1 Division of Gastrointestinal Pathology, Department of Pathology and Laboratory Medicine, Emory University, Atlanta, Georgia 30322; and 2 Department of Anesthesiology, Brigham and Women's Hospital, Boston, Massachusetts 02115
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ABSTRACT |
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Neutrophil (PMN) transepithelial migration is a major effector of epithelial defense in inflammatory diseases involving mucosal surfaces. However, major receptor-ligand interactions between epithelial cells and PMN remain incompletely characterized. To better define the molecular events involved in PMN interactions with epithelial cells, we produced a monoclonal antibody called g82 that inhibited PMN transepithelial migration in the physiological basolateral-to-apical direction. The g82 antigen localized to the apical surface of human colonic epithelium and was significantly upregulated under inflammatory conditions. Immunoprecipitation revealed two polypeptides of Mr 207 and 32 kDa. F(ab')2 fragments from g82 IgG had no effect on transmigration, suggesting Fc dependence. Further experiments confirmed dependence on the PMN Fc receptor CD32A and that the observed effects were secondary to a failure of PMN to detach from the apical epithelial surface. These Fc-mediated events were epitope specific since binding, isotype-matched antibodies did not affect detachment. These results identify a new mechanism for retention of PMN at the apical epithelial surface following transepithelial migration. This pathway may be important in pathogen clearance and mucosal pathophysiology associated with autoimmunity.
intestine; inflammation; ulcerative colitis; epithelial cells
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INTRODUCTION |
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POLYMORPHONUCLEAR LEUKOCYTE (PMN) migration across columnar epithelia is an important component of mucosal inflammation (22, 31, 46). When PMN reach their target, there is a potent respiratory burst, enzyme release, and phagocytosis that destroys invading pathogens (4, 13, 18) and may lead to tissue injury. Although PMN migration across the vascular endothelium has been extensively characterized on a molecular level (20), much less is known about the sequence of events that regulate PMN-epithelial interactions (34).
Previous studies have shown that PMN transmigration across intestinal
(2, 38), lung (43), and urinary tract
(1) epithelia is dependent on the leukocyte-specific
2-integrin CD11b/CD18. Although this CD11b/CD18-mediated
interaction appears to represent an initial adhesive event in
PMN-epithelial interactions (36), the epithelial ligand
pair for this integrin remains undefined and does not appear to be
intracellular cell adhesion molecule-1 (7, 36). At a step
following initial
2-integrin-dependent adhesion, PMN
migration into the intraepithelial space is regulated by the membrane
glycoprotein CD47 (9, 37). Although CD47 has been studied
in a variety of cell systems (6, 48), the mechanism by
which CD47 regulates PMN migration remains obscure.
The apical epithelial surface is a key site for host-pathogen interactions and represents the final destination of migrating leukocytes before they enter the intestinal lumen (27, 49). During active intestinal inflammation, when migrating PMN are retained in close proximity to the apical epithelial surface, crypt abscesses are formed. There is also release of inflammatory mediators that results in alterations in intestinal permeability and epithelial injury (27, 30, 32). Thus aberrant triggering of accumulation of PMN within intestinal crypts could result in further injury and may participate in disease flares characteristic of ulcerative colitis and Crohn's disease (5, 24, 39).
Using a monoclonal antibody (MAb) approach to study epithelial-leukocyte interactions, we report the identification and characterization of an IgG1 MAb (g82) that, following N-formylpeptide (fMLP)-stimulated PMN migration across polarized monolayers of T84 epithelial cells, inhibits detachment/release of PMN into the lumen. The antigen consists of one or two proteins with relative molecular mass (Mr) of 207 and 32 kDa, is expressed on the apical epithelial surface, and is upregulated under inflammatory conditions. Here we show that the inhibitory effect of g82 IgG on basolateral-to-apical PMN transepithelial migration results from the failure of PMN to detach from the apical epithelial surface. This inhibition is Fc (CD32A)-dependent and unique to the epitope defined by g82, since antibodies reactive with other apically expressed antigens have no effect on PMN transmigration. Our results indicate that transepithelial migration of PMN is not complete after crossing apical intercellular tight junctions but also requires release from the apical domain into the lumen. Control of the latter step might influence critical PMN effector mechanisms, such as clearance of bound pathogens from the apical surface, and mucosal injury in diseases associated with autoimmunity, such as ulcerative colitis (10, 15, 17).
