In vitro interaction of Mycobacterium avium with intestinal epithelial cells

Mapother, M E; Songer, J. Glenn

doi:10.1128/iai.45.1.67-73.1984

Cited by 54 publications

(18 citation statements)

References 21 publications

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“…Host response to M. avium-M. intracellulare may also involve these cells. Human intestinal epithelial cell monolayers have been shown to bind M. avium-M. intracellulare (25), which may explain the common intestinal infestation of AIDS patients with M. avium-M. intracellulare (41). Identification of a receptor for M. avium-M. intracellulare offers additional avenues by which infection due to M. avium-M. intracellulare could be treated or prevented by developing drugs which would compete with the bacterial ligand for the receptor or prevent adhesion to host tissue.…”

Section: Discussionmentioning

confidence: 99%

Mycobacterium avium-M. intracellulare binds to the integrin receptor alpha v beta 3 on human monocytes and monocyte-derived macrophages

1993

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Mycobacterium avium-M. intracellulare is an intracellular pathogen responsible for the highest incidence of disseminated bacterial infection in patients with AIDS. Treatment of the infection is difficult and has been of limited efficacy. Attachment of the organism to macrophages is a critical early step in the establishment of the disease. In the present study, we isolated and identified a receptor that mediates attachment of M. avium-M. intracelulare to human peripheral blood monocytes and monocyte-derived macrophages. On Western blotting, (immunoblotting), the receptor was found to cross-react with antibodies against a human vitronectin receptor (aEv33). The receptor could be purified from monocyte extracts by using monoclonal antibodies (MAbs) against the av subunit of vitronectin receptor coupled to CNBr-Sepharose 4B, as well as with the adhesive tripeptide sequence arginine-glycine-aspartic acid (RGD) coupled to CNBr-Sepharose 4B. Surface-bound MAbs directed against aV143 were found to inhibit the attachment of M. avium-M. intraceflulare to monocyte-derived macrophages in an in vitro inhibition assay, while MAbs directed against CD14, CD18, a21 and platelet glycoprotein gpIIb/IHIa receptors did not inhibit this attachment. These observations suggest that (VO3 on the surface of human monocytes and monocyte-derived macrophages may function as a receptor for M. avium-M. intraceldulare. Identification of a receptor for M. avium-M. intracellulare on macrophages may offer new approaches to the prevention and control of M. avium-M. intracelulare infection at the cellular level. receptors involved have been identified (5). However, monocytes, macrophages, and stimulated polymorphonuclear cells (but not lymphocytes or unstimulated polymorphonuclear cells) have also been shown to bind M. avium-M. intracellulare in the absence of complement, serum, or serum proteins. This binding was shown to be mediated by a proteinaceous receptor on macrophages (8). Here we report that the integrin receptor CLV,3 on monocytes and monocytederived macrophages (MDM) which is also known as vitronectin receptor (20,28,31), binds to M. avium-M. intracellulare in the absence of complement. MATERIALS AND METHODSGrowth and labeling of M. avium-M. intracelulare. M. avium-M. intracellulare obtained from the American Type Culture Collection (ATCC 25291, Rockville, Md.) was cultured in Middlebrook 7H11 broth (Difco Laboratories, Detroit, Mich.) at 37°C in 5% CO2 for 7 to 14 days with vigorous agitation once a day. Bacteria were harvested by centrifuging the medium at 800 x g for 15 min. In order to label the bacteria, the pellet was resuspended in a 1-mg/ml solution of fluorescein isothiocyanate (FITC) (Sigma Chemical Co., St. Louis, Mo.) in 50 mM Na2CO3-100 mM NaCl (pH 9.2) and incubated in the dark for 30 min at room temperature. The suspension was then washed two times with phosphatebuffered saline (PBS) containing 1 mM CaCl2 and 1 mM MgCl2 (Dulbecco's PBS or DPBS) and resuspended in the same buffer. The final suspension was sonicated for 4 s,...

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Section: Discussionmentioning

confidence: 99%

Mycobacterium avium-M. intracellulare binds to the integrin receptor alpha v beta 3 on human monocytes and monocyte-derived macrophages

1993

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“…Mycobacteria exhibit a tropism for the lungs, and interactions of the tubercle bacillus with alveolar macrophages have been extensively documented (Schlesinger et al, 1990;Schlesinger, 1993;Stokes et al, 1993). However, several reports indicate that mycobacteria are also able to bind to, invade and replicate within epithelial cells (Shepard, 1955;Mapother and Sanger, 1984;Bermudez and Goodman, 1996). Therefore, epithelial cells may represent a niche for the mycobacteria to persist and establish the infection, avoiding the hostile environment of the macrophage.…”

Section: Introductionmentioning

confidence: 99%

Mycobacterium smegmatis laminin‐binding glycoprotein shares epitopes with Mycobacterium tuberculosis heparin‐binding haemagglutinin

