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A revived interest in intraepithelial lymphocytes (IEL) has been elicited by several recent reports suggesting that murine and avian intestinal epithelium contains mainly CD3+ CD8+ cells expressing the γ/δ T-cell receptor (TcR) for antigen; this contrasts with systemically distributed T cells which preferentially employ the TcRα/β. An anatomical dichotomy in the distribution of these two T-cell lineages has hence been proposed Here we report that this concept does not hold true in man. In situ studies with monoclonal TcR-framework antibodies showed that most (70-90%) human intestinal IEL (which are mainly CD3+CD8) expressed TcRα/β, Moreover, almost half of the intraepithelial CD3+ cells were positive for the smallest (180 kDa) CD45 molecule (UCHLl); this probably reflected that they are antigen-primed and thus represent traditional CD3+ CD8+ α/β+ memory T cells. [ABSTRACT FROM AUTHOR]

Extensively washed, ethanol fixed and paraffin embedded colonic specimens from 15 patients with ulcerative colitis (UC) and nine patients with Crohn's disease (CD) of the colon, ileal specimens from six patients with CD of the ileum, and histologically normal control specimens obtained from 10 patients operated for colonic carcinoma, were examined by immunohistochemistry with a monoclonal antibody specific for a neoepitope in the C9 part of the terminal complement complex (TCC). The submucosal blood vessels in inflammatory bowel disease (IBD) showed significantly more TCC positivity than the controls, and vascular TCC deposition was statistically related (p less than 0.001) to degree of inflammation. Five of the six ileal CD specimens contained likewise vascular TCC deposits. In addition, five UC specimens and one colonic CD specimen contained TCC-positive fibrils in the muscularis mucosae or submucosa. There was no significant difference in vascular TCC deposits between UC and CD. The results suggested that terminal complement activation takes place in the intestinal lesions of IBD. [ABSTRACT FROM PUBLISHER]

Interactions between homing receptors on circulating leucocytes and endothelial addressins regulate tissue-specific cellular extravasation. Although integrin α4β7 appears to be the main receptor for gut-homing T lymphocytes, less is known about molecules mediating mucosal B cell homing. Expression of integrin α4β7 on B lymphocytes, B cell blasts, and plasma cells in human gut-associated lymphoid tissue (GALT; the Peyer's patches and appendix) and lamina propria was studied by multi-colour immunofluorescence applied on cryosections. Isolated mononuclear cells from the same tissue compartments were examined by flow cytometry and compared with peripheral blood B cells. Integrin α4β7 was expressed by IgA+ B cell blasts and plasma ceils (CD38high) in the lamina propria, B cell blasts in GALT, and sIgD+ B lymphocytes in peripheral blood. In contrast, GALT sIgD+ B lymphocytes were negative or only weakly positive for β4β7. These results suggested that B lymphocytes down-regulate α4β7 upon extravasation in GALT but up-regulate this integrin after antigen-priming. Thus, α4β7 may be a homing receptor also for B cell blasts extravasating in the gut lamina propria, where this integrin is maintained on plasma cells, perhaps as a local retention factor. [ABSTRACT FROM AUTHOR]

Jejunal biopsy specimens from 10 patients with treated coeliac disease and seven non-coeliac controls were challenged in vitro with peptic-tryptic gluten digest. Mucosal T cells were examined in situ by three-colour immunofluorescence staining for expression of the activation marker CD25 (the p55 α-chain of interleukin-2 receptor) and the nuclear proliferation marker revealed by monoclonal antibody Ki-67. Intraepithelial T cells expressed CD25 rarely whereas the proportion of activated lamina propria T cells increased (P < 0.002) from median 2.8% (cultured with 20% fetal calf serum alone for 24-48 h) to 10.0% after 24 h with gluten (n = 10; range 1.1-17.4%) and to 10.4% after 48 h (n = 7; range 1.4-17.5%). Such gluten-induced increase of CD25+ T cells was not observed in specimens from non-coeliac control subjects. Crypt-cell hyperplasia and T-cell proliferation (Ki-67+) were observed neither in the coeliac nor in the control mucosae after gluten stimulation. Three-colour staining combining a polyclonal antibody reagent to CD3 and a monoclonal antibody to CD25 with a monoclonal antibody to CD45R0, CD4, CD8, the p75 β-chain of interleukin-2 receptor, integrin αEβ7, or HLA-DR showed that most of the CD25+ T cells (> 90%) were CD4+CD8-, co-expressed CD45R0 and the p75 β-chain, and often also the integrin αEβ7 but not HLA-DR. In addition to these activated T cells, a dominating population of CD25+ CD3- CD4+ subepithelial pan-HLA-class II+ macrophages (CD68+) with variable expression of the p75 β-chain was often induced by gluten challenge. [ABSTRACT FROM AUTHOR]