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MATERIALS AND METHODS |
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Cell culture.
T84 intestinal epithelial cells (11) (passages
55-80) were grown in a 1:1 mixture of DMEM and Ham's F-12
medium supplemented with 15 mM of HEPES buffer (pH 7.5), 14 mM
NaHCO3, 8 µg/ml ampicillin, 90 µg/ml streptomycin, 40 µg/ml penicillin, and 5% newborn calf serum as previously described
(37). For apical-to-basolateral and basolateral-to-apical
transmigration experiments, T84 cells were grown on collagen-coated,
permeable polycarbonate filters (5 µm pore size) with a surface area
of 0.33 cm2 (Costar, Cambridge, MA) in both standard and
inverted configurations as previously described (2, 37,
38). For ELISA experiments, T84 epithelial cells were cultured
to confluence (7 days) in 96-well tissue culture plates. In some
experiments, cell monolayers were exposed to recombinant human
interferon (IFN)- (106 U/ml; kind gift of Dr.
Randall Mrsny, Genentech, San Francisco, CA) for 48 h before use
as previously described (7).
Buffers.
Hanks' balanced salt solution (HBSS) contains (in g/l) 0.185 CaCl2, 0.098 MgSO4, 0.4 KCl, 0.06 KH2PO4, 8.0 NaCl, 0.048 Na2PO4, 1 glucose, and 10 mM HEPES, pH 7.4. Modified HBSS() was prepared as above, except without
CaCl2 and MgSO4. Immunoprecipitation wash
buffer consisted of 400 mM NaCl, 100 mM NaF, 1 mM EDTA, 1% Triton
X-100, and 10 mM Na2HPO4, pH 7.4. For
immunoprecipitation experiments, cell lysis buffer contained 100 mM
KCl, 30 mM NaCl, 2 mM EDTA, 10 mM HEPES, pH 7.4, and 2% Triton X-100.
PMN isolation.
PMN were isolated from whole blood (anticoagulated with
citrate/dextrose) obtained from normal human volunteers by using a previously described gelatin sedimentation procedure (16).
PMN were suspended in modified HBSS() at a concentration of 5 × 107 cells/ml (4°C) and used in transmigration experiments.
Antibodies. T84 cell membranes (21) were used to produce monoclonal antibodies and were screened for inhibition of PMN transepithelial migration as previously described (37). Briefly, female BALB/c mice were immunized by intraperitoneal injection of 1 × 107 cell equivalents emulsified with complete Freud's adjuvant (GIBCO BRL) followed by two subsequent injections of 107 cell equivalents emulsified in incomplete Freud's adjuvant over a 6-wk period. Mice with high titers of antibodies recognizing epithelial surfaces were given a final intravenous immunization by tail vein with 3 × 106 cell equivalents of T84 cell membranes in HBSS. Splenocytes were subsequently harvested (4 days later) and fused with P3U1 myeloma cells. Ten days after fusion, hybridoma supernatants were collected and screened for reactivity with T84 cells compared with a nonepithelial cell type. Hybridoma supernatants demonstrating preferential epithelial reactivity were then screened for inhibition of PMN transepithelial migration. Hybridomas were then subcloned twice by limiting dilution and weaned from selection media, and antibodies were purified by using protein A-Sepharose (Sigma, St. Louis, MO) followed by dialysis against 150 mM NaCl and 10 mM HEPES buffer, pH 7.4. F(ab')2 was commercially prepared (Lampire Biological Laboratories, Coopersburg, PA) by pepsin digestion (100 U/mg, 6 h, 37°C) (33). Purity of antibody digest was confirmed by SDS-PAGE. F(ab') fragments of inhibitory anti-CD32A IgG (26) and anti-CD16B IgG (42) were also commercially prepared by papain digestion in an analogous fashion (Lampire Biological), and binding activity was confirmed by flow cytometry. Control antibodies used in transmigration assays included MAb 44a (inhibitory anti-CD11b), W6/32 (MHC class I, noninhibitory IgG2; antigen is expressed on the apical and basolateral surface of T84 cells), and CLT/479 (IgG1; antigen is expressed on the apical surface of T84 cells) (3, 35, 38, 40, 47).
