. avium subsp. paratuberculosis and integrins on M cells.

scholarly journals Comparison of Early Ileal Invasion by Salmonella enterica Serovars Choleraesuis and Typhimurium

2003 ◽  
Vol 40 (4) ◽  
pp. 371-375 ◽  
Author(s):  
D. K. Meyerholz ◽  
T. J. Stabel
Keyword(s):  
Goblet Cells ◽  
Cell Swelling ◽  
M Cells ◽  
M Cell ◽  
Phenotypic Differences ◽  
Cellular Invasion ◽  
Cell Internalization ◽  
Serovar Typhimurium ◽  
A Site ◽  
Cell Extrusion

The mechanisms of Salmonella serovar-host specificity are not well defined. Pig ileal loops were used to compare phenotypic differences in early cellular invasion between non-host-adapted Salmonella serovar Typhimurium (SsT) and host-adapted Salmonella serovar Choleraesuis (SsC). By 10 minutes postinoculation, both serovars invaded a small number of M cells, enterocytes, and goblet cells. Multiple SsC organisms (up to 6 per cell) simultaneously invaded M cells, whereas SsT often invaded as one to two organisms per M cell. Internalization of both serovars resulted in vacuoles containing a single bacterium. The follicle-associated epithelium (FAE) of SsC-inoculated loops responded with more filopodia and lamellipodia although exhibiting less cell swelling than SsT. Additionally, SsT showed an enhanced affinity for sites of cell extrusion compared with SsC at 60 minutes. These results suggest: 1) both SsC and SsT exhibit non-cell-specific invasion as early as 10 minutes postinoculation, 2) Salmonella serovars exhibit differences in early invasion of FAE and M cells, and 3) cells undergoing extrusion may provide a site for preferential adherence by SsT and SsC.

scholarly journals Differential expression of lectin-binding sites defines mouse intestinal M-cells.

1993 ◽  
Vol 41 (11) ◽  
pp. 1679-1687 ◽  
Author(s):  
M A Clark ◽  
M A Jepson ◽  
N L Simmons ◽  
T A Booth ◽  
B H Hirst
Keyword(s):  
Electron Microscope ◽  
Laser Scanning ◽  
Lectin Binding ◽  
Fluorescein Isothiocyanate ◽  
Goblet Cells ◽  
Confocal Laser ◽  
M Cells ◽  
M Cell ◽  
Ulex Europaeus ◽  
Transmission Electron

We investigated the binding of four lectins to the follicle-associated epithelium (FAE) overlying fixed mouse small intestinal Peyer's patches to identify M-cell-specific surface markers. Wheat germ agglutinin and peanut agglutinin displayed heterogeneous staining patterns, binding most avidly to the intestine goblet cells. In contrast, the lectins Ulex europaeus 1 (UEA 1) and Psophocarpus tetragonolobus (winged bean; WBA) were almost exclusively M-cell specific. When confocal laser scanning images of tissues stained with fluorescein isothiocyanate (FITC)-conjugated UEA1 or WBA were compared with the appearance of the same tissues under the scanning electron microscope (SEM), UEA1 strongly stained 97.2% (106/109) of M-cells, 0.6% (3/516) enterocytes, and 0% (0/28) goblet cells, whereas WBA stained 100% (83/83) M-cells, 1.7% (6/361) enterocytes, and 5.3% (1/19) goblet cells. The M-cell specificity of the lectin binding was further demonstrated by localization of horseradish peroxidase (HRP)-conjugated lectins under the transmission electron microscope (TEM). This is the first demonstration of carbohydrates in the glycocalyx of M-cells that are not expressed elsewhere on the FAE surface. These carbohydrates not only provide a means to identify mouse M-cells by LM but may also contribute to the occurrence of specific interactions between microorganisms and the M-cell apical membrane.

scholarly journals Cloning, expression, and purification of the M cell targeting peptide CPE16 derived from C-terminus of Clostridium perfringens enterotoxin and the binding evaluation with Claudin-r4

2019 ◽  
Vol 3 (1) ◽  
pp. 38-45
Author(s):  
Huynh Kien Quang ◽  
Mai Quoc Gia ◽  
Nguyen Hoang An ◽  
Vo Thi Thanh Ha ◽  
Tran Van Hieu
Keyword(s):  
Clostridium Perfringens ◽  
Recombinant Plasmid ◽  
Oral Vaccine ◽  
Restriction Enzymes ◽  
Soluble Form ◽  
M Cells ◽  
M Cell ◽  
Binding Ability ◽  
Clostridium Perfringens Enterotoxin ◽  
C Terminus

Developing the oral vaccine that stimulates the mucosal immune system in order to prevent the gastro-intestinal infection is an indispensable demand nowadays. Targeting the M cells, which is a sampling antigen cell, is a highly efficient solution to prevent the dispersion of antigens. Many researches demonstrate that C-terminus Clostridium perfringens enterotoxin bounds to the Claudin- 4 receptor on the M cell surface. By using bioinformatics methods, the peptide CPE16 (16 amino acid of C-terminus of Clostridium perfringens enterotoxin) was predicted to have a high affinity to Claudin-4 receptor on M cells. In this present study, CPE16-GFP was produced as a resource to assess the binding ability to M cells. Recombinant plasmid pET22b-cpe16-gfp was constructed through cloning cpe16-gfp gene into pET22b by two restriction enzymes, NdeI and XhoI, respectively. The recombinant plasmid was transformed into E. coli BL21 (DE3) strain. The expression of protein CPE16-GFP was induced by 0.5 mM IPTG and confirmed by SDS-PAGE analysis and Western blot probed with anti-6xHis antibody. CPE16-GFP protein was expressed in soluble form. CPE16- GFP was purified by using immobilized-metal affinity chromatography with the purity up to 94.14 percent. Finally, CPE16 was tested for the binding ability to recombinant GST-claudin-R4 with the use of silicon nanowire (SiNW-FET). The result showed that CPE16 interacted with GST-claudin-R4 presented by the change of the current through nanowire, compared to its counterpart control GST.  

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