Pethe

Puech

Daffé

et al. 2001

Molecular Microbiology

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SummaryMycobacterium tuberculosis, the causative agent of tuberculosis, produces a heparin-binding haemagglutinin adhesin (HBHA), which is involved in its epithelial adherence. To ascertain whether HBHA is also present in fast-growing mycobacteria, Mycobacterium smegmatis was studied using anti-HBHA monoclonal antibodies (mAbs). A cross-reactive protein was detected by immunoblotting of M. smegmatis whole-cell lysates. However, the M. tuberculosis HBHA-encoding gene failed to hybridize with M. smegmatis chromosomal DNA in Southern blot analyses. The M. smegmatis protein recognized by the anti-HBHA mAbs was purified by heparin±Sepharose chromatography, and its amino-terminal sequence was found to be identical to that of the previously described histone-like protein, indicating that M. smegmatis does not produce HBHA. Biochemical analysis of the M. smegmatis histone-like protein shows that it is glycosylated like HBHA. Immunoelectron microscopy demonstrated that the M. smegmatis protein is present on the mycobacterial surface, a cellular localization inconsistent with a histone-like function, but compatible with an adhesin activity. In vitro protein interaction assays showed that this glycoprotein binds to laminin, a major component of basement membranes. Therefore, the protein was called M. smegmatis laminin-binding protein (MS-LBP). MS-LBP does not appear to be involved in adherence in the absence of laminin but is responsible for the laminin-mediated mycobacterial adherence to human pneumocytes and macrophages. Homologous laminin-binding adhesins are also produced by virulent mycobacteria such as M. tuberculosis and Mycobacterium leprae, suggesting that this adherence mechanism may contribute to the pathogenesis of mycobacterial diseases.

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“…The mechanisms by which M. avium interacts with the epithelial mucosa are complex. Our, as well as other, laboratories have shown that M. avium binds to and invades intestinal epithelial cells such as HT‐29 and Caco‐2 cells ( Manpother and Sanger, 1984; Bermudez and Young, 1994) as well as laryngeal epithelial cells (HEp‐2 cells) in vitro . Subsequent to oral ingestion, M. avium has been shown to colonize preferentially the terminal ileum and the ascending colon in a mouse model of infection ( Bermudez et al ., 1992 ).…”

Section: Introductionmentioning

confidence: 99%

Mycobacterium avium enters intestinal epithelial cells through the apical membrane, but not by the basolateral surface, activates small GTPase Rho and, once within epithelial cells, expresses an invasive phenotype

2000

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SummaryMycobacterium avium is a common pathogen in AIDS patients that is primarily (but not exclusively) acquired through the gastrointestinal tract, leading to the development of bacteraemia and disseminated disease. To cause infection through the gut, binding and invasion of the intestinal epithelial barrier are required. To characterize this process further, we determined the cell surface(s) (basolateral vs. apical membrane) that M. avium interacts with in intestinal mucosal cells in vitro. The level of binding and invasion of both HT-29 and Caco-2 intestinal cell monolayers by M. avium were similar when the assay was performed with control medium in the presence of Ca 21 (when only the apical surface was exposed), with Ca 21 -depleted medium or with Ca 21 -depleted medium 1 1 mM EGTA (exposure of both apical and basolateral membranes), suggesting that the bacterium enters the apical surface of the epithelial lining. These observations were confirmed by assays in a transwell system and by using fluorescent microscopy. Real-time video microscopy showed that M. avium entry was not associated with membrane ruffling and the use of pharmacological inhibitors of the small GTPases demonstrated that M. avium invasion is dependent on the activation of the small GTPases Rho, but not on Rac or Cdc42. Passage of M. avium through HT-29 cells led to a phenotypic change (intracellular growth; IG) that was associated with a significantly greater (between five-and ninefold) ability to bind to and invade new monolayers of epithelial cells or macrophages when compared with the invasion by M. avium grown on agar (extracellular growth; EG). IG phenotype invasion of HT-29 cells also takes place only by the apical surface. M. avium enters intestinal epithelial cells by the apical surface and, once within the cells, changes phenotype, becoming more invasive towards both macrophages and other epithelial cells.

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In vitro interaction of Mycobacterium avium with intestinal epithelial cells

Cited by 54 publications

References 21 publications

Mycobacterium avium-M. intracellulare binds to the integrin receptor alpha v beta 3 on human monocytes and monocyte-derived macrophages

Mycobacterium avium-M. intracellulare binds to the integrin receptor alpha v beta 3 on human monocytes and monocyte-derived macrophages

Mycobacterium smegmatis laminin‐binding glycoprotein shares epitopes with Mycobacterium tuberculosis heparin‐binding haemagglutinin

Mycobacterium avium enters intestinal epithelial cells through the apical membrane, but not by the basolateral surface, activates small GTPase Rho and, once within epithelial cells, expresses an invasive phenotype

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