PMN transmigration and cell adhesion assays. PMN transepithelial migration experiments were performed in both the apical-to-basolateral and basolateral-to-apical directions with the use of T84 monolayers (34, 38). PMN adhesion to confluent T84 monolayers was also assessed as previously described (35).
Quantitation of PMN adherent to monolayers after basolateral-to-apical transmigration. To quantify PMN associated with epithelial monolayers in basolateral-to-apical transmigration assays, a modified monolayer-washing procedure was developed. After completion of transmigration assays (110 min), T84 monolayers were gently removed from the original reservoirs of 24-well tissue culture plates and transferred to new 24-well tissue culture plates containing 1 ml of HBSS in each well. With the use of a rotor adapted for tissue culture plates (Beckman), the transferred, inverted monolayers were subjected to low-speed centrifugation for 5 min (50 g, 4°C). PMN that detached from the epithelial surface of T84 cell monolayers were then quantified by myeloperoxidase (MPO) assay (38) and reported as adherent PMN. PMN remaining within the filter/monolayer and in the original transmigration chambers were also quantified by MPO assay after nonmigrated PMN were rinsed away and were reported as filter-associated and migrated PMN, respectively.
Immunofluorescence. T84 monolayers were fixed in 3.7% paraformaldehyde in HBSS (10 min, 20°C), washed and incubated in HBSS containing 5% normal goat serum, followed by primary antibody for 2 h (10 µg/ml in 5% normal goat serum) as previously described (36). Labeling was also performed on 4-µm frozen tissue sections of human colonic mucosa, both normal and with active ulcerative colitis, obtained from fresh surgical resection specimens. Tissue sections were mounted on glass coverslips, air dried, and fixed in 3.7% paraformaldehyde. After being washed, monolayers and tissue sections were incubated with FITC-conjugated 2° antibody (Cappel, Durham, NC) for 1 h at 20°C. To visualize nuclei, both monolayers and tissue sections were labeled with propidium iodide (1:1,000 dilution, 20 min, 20°C). Additional control stains for basolateral membrane/tight junction [junction adhesion molecule (JAM); MAb J10.4] (25) and nonspecific murine IgG1 were performed in parallel.
Immunoprecipitation. Immunoprecipitation experiments were performed with the use of biotin surface-labeled T84 cell monolayers as previously described (36, 37). Briefly, monolayers were surface labeled with 1 mM sulfo-NHS-biotin (Pierce Chemical) in HBSS for 20 min (4°C) followed by quenching with 150 mM NH4Cl (36). Monolayers were solubilized in cell lysis buffer containing protease inhibitors followed by centrifugation. The supernatants were then precleared for 2 h with normal mouse IgG-Sepharose [IgG coupled to cyanogen bromide-activated Sepharose at a protein-Sepharose density of 3 mg/ml according to the manufacturer's instructions (Pharmacia, Uppsala, Sweden)]. This was followed by incubation for 2 h (4°C) with 30 µl g82-Sepharose. Immunoprecipitates were then washed with 1% octylglucoside-100 mM sodium phosphate, pH 7.4, denatured by boiling in sample buffer, and subjected to SDS-PAGE followed by Western blot. Nitrocellulose blots were probed with avidin peroxidase and visualized by enhanced chemiluminescence (Amersham, Piscataway, NJ).
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RESULTS |
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Antibody production and screening. A fusion with splenocytes from mice immunized with T84 cell membranes yielded 915 antibody-producing clones that were screened for binding to T84 epithelial cells and to PMN. By using this approach, 132 clones were identified that preferentially labeled the surface of T84 cells by ELISA. Transepithelial migration experiments were then performed by pooling supernatants in groups of three and adding 50 µl to the upper chamber(s) of the Transwell device as previously described (37). Of the pooled supernatants that initially inhibited migration, one gave consistent inhibition in the screening assays and was subcloned to produce MAb g82 (IgG1). In initial transmigration assays with g82 hybridoma supernatants, antibody was added to the upper chamber (apical surface) in apical-to-basolateral transmigration assays and ~50% inhibition of migration was observed (data not shown). However, the addition of the antibody to the upper chamber (basolateral surface) in basolateral-to-apical transmigration assays resulted in no significant inhibition of migration (data not shown) for reasons that are shown in Fig. 5.
Binding of the antibody to the surface of T84 cell monolayers could be readily detected by ELISA (Fig. 1), and such binding was increased approximately twofold after treatment of T84 cells with IFN-
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g82 IgG immunoprecipitates two proteins from surface-labeled T84
cells.