T-cell receptor γ/&delta+ (TcRγ/&delta+) lymphocytes in human Peyer's patches (PP) adjoining ileal mucosa were studied by monoclonal antibodies with paired immunofluorescence staining in situ and by flow-cytometric phenotyping of isolated cells. The proportion of γ/&delta+ T cells in the follicle-associated epithelium outside the M-cell areas (median 4.1%, range 2.2-30.1%) was similar to that in mucosal villous epithelium (median 4.4%, range 0.5-30.5%). Most intraepithelial γ/&delta) cells (∼90%) expressed CD45R0 but only a few expressed CD8 (< 10%) and none L-selectin; a dominating subset (median 46%) employed the V&delta1/J&delta1 gene product (range 22-100%). The M-cell areas lacked γ/δ cells but contained clustered CD20+ and CD3+ lymphocytes. The subepithelial PP dome area and interfollicular (T-cell) zones, as well as the mucosal lamina propria, contained very few γ/&delta cells (median 1.7%, range 0.4-8.9%.) which were dominated (88-100%) by the Vδ2-encoded subset. Those in the dome area and lamina propria were often (∼75%) CD45R0+ (range 44-90%) while very few (∼2%) expressed L-selectin (range 0-15%). By contrast, CD45R0 expression on γ/&delta cells in the PP T-cell zones was relatively low (∼46%) and that of L-selectin relatively high (∼43%). In conclusion, TcR γ/&delta+ cells are quite rare in human PP and belong mainly to the V&delta2-encoded subset, thus being different from most intraepithelial γ/&delta cells that probably have another origin. The L-selectin+ fraction of PP γ/&delta cells presumably represent newly recruited 'naive' T lymphocytes while CD45R0+ γ/&delta cells both in PP and lamina propria are probably antigen-primed. [ABSTRACT FROM AUTHOR]

Expression of the γ/δ T-cell receptor (TcR) for antigen on CD3+ intraepithelial lymphocytes (IEL) was studied in situ by two-colour immunofluorescence on jejunal tissue sections from 24 patients with celiac disease and 17 controls. The proportion of intraepithelial TcRγ/delta; + cells was significantly increased (P<0.002) in untreated (median 20%, range 11-53%) as well as in treated (gluten-free diet) celiac disease (median 23%, range 16-55%) compared with controls (median 2%, range 0-39%). Although TcRα/β + IEL dominated both in controls and celiac disease, T cells expressing the TcRγ/δ were preferentially located within the epithelium rather than in the lamina propria. Paired staining for TcRγ/δ + and CD8 revealed that most (sim;90%) intraepithelial TcRγ/δ + lymphocytes in celiac disease were CD8-. A remarkably large fraction (median 67%, range 58-94%) of intraepithelial TcRγ/δ + cells expressed the Vδ1/Jδ1-encoded epitope revealed by monoclonal antibody &deta;TCSI. Our results suggested that increase of the intraepithelial TcRγ/δ + CD8- subset of T cells is particularly related to celiac disease. [ABSTRACT FROM AUTHOR]