Labeling and immunoprecipitation experiments were performed using g82
IgG. T84 cell monolayers were surface labeled with biotin, detergent
solubilized, and immunoprecipitated with g82 IgG as detailed in
MATERIALS AND METHODS. Western blots were then analyzed by
gel scanning. Compared with protein standard controls, two proteins of
~207 and ~32 kDa Mr (Fig.
2) were immunoprecipitated by g82
IgG under both reducing and nonreducing conditions.
Immunoprecipitation experiments with IFN--stimulated T84 cells
produced two proteins with identical Mr values
to those obtained from nonstimulated epithelial cells (data not shown).
Western blotting of T84 cell extracts or immunoprecipitates with g82
IgG were unsuccessful. Thus it is not clear which of the two proteins,
or both, represents the antigen.
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The antigen for g82 is expressed on the apical surface of T84 cells
and human colonic epithelium.
To localize the epitope defined g82 IgG, immunofluorescence labeling
was performed using confluent monolayers of T84 cells cultured on
permeable supports. Fig. 3, A
and B, represent computer-reconstructed confocal micrographs
of g82 IgG staining in the x-z plane. Nuclear staining is
shown in red. An intense linear staining pattern consistent with apical
surface expression of the g82 antigen is observed, which is
significantly increased after IFN- treatment (Fig. 3B). For comparison, staining for the tight junction/basolateral membrane protein JAM (25) is shown in Fig. 3C.
Background fluorescence from nonspecific isotype-matched IgG was not
significant (not shown).
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Inhibition of detachment of PMN from the apical surface of
epithelia is specific for the g82 antigen and mediated by Fc receptors.
We surmised that the results in Fig. 5 could be explained by PMN Fc
receptors interacting with g82 IgG bound to the apical surface. To test
this hypothesis, the Fc portion of g82 IgG was removed to form
F(ab')2 fragments, which were then used in transmigration assays. F(ab')2 fragments produced from g82 IgG showed
similar binding characteristics to T84 cells as intact IgG, thus
indicating preservation of specific binding affinity (data not shown).
As indicated in Fig. 6A, the
data are normalized with respect to migration in the absence of
antibody. Although apical surface detachment was inhibited 93.8% by
intact g82 IgG, there was no significant inhibition of surface
detachment by the g82 F(ab')2.
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Effects of g82 IgG on PMN adhesion to T84 monolayers.
The results of the above migration experiments suggested that
Fc-mediated PMN adhesion to the apical surface of T84 monolayers could
have significant effects on terminal events in the PMN transepithelial migration cascade. Therefore, we examined the ability of g82 IgG to
promote adhesion of PMN when directly applied to the apical surface of
confluent T84 monolayers. As shown in Fig. 6B, PMN adhesion
to monolayers pretreated with binding-control antibodies was not
significantly increased compared with binding in the absence of applied
antibody. In contrast, PMN adhesion to T84 monolayers pretreated with
g82 IgG was increased nearly threefold. Furthermore, when T84
monolayers were stimulated with IFN-, there was an additional twofold increase in adhesion observed with g82 IgG compared with controls (data not shown). These findings parallel previous
observations of generally increased PMN adhesion to T84 monolayers
after stimulation with IFN-
(8). Lastly, as can be seen
in Fig. 6B, the g82 IgG-induced increase in adhesion was
reduced to control levels by pretreatment of PMN with anti-CD32A F(ab')
but not by anti-CD16B F(ab').
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DISCUSSION |
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In this report, we demonstrate that Fc-mediated adhesive interactions can modulate the PMN transepithelial migration response to the chemotactic peptide fMLP. In particular, we show that g82 IgG specifically inhibits PMN detachment following basolateral-to-apical transepithelial migration and is dependent on CD32A, one of the low-affinity Fc receptors on PMN (13, 18). Since this Fc-mediated effect appears to be epitope specific, it suggests that geographical constraints (antigen clustering, compartmentalization, etc.) and/or signaling events at the level of the apical membrane might influence PMN detachment from the epithelial surface after transmigration. In addition, signaling events from such contact could result in altered epithelial functions such as permeability that, in turn, may modulate subsequent PMN migration.
Although we were not able to demonstrate direct adhesive interactions between the g82 antigen and PMN, it has been suggested that induction of other apical ligands for PMN such as intracellular cell adhesion molecule-1 might function to promote clearance of apically attached bacteria (19, 36). If this is true, then one could envision similar Fc-mediated events to those described here. In particular, since the apical epithelial surface is the initial site of pathogen adherence (14, 28, 29, 49), opsonization followed by antibody-mediated adhesion/phagocytosis would facilitate clearance of offending microorganisms. Indeed, a characteristic histological feature of infectious colitis is the presence of abundant crypt abscesses, collections of PMN within colonic crypts (22, 23), in close proximity to the apical epithelial surface.
IgG-mediated PMN-epithelial adhesive interactions are also likely to play a role in the pathophysiology of mucosal diseases associated with autoantibodies such as ulcerative colitis. In the active phase of this disease, when there are abundant crypt abscesses, patient symptoms are most pronounced. Similarly, studies have shown that ulcerative colitis is associated with IgG autoantibodies directed against intestinal epithelial cells in a majority of patients (10, 15, 17). In one of these studies, abundant IgG1 binding to the apical intestinal epithelial surface was found in 7 of 11 patients with active ulcerative colitis (15). Thus, although we do not have evidence for a direct association between antibodies against the antigen for g82 and human disease, our findings provide a mechanism for PMN retention in colonic crypts under the above conditions.
The tissue/epithelial distribution studies in this report provide
additional evidence supporting the physiological relevance of our
findings. First, the antigen for g82 is similarly expressed at the
apical surface in both T84 cells and natural human colonic epithelium.
Secondly, such expression is significantly upregulated in T84
monolayers after stimulation with the inflammatory cytokine IFN-. In
parallel, the antigen is similarly upregulated in colonic epithelium
from patients with active ulcerative colitis. Thus, under inflammatory
conditions associated with enhanced PMN-epithelial interactions and
crypt abscess formation, there is appropriate upregulation of a crypt
surface antigen that supports Fc-mediated PMN adhesion.
Although we have illustrated the potential importance of IgG antibodies against specific antigens in the gastrointestinal tract during disease, how would IgG gain access to such luminal antigens? Although IgA is the predominant secreted immunoglobulin, recent studies by Dickinson et al. (12) have demonstrated that IgG antibodies are actively transported across epithelial barriers in the basolateral-to-apical direction. IgG movement into the lumen would be further enhanced under conditions associated with disruption of epithelial barrier function. This is indeed the case for ulcerative colitis, which is characterized by widespread mucosal ulceration leading to abundant seepage of serum components into the intestinal lumen (22, 23, 41). These observations provide a mechanism by which IgG can gain access to apically restricted antigens that is enhanced under specific inflammatory conditions.
As depicted in Fig. 7, the results from
the current study can be incorporated into a multistep model of PMN
transepithelial migration and crypt abscess formation. In this model,
extravasated PMN migrate to the basolateral aspect of the crypt
epithelium, where initial contact is dependent on the PMN
2-integrin CD11b/CD18 (2, 38).
Subsequently, PMN migrate across the epithelium by CD47-dependent
processes (37) to reach the apical aspect (lumen) of the
epithelium. Here, in the presence of apically bound IgG, PMN are
retained in close proximity to the epithelium, contributing to crypt
abscess formation. Close apposition of PMN to apical epithelial
membranes would facilitate the action of paracrine factors released
from PMN on epithelial function such as cAMP-mediated chloride
secretion (27, 44, 45). The above scenario thus provides a
mechanism for secretory diarrhea that is commonly observed in
individuals with abundant crypt abscesses.
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Results of our immunoprecipitation experiments indicate that g82 IgG recognizes at least one of two proteins with Mr values of 207 and 32 kDa. However, since the antibody does not Western blot, it remains unclear which protein harbors the recognized epitope. Further characterization of the epitope awaits purification/sequencing and/or molecular cloning of the proteins recognized.
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ACKNOWLEDGEMENTS |
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We thank Denice Esterly for expert tissue culture and technical assistance.
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FOOTNOTES |
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This research was supported by Grants HL-60540, HL-54229, HL-60569, DK-50189, DK-47662, DK-35932, and DK-57827 from the National Institutes of Health and a Biomedical Sciences Grant from the Arthritis Foundation.