Work-associated lower airway inflammation in waste collectors was examined by induced sputum and correlated with the bioaerosol exposure. Organic waste collectors (n=25) underwent induced sputum collection and spirometry before work on Monday and the following Thursday. Total cells, cell differentials, interleukin (IL)-8 and eosinophilic cationic protein were determined. Personal full-shift exposure measurements were performed Monday, Tuesday and Wednesday and analysed for total bacteria, fungal spores, endotoxins and beta(1-3)-glucans. The percentage of neutrophils (46-58%) and the IL-8 concentration (1.1-1.4 ng x mL(-1)) increased from Monday to Thursday. Forced expiratory volume in one second (FEV1) was significantly reduced on Thursday, and the decrease in FEV1/forced vital capacity correlated with the increase in the percentage of neutrophils. The median exposure to endotoxin (range 7-180 EU x m(-3)) and beta(1-3)-glucan (range 5-220 ng x m(-3)) was correlated with the increase in IL-8. Bioaerosol exposure during waste collection induced an inflammatory response in the lower airways, characterised by neutrophils and interleukin-8 secretion, that influenced the lung function. The inflammatory response was related to microbial components in the bioaerosol and was more pronounced for endotoxin than beta(1-3)-glucan exposure. No associations were found for mould spores or bacteria.

Harmless antigens encountered on the mucosal surface are normally tolerated in the sense that they do not induce inflammatory immune responses. Oral tolerance is the type of mucosal immune regulation that prevents inflammatory reactions to food proteins. However, parasites and invasive microorganisms at the mucosal surfaces must be recognised and dealt with in a proper manner. The immune system does so by cross-regulating the response where it either produces IgA to exclude invasion, IgE to fight parasites, or IgG to destroy the invasive organisms. Allergy is an anti-parasitic reaction to a misinterpreted but harmless antigen. This lack of tolerance induction is influenced by genetic factors controlling the amount of interleukin (IL)-4 produced initially in the immune response. IL-4 directs B-cells to produce IgE, induces naive T-cells to become IL-4 producing T-helper cells (Th2 cells) and prevent other T-cells from entering into the IFN-γ-producing Th1 pathway. Long lasting Th2 clones lose their IL-12 responsiveness and can no longer be induced to produce IFN-γ thus they are locked in an allergy inducing Th2 phenotype. Environmental factors play on the genetic background and influence the outcome of the immune response. Mucosal tolerance depends on an intact mucosal surface, is influenced by the age of the subject, and can be manipulated through antigen dose and place of entry. Immune-manipulating therapy may be more successful in primary than in secondary prevention of allergy.

In mice, integrin alpha 4 beta 7 is the main receptor used by lymphocytes that home to the Peyer's patches, although L-selectin contributes to the initial interaction with high endothelial venules. Less is known about the expression and function of these adhesion molecules in humans. The distribution of L-selectin and alpha 4 beta 7 on various B- and T-cell subsets was examined in human Peyer's patches (n = 8) and appendix (n = 4), collectively called gut-associated lymphoid tissue. Multicolor immunophenotyping was performed on cryosections, and dispersed cells were examined by flow cytometry. In cryosections, CD45RA+ T cells around and within interfollicular high endothelial venules, as well as surface (s)IgD+ B lymphocytes in the follicle mantles, often expressed abundant L-selectin but only intermediate levels of alpha 4 beta 7. CD45RO+ T cells and sIgD- B cells expressed higher levels of alpha 4 beta 7 and were often located near putative efferent lymphatics; only a small fraction (< 20%) of such memory cells expressed L-selectin. By flow cytometry, considerably more T than B lymphocytes co-expressed L-selectin and alpha 4 beta 7 (40% versus 25% and 67% versus 39%, respectively). In samples with many L-selectin+ cells (> 30%), more of these lymphocytes co-expressed alpha 4 beta 7 than in samples with few L-selectin+ cells. Because L-selectin and alpha 4 beta 7 were co-expressed on lymphocytes located near high endothelial venules, and because such co-expression was relatively common when many L-selectin+ cells were present, both of these molecules might participate in homing to human gut-associated lymphoid tissue. Such homing is probably most pronounced for T lymphocytes that were found to express L-selectin and alpha 4 beta 7 more often than B lymphocytes. The selective and relatively high expression of alpha 4 beta 7 on memory cells located near efferent lymphatics indicated a different migratory capacity; after exit from gut-associated lymphoid tissue, such stimulated cells might home mainly to mucosal effector sites.