Address for reprint requests and other correspondence: T. A. Reaves, Dept. of Pathology and Laboratory Medicine, Emory Univ., 1639 Pierce Drive, Atlanta, GA 30322 (E-mail: treaves{at}emory.edu).
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Received 26 May 2000; accepted in final form 14 November 2000.
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REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Agace, WW.
The role of the epithelial cell in Escherichia coli induced neutrophil migration into the urinary tract.
Eur Respir J
9:
1713-1728,
1996
2.
Balsam, LB,
Liang TW,
and
Parkos CA.
Functional mapping of CD11b/CD18 epitopes important in neutrophil-epithelial interactions: a central role of the I domain.
J Immunol
160:
5058-5065,
1998
3.
Barnstable, CJ,
Bodmer WF,
Brown G,
Galfre G,
Milstein C,
Williams AF,
and
Ziegler A.
Production of monoclonal antibodies to group A erythrocytes, HLA and other human cell surface antigensnew tools for genetic analysis.
Cell
14:
9-20,
1978[ISI][Medline].
4.
Borregaard, N,
and
Cowland JB.
Granules of the human neutrophilic polymorphonuclear leukocyte.
Blood
89:
3503-3521,
1997
5.
Brown, MO.
Inflammatory bowel disease.
Prim Care
26:
141-170,
1999[ISI][Medline].
6.
Chen, D,
Guo K,
Yang J,
Frazier WA,
Isner JM,
and
Andres V.
Vascular smooth muscle cell growth arrest on blockade of thrombospondin-1 requires p21(Cip1/WAF1).
Am J Physiol Heart Circ Physiol
277:
H1100-H1106,
1999
7.
Colgan, SP,
Parkos CA,
Delp C,
Arnaout MA,
and
Madara JL.
Neutrophil migration across cultured intestinal epithelial monolayers is modulated by epithelial exposure to IFN-gamma in a highly polarized fashion.
J Cell Biol
120:
785-798,
1993[Abstract].
8.
Colgan, SP,
Parkos CA,
Matthews JB,
Awtrey DALCS,
Lichtman AH,
Delp-Archer C,
and
Madara JL.
Interferon-gamma induces a cell surface phenotype switch on T84 intestinal epithelial cells.
Am J Physiol Cell Physiol
267:
C402-C410,
1994
9.
Cooper, D,
Lindberg FP,
Gamble JR,
Brown EJ,
and
Vadas MA.
Transendothelial migration of neutrophils involves integrin-associated protein (CD47).
Proc Natl Acad Sci USA
92:
3978-3982,
1995
10.
Das, KM,
Sakamaki S,
Vecchi M,
and
Diamond B.
The production and characterization of monoclonal antibodies to a human colonic antigen associated with ulcerative colitis: cellular localization of the antigen by using the monoclonal antibody.
J Immunol
139:
77-84,
1987
11.
Dharmsathaphorn, K,
and
Madara JL.
Established intestinal cell lines as model systems for electrolyte transport studies.
Methods Enzymol
192:
354-389,
1990[Medline].
12.
Dickinson, BL,
Badizadegan K,
Wu Z,
Ahouse JC,
Zhu X,
Simister NE,
Blumberg RS,
and
Lencer WI.
Bidirectional FcRn-dependent IgG transport in a polarized human intestinal epithelial cell line.
J Clin Invest
104:
1-10,
1999
13.
Edberg, JC,
Moon JJ,
Chang DJ,
and
Kimberly RP.
Differential regulation of human neutrophil FcgammaRIIa (CD32) and FcgammaRIIIb (CD16)-induced Ca2+ transients.
J Biol Chem
273:
8071-8079,
1998
14.
Gewirtz, AT,
Siber AM,
Madara JL,
and
McCormick BA.
Orchestration of neutrophil movement by intestinal epithelial cells in response to Salmonella typhimurium can be uncoupled from bacterial internalization.
Infect Immun
67:
608-617,
1999
15.
Halstensen, TS,
Mollnes TE,
Garred P,
Fausa O,
and
Brandtzaeg P.
Epithelial deposition of immunoglobulin G1 and activated complement (C3b and terminal complement complex) in ulcerative colitis.
Gastroenterology
98:
1264-1271,
1990[ISI][Medline].
16.
Henson, PM,
and
Oades ZG.
Stimulation of human neutrophils by soluble and insoluble immunoglobulin aggregates. Secretion of granule constituents and increased oxidation of glucose.
J Clin Invest
56:
1053-1061,
1975[ISI][Medline].
17.
Hibi, T,
Aiso S,
Ishikawa M,
Watanabe M,
Yoshida T,
Kobayashi K,
Asakura H,
Tsuru S,
and
Tsuchiya M.
Circulating antibodies to the surface antigens on colon epithelial cells in ulcerative colitis.
Clin Exp Immunol
54:
163-168,
1983[ISI][Medline].
18.
Hoffmeyer, F,
Witte K,
and
Schmidt RE.
The high-affinity Fc gamma RI on PMN: regulation of expression and signal transduction.
Immunology
92:
544-552,
1997[ISI][Medline].
19.
Huang, GT,
Eckmann L,
Savidge TC,
and
Kagnoff MF.
Infection of human intestinal epithelial cells with invasive bacteria upregulates apical intercellular adhesion molecule-1 (ICAM-1) expression and neutrophil adhesion.
J Clin Invest
98:
572-583,
1996
20.
Imhof, BA,
and
Dunon D.
Basic mechanism of leukocyte migration.
Horm Metab Res
29:
614-621,
1997[ISI][Medline].
21.
Kaoutzani, P,
Parkos CA,
Delp-Archer C,
and
Madara JL.
Isolation of plasma membrane fractions from the intestinal epithelial model T84.
Am J Physiol Cell Physiol
264:
C1327-C1335,
1993
22.
Kumar, NB,
Nostrant TT,
and
Appelman HD.
The histopathologic spectrum of acute self-limited colitis (acute infectious-type colitis).
Am J Surg Pathol
6:
523-529,
1982[ISI][Medline].
23.
Lamm, ME.
Interaction of antigens and antibodies at mucosal surfaces.
Annu Rev Microbiol
51:
311-340,
1997[ISI][Medline].
24.
Lewin, KJ.
Inflammatory Bowel Disease. New York: IGaku-Shoin Medical, 1992.
25.
Liu, Y,
Nusrat A,
Schnell FJ,
Reaves TA,
Walsh S,
Pochet M,
and
Parkos CA.
Human junction adhesion molecule regulates tight junction resealing in epithelia.
J Cell Sci
113:
2363-2374,
2000
26.
Looney, RJ,
Abraham GN,
and
Anderson CL.
Human monocytes and U937 cells bear two distinct Fc receptors for IgG.
J Immunol
136:
1641-1647,
1986
27.
Madara, JL,
Patapoff TW,
Gillece-Castro B,
Colgan SP,
Parkos CA,
Delp C,
and
Mrsny RJ.
5'-adenosine monophosphate is the neutrophil-derived paracrine factor that elicits chloride secretion from T84 intestinal epithelial cell monolayers.
J Clin Invest
91:
2320-2325,
1993[ISI][Medline].
28.
McCormick, B,
Gewirtz A,
and
Madara JL.
Epithelial cross talk with bacteria and immune cells.
Curr Opin Gastroenterol
14:
492-497,
1998[ISI].
29.
McCormick, BA,
Parkos CA,
Colgan SP,
Carnes DK,
and
Madara JL.
Apical secretion of a pathogen-elicited epithelial chemoattractant activity in response to surface colonization of intestinal epithelia by Salmonella typhimurium.
J Immunol
160:
455-466,
1998
30.
Nash, S,
Parkos C,
Nusrat A,
Delp C,
and
Madara JL.
In vitro model of intestinal crypt abscess. A novel neutrophil-derived secretagogue activity.
J Clin Invest
87:
1474-1477,
1991[ISI][Medline].
31.
Nostrant, TT,
Kumar NB,
and
Appelman HD.
Histopathology differentiates acute self-limited colitis from ulcerative colitis.
Gastroenterology
92:
318-328,
1987[ISI][Medline].
32.
Nusrat, A,
Parkos CA,
Liang TW,
Carnes DK,
and
Madara JL.
Neutrophil migration across model intestinal epithelia: monolayer disruption and subsequent events in epithelial repair.
Gastroenterology
113:
1489-1500,
1997[ISI][Medline].
33.
Parham, P.
Preparation and purification of active fragments from mouse monoclonal antibodies.