The L1 antigen (calprotectin) is present in circulating monocytes but is restricted to certain subsets of tissue macrophages. Its expression is significantly increased in inflammatory bowel disease, apparently because of newly recruited monocytes. In vitro experiments were performed to substantiate lack of L1 upregulation in tissue macrophages, thereby justifying the use of this marker to detect newly recruited cells. Its reliability was further evaluated by studying mononuclear cell infiltrates characteristic of acute kidney rejection. After pro-inflammatory stimulation, monocytes matured in vitro (n = 12) as well as adherent mononuclear cells from normal small intestinal mucosa (n = 5) were examined for L1 expression by immunocytochemistry and by ELISA (cell lysates). In addition, peritubular mononuclear L1+ cells were examined by immunohistochemistry in routine biopsy specimens from transplanted kidneys with (n = 11) or without (n = 14) histopathologically diagnosed acute rejection. L1 was not upregulated in monocytes matured in vitro, nor in mucosal macrophages after stimulation with interferon-gamma, LPS, phorbol ester, or supernatant from activated leucocytes. In transplanted kidneys with signs of acute rejection, the fraction of L1+ macrophages was significantly increased (P < 0.001). Because L1 is persistently downregulated in mature tissue macrophages and is formalin-resistant, it identifies young infiltrating macrophages in routinely processed biopsy material. L1 should therefore be a valuable adjunct in the diagnosis of kidney rejection.

Two alternative integrins involved in mucosal homing (alpha 4 beta 7) or epithelial retention (alpha E beta 7) of lymphocytes were examined in the human gut. The distribution of the beta 7 subunit [monoclonal antibody (mAb) M301] was bimodal in that it was strongly expressed by alpha E beta 7 + cells but weakly by alpha 4 beta 7 + cells. More than 90% of intraepithelial lymphocytes (IEL), including the minor subsets of CD4+, T-cell receptor (TCR) gamma/delta +, and CD3- cells, expressed alpha E beta 7 as did most lamina propria CD8+ (88%) and a fraction (36%) of CD4+ lymphocytes. Conversely, B-lineage cells (CD19+) and macrophages (CD68+) were negative. In gut-associated lymphoid tissue (GALT: Peyer's patches and appendix) only a few (< 5%) cells were positive for alpha E beta 7 (confined to CD8+ lymphocytes and CD11c+ putative dendritic cells). A relatively small fraction of IEL (30-50%) expressed alpha 4 beta 7 (mAb Act-1), while most (70%) lamina propria T and B lymphocytes, blasts, plasma cells and macrophages were positive. In GALT, T lymphocytes expressed similar levels of alpha 4 beta 7 as in the lamina propria whereas relatively few B lymphocytes (< 50%) were positive. Isolated lamina propria CD8+, CD4+, CD19+, and CD38+ cells contained mRNA for alpha 4 and the former three subsets as well as appendix CD8+ cells also for beta 7 while only lamina propria CD8+ cells had mRNA for alpha E. Together, the results suggested that alpha E beta 7 and alpha 4 beta 7 are differentially regulated in inductive sites and effector sites of the human gut. Because lymphoid cells at both sites expressed mainly alpha 4 beta 7, this integrin may be a homing receptor on memory and effector cells bound for lamina propria as well as on naive lymphocytes extravasating in GALT. Conversely, because alpha E beta 7 was mainly expressed by CD8+ cells in epithelium and lamina propria, it was probably induced after extravasation, in agreement with the observation that IEL and a fraction of lamina propria T lymphocytes (mainly CD8+ cells) generally expressed higher levels of beta 7 than most CD4+ and B cells. Also a subset of putative dendritic cells located near the follicle-associated epithelium of GALT expressed alpha E beta 7, perhaps reflecting epithelial interaction during primary immune responses.