In: Immunological Methods in Biomedical Sciences, edited by Weir CBDM,
and Herznberg L.. Oxford, UK: Blackwell, 1988, p. 14.1-14.2.
34.
Parkos, CA.
Molecular events in neutrophil transepithelial migration.
Bioessays
19:
865-873,
1997[ISI][Medline].
35.
Parkos, CA,
Colgan SP,
Bacarra AE,
Nusrat A,
Delp-Archer C,
Carlson S,
Su DH,
and
Madara JL.
Intestinal epithelia (T84) possess basolateral ligands for CD11b/CD18-mediated neutrophil adherence.
Am J Physiol Cell Physiol
268:
C472-C479,
1995
36.
Parkos, CA,
Colgan SP,
Diamond MS,
Nusrat A,
Liang TW,
Springer TA,
and
Madara JL.
Expression and polarization of intercellular adhesion molecule-1 on human intestinal epithelia: consequences for CD11b/CD18-mediated interactions with neutrophils.
Mol Med
2:
489-505,
1996[ISI][Medline].
37.
Parkos, CA,
Colgan SP,
Liang TW,
Nusrat A,
Bacarra AE,
Carnes DK,
and
Madara JL.
CD47 mediates postadhesive events required for neutrophil migration across polarized intestinal epithelia.
J Cell Biol
132:
437-450,
1996[Abstract].
38.
Parkos, CA,
Delp C,
Arnaout MA,
and
Madara JL.
Neutrophil migration across a cultured intestinal epithelium. Dependence on a CD11b/CD18-mediated event and enhanced efficiency in physiological direction.
J Clin Invest
88:
1605-1612,
1991[ISI][Medline].
39.
Robinson, M.
Medical therapy of inflammatory bowel disease for the 21st century.
Eur J Surg Suppl
582:
90-98,
1998[Medline].
40.
Sakamoto, JC-CC,
Friedman E,
Finstad CL,
Enker WE,
Melamed MR,
Lloyd KO,
Oettgen HF,
and
Old LJ.
Antigens of normal and neoplastic human colonic mucosa cells (HCMC) defined by monoclonal antibodies (mAB) (Abstract).
Proc Am Assoc Cancer Res
24:
225,
1983[ISI].
41.
Scotiniotis, I,
Rubesin SE,
and
Ginsberg GG.
Imaging modalities in inflammatory bowel disease.
Gastroenterol Clin North Am
28:
391-421,
1999[ISI][Medline].
42.
Selvaraj, P,
Rosse WF,
Silber R,
and
Springer TA.
The major Fc receptor in blood has a phosphatidylinositol anchor and is deficient in paroxysmal nocturnal haemoglobinuria.
Nature
333:
565-567,
1988[ISI][Medline].
43.
Stockley, RA.
Role of inflammation in respiratory tract infections.
Am J Med
99:
8S-13S,
1995[Medline].
44.
Strohmeier, GR,
Lencer WI,
Patapoff TW,
Thompson LF,
Carlson SL,
Moe SJ,
Carnes DK,
Mrsny RJ,
and
Madara JL.
Surface expression, polarization, and functional significance of CD73 in human intestinal epithelia.
J Clin Invest
99:
2588-2601,
1997
45.
Strohmeier, GR,
Reppert SM,
Lencer WI,
and
Madara JL.
The A2b adenosine receptor mediates cAMP responses to adenosine receptor agonists in human intestinal epithelia.
J Biol Chem
270:
2387-2394,
1995
46.
Surawicz, CM,
Haggitt RC,
Husseman M,
and
McFarland LV.
Mucosal biopsy diagnosis of colitis: acute self-limited colitis and idiopathic inflammatory bowel disease.
Gastroenterology
107:
755-763,
1994[ISI][Medline].
47.
Todd, RF, III,
Nadler LM,
and
Schlossman SF.
Antigens on human monocytes identified by monoclonal antibodies.
J Immunol
126:
1435-1442,
1981
48.
Wang, XQ,
and
Frazier WA.
The thrombospondin receptor CD47 (IAP) modulates and associates with alpha2 beta1 integrin in vascular smooth muscle cells.
Mol Biol Cell
9:
865-874,
1998
49.
Wick, MJ,
Madara JL,
Fields BN,
and
Normark SJ.
Molecular cross talk between epithelial cells and pathogenic microorganisms.
Cell
67:
651-659,
1991[ISI][Medline].