In conclusion, it appears that gluten induces a non-proliferative activation of CD4+ lamina propria T cells, but a proliferative activation in the TCR alpha/beta +CD8+ and TCR gamma/delta + IEL subsets. Although the increased density of TCR alpha/beta +CD8+ and TCR gamma/delta + IEL in the coeliac lesion may be secondary to T cell mediated immune activation in the lamina propria, a direct and primary antigen-driven proliferation of IEL (perhaps CD1-dependent29) cannot be excluded.

Multi-colour immunofluorescence can be a potent tool in investigating the cause of chronic inflammation of unknown etiology. The immune system uses two main mechanisms for target destruction; antibody-induced, complement-mediated or cell-mediated lysis. Immunohistochemical tracing of complement activation and immunoglobulin deposition in intestinal lesions from patients with ulcerative colitis and Crohn's disease show that ulcerative colitis may be an autoimmune disease characterized by an autoantibody of the IgG1 subclass to an apically located colonic autoantigen, which induce in situ complement activation and cellular lysis. The apical immune complex deposits co-localized with a previous identified putative autoantigen of 40kD, with peptide sequence homology with tropomyosin. Coeliac disease is a mucosal hypersensitivity to the wheat protein gluten. The intestinal lesion shows a peculiar disease specific increased percentage of T cell receptor (TCR)gamma/delta cells in the epithelium; differential use of CD45-isoforms and proliferative activation of intraepithelial T cells. T-helper(CD4) cells in the lamina propria show a non proliferative activation (CD25) which can be induced after gluten challenge in vitro. Immunomagnetic isolation of such in vitro activated T cells renders it possible to establish gluten reactive T cell clones that respond to gluten presented in the context of the disease associated human leucocyte antigen HLA-DQ2.

In conclusion, the "M(r) 40kD" antigen is expressed on the apical face of colonic enterocytes often in spatial relation to immune complexes in active UC, apparently targeting an IgG1-mediated autoimmune attack. Thus, our findings support the notion that an autoimmune response to the "M(r) 40kD" antigen, with local production of specific IgG1, is a possible immunopathological mechanism(s) in UC.

Expression of the gamma/delta T cell receptor (TCR) on CD3+ intraepithelial lymphocytes (IELs) was studied by two-colour immunofluorescence in duodenal tissue sections from healthy (n = 6) or infection-prone (n = 7) subjects with selective IgA deficiency (IgAD), and subjects (n = 4) with combined IgAD and IgG subclass deficiency. TCR gamma/delta+ IEL proportions in selective IgAD subjects (median 6.3%, range 1.0-41%) and in those with combined deficiency (median 4.5%, range 1.2-33%) were well within the range (0.3-38%) for histologically normal controls (n = 11), but the healthy IgAD subgroup tended to show raised TCR gamma/delta+ IEL proportions (median 13.6%) compared with the other two subgroups. Also the number of TCR gamma/delta+ IELs per intestinal length unit was relatively high (median 13.9/mm) in the healthy IgAD subjects, and significantly raised (P < 0.03) compared with controls (median 3.2/mm). Paired staining revealed that most TCR gamma/delta+ IELs in both selective IgAD (98%) and combined deficiency (99%) were CD8-, and a large fraction (median 84% and 63%, respectively) expressed the V delta 1/J delta 1-encoded epitope. The total number of CD3+ IELs (mostly CD8+) was similar to controls. IgAD subjects, and especially the healthy subgroup, had significantly increased serum concentrations of soluble CD8 (P < 0.0002), neopterin (P < 0.005), and beta 2-microglobulin (P < 0.007), which was similar to our previous observations in common variable immunodeficiency, and probably reflected stimulation of cell-mediated immunity. In addition, the increased TCR gamma/delta+ IELs might reflect a component of compensatory surface protection in the healthy IgAD subgroup.

Expression of the gamma/delta T-cell receptor (TCR) on CD3+ intraepithelial lymphocytes (IEL) was studied in situ by two-color immunofluorescence on duodenal tissue sections from 34 infection-prone, adult patients with various types of primary hypogammaglobulinemia, classical Bruton's, Bruton-like or congenital, and common variable immunodeficiency. TCR gamma/delta+ IEL proportions (median 4.3%, range 0.3-43.3%) were within the range (0.3-38.3%) for histologically normal controls (n = 11), and there was no significant difference between the three patient categories. The total number of CD3+ IEL (mostly CD8+) per intestinal length unit was significantly higher (P < 0.004) in patients than in controls. In addition, TCR gamma/delta+ IEL per length unit, as well as TCR gamma/delta+ IEL proportions, were significantly increased (P < 0.008 and P < 0.05) in 14 patients with intestinal villous atrophy. Paired staining revealed that most (approximately 94%) TCR gamma/delta+ IEL in B-cell deficiency were CD8-, and a large fraction (approximately 67%) expressed the V delta 1/J delta 1-encoded epitope. No relationship was found between CD3+ or TCR gamma/delta+ IEL and the number of CD3+, CD4+, CD8+, and B lymphocytes or the CD4:CD8 ratio in peripheral blood. TCR gamma/delta+ IEL thus appeared to maintain a normal distribution in B-cell deficiency, except for being increased in patients with villous atrophy. Enhanced T-cell-mediated immunity, as possibly reflected by the numerical increase of CD3+ IEL, might compensate for a deficient mucosal B-cell system.

In order to identify any dominating subset of activated T cells in the celiac lesion, we examined CD3+, CD4+, CD8+ and T cell receptor (TcR) gamma/delta+ lymphocytes in jejunal cryosections from 25 patients with celiac disease and 10 controls by three-color immunofluorescence staining for expression of the nuclear proliferation marker detected by monoclonal antibody (mAb) Ki-67 and the p55 alpha chain of interleukin-2 receptor (CD25). mAb Ki-67+ intraepithelial lymphocytes (IEL) were exclusively observed in celiac patients. The median proportion of CD3+ IEL positive for Ki-67 increased from nil in controls to 4.5% in partly treated (range 0-19.0%; n = 10; p = < 0.05) and 12.8% in untreated celiac disease (range 4.0-30.7%; n = 15; p < 0.005). Only 1.5% of CD3+ subepithelial T cells expressed the Ki-67 marker in celiac disease (range 0-9.5%). Two- and three-color staining combining mAb to CD3 and Ki-67 with mAb to CD4, CD8 or TcR delta showed that both TcR alpha/beta+ CD8+ and TcR gamma/delta+ (but not CD4+) mucosal T cells proliferated in the epithelium. By contrast, CD25 were almost exclusively expressed on CD4+ T cells in the lamina propria. The percentage of CD25+ T cells increased significantly from 1.7% in controls (range 0-2.9%) to 7.5% in partly treated (range 0.8-17.8%, p < 0.002), and to 14.65% in untreated celiac disease (range 3.9-21%, p < 0.002). These results suggest that gluten ingestion in celiac disease induces proliferative activation of TcR alpha/beta+ CD8+ and TcR gamma/delta+ IEL but non-proliferative activation (lymphokine production?) of lamina propria CD4+ T cells.

Patients with celiac disease produce not only immunoglobulin A (IgA) but also immunoglobulin G (IgG) and M (IgM) antibodies to gluten. Intake of dietary gluten may hence induce local complement activation and mucosal damage. Jejunal tissue sections from adult patients with celiac disease were examined by immunofluorescence with monoclonal antibodies to activation neoepitopes in C3b and the terminal complement complex (TCC). Subepithelial deposition of TCC was observed in 93% of 28 untreated and in 57% of 23 partly treated study subjects. The immunofluorescence staining intensity was well correlated with the serum level of gluten-specific IgG and IgM (but not IgA), the number of mucosal IgG-producing cells, and the degree of villous atrophy. Similar immune deposits were not observed in 5 successfully treated patients with celiac disease, 5 patients with dermatitis herpetiformis without jejunal villous atrophy, and 90% of 21 control patients with histologically normal jejunal mucosa. Gluten challenge increased the amount of subepithelial TCC and produced additional C3b deposition, suggesting recent complement activation. Ingested gluten might thus, via Ig-mediated subepithelial complement activation, damage the surface epithelium in celiac disease and induce compensatory crypt hyperplasia.

Vitronectin stores were localized in human platelets by immunofluorescence, confocal laser microscopy, and immunoelectron microscopy. In ultrathin cryosections, the immunogold labelling was mainly observed in the alpha-granules, although occasional immunolabelling of the platelet surface was present. The interaction between thrombin-stimulated platelets and exogenously added vitronectin was also explored. This vitronectin had been purified by heparin-affinity chromatography. Binding of vitronectin to thrombin-stimulated platelets was demonstrated by flow cytometry. The immunoelectron microscopical studies revealed that this binding was mainly restricted to clumps of released alpha-granular material on the platelet surface. The possible significance of the interaction between exogenous vitronectin and released platelet proteins is discussed.

Epithelial HLA class II is differentially expressed (DR >> DP) only after birth in salivary glands and small intestinal mucosa, in contrast to class I determinants and secretory component (SC) which appear early in gestation. However, there is a brisk postnatal increase in SC expression along with the class II induction, suggesting stimulation by cytokines from activated immune cells. T lymphocytes remain quite scanty in postnatal salivary glands, and the striking SC and class II expression might reflect a synergistic effect of IFN-gamma and TFN-alpha on immature epithelial cells. Enhanced epithelial expression of both SC and class II in salivary glands from sudden infant death victims could be the effect of immunostimulation caused by an infectious agent. Strikingly upregulated SC and epithelial class II expression (DR > DP > DQ) is seen in various inflammatory lesions such as obstructive sialadenitis, Sjögren's syndrome, chronic gastritis, and celiac disease. IFN-gamma and TNF-alpha are most likely involved as the expression patterns can be reproduced with these cytokines in vitro on colonic epithelial cell lines. However, these molecules of the Ig supergene family do not show a selective response in epithelia of inflammatory lesions because increased expression is also seen for lysozyme, lactoferrin and some other proteins. ICAM-1 can be upregulated on epithelial cells by various cytokines in vitro although the situation remains uncertain in mucosal inflammation. The expression pattern in IBD is complicated by dysplastic epithelial changes leading to reduced SC levels which may thus, in turn, jeopardize the poly-Ig transport mechanism. Epithelial class II molecules appear to have antigen-presenting properties, but the immunopathologic role of their increased expression in inflammatory disease in terms of induction of autoimmunity and/or abrogation of oral tolerance is a matter of continuing dispute.

To trace sites for local complement activation in inflammatory bowel disease, an indirect two-colour immunofluorescence method was applied on prewashed and directly ethanol-fixed mucosal specimens from patients with ulcerative colitis, Crohn's colitis, or terminal ileitis. Monoclonal antibodies to the IgG subclasses and to neoepitopes of activated complement C3b and the terminal complement complex (TCC) were used in combination with rabbit antiserum to immunoglobulins and various complement components. Deposits of activated C3b were found on the luminal face of the surface epithelium in the most affected ulcerative colitis specimens from 91% of 23 studied patients, together with cytolytic TCC in 81%. Furthermore, there was a selective deposition of the immunoglobulin G subclass 1 (IgG1) within the epithelial immune complexes in 63% of 11 studied patients. These results suggested that IgG1 autoantibodies to brush-border antigen(s) induce a complement-mediated attack on the epithelium in ulcerative colitis. The epithelial complement deposition seen in Crohn's disease tended to be more granular and was observed in 5 of 10 patients with colitis and in 4 of 10 with ileitis. No co-localization of IgG was observed, suggesting that complement activation had been induced by the alternative pathway. Type III immune reaction may, in addition, take place in both diseases since there was evidence of continuous vascular complement activation in submucosal blood vessels.

Expression of CD45RO on intraepithelial lymphocytes (IEL) bearing the T-cell receptor (TcR) alpha/beta or gamma/delta was studied in situ by three-colour immunofluorescence on jejunal tissue sections from 21 patients with coeliac disease and eight controls. CD45RA-TcR alpha/beta+ IEL expressed CD45RO significantly more often (75%) than the preferentially expanded TcR gamma/delta+ counterpart (59%). Triple staining for CD3, CD4/8 and CD45RA or CD45RB revealed that all CD3 + 4 - 8 - IEL (taken to be TcR gamma/delta+) expressed CD45RB and none were CD45RA. CD45RO positivity was of the same magnitude (66%) on the predominating monoclonal antibody delta TCS1-reactive fraction of TcR gamma/delta+ cells as on the remainder of the TcR gamma/delta+ subset. These results suggest that gluten exposition in patients with coeliac disease leads to accumulation of CD45RA-, putative antigen-primed memory cells of both TcR phenotypes. The less marked CD45RO expression within the preferentially expanded TcR gamma/delta+ subset of IEL may be of particular biological interest.

Expression of various CD45 isoforms (RA, RB and R0) on CD3+, CD4+ and CD8+ intraepithelial and lamina propria T cells was examined in situ by a three-color immunofluorescence technique in jejunal biopsy specimens from 32 patients with celiac disease and 18 controls. The median percentage of CD3+ intraepithelial lymphocytes (IEL) that expressed CD45R0 increased from 52% in controls to 69% in untreated celiac disease (p less than 0.01). Furthermore, the percentages of CD4+ and CD8+ IEL strongly positive for CD45R0 rose respectively from 94% and 24% in controls to 100% and 55% in untreated celiac disease. Conversely, CD45R0 was strongly expressed on most CD3+ lamina propria lymphocytes (LPL) both in control (81%) and diseased (77%-81%) mucosa. A variable fraction of the intraepithelial and lamina propria CD3+ T cells expressed mainly CD45RB (controls, 46% and 20%, respectively; celiac disease, 29% and 15%). Only 2% IEL and 4% LPL were positive for CD45RA. Expression of different CD45R isoforms thus identified three distinct CD8+ T cell subsets in human intestinal mucosa. In addition, our results suggested that antigen-primed CD8+CD45R0+ memory cells accumulate in the jejunal epithelium of patients with untreated celiac disease.

The epithelial destruction seen in ulcerative colitis remains unexplained. Complement activation has been proposed to be involved, but no definite evidence has been available to this end. In the present study, we examined immunohistochemically ulcerative colitis lesions with monoclonal antibodies to activation neoepitopes in the complement component C3b and in the cytolytically active terminal complement complex. Colonic tissue specimens from 23 patients with ulcerative colitis were examined by indirect two-color immunofluorescence staining with monoclonal antibodies to the four human immunoglobulin G subclasses and to activated complement C3b or terminal complement complex. All except two patients had activated C3b deposited apically on the surface epithelium of involved mucosa. Immunoglobulin G1 was found on the epithelium in extensively prewashed specimens from 7 of 11 patients, and a striking colocalization of immunoglobulin G1, C3b, and terminal complement complex was observed in 4. Immune deposits were not observed in 31 noninflamed specimens from the same ulcerative colitis patients. Only 1 of 44 histologically normal mucosae from 17 controls and 1 of 10 colonic adenomas contained some epithelial complement deposits. It is concluded that activated complement is often deposited along the brush border of the surface epithelium in active ulcerative colitis lesions and may be associated with immunoglobulin G1 autoantibody.

Extensively washed and ethanol-fixed colonic specimens from 10 patients with ulcerative colitis, 3 patients with Crohn's disease of the colon, and 8 histologically normal controls were examined by two-color immunohistochemistry with monoclonal antibody to a neoepitope in the terminal complement complex combined with antiserum to factor VIII-related antigen (von Willebrand's factor), C3c, C3d, or C5. An alternative combination was monoclonal antibody to S-protein and antiserum to C9. Submucosal vessel walls in both normal and diseased colon showed parallel positivity for C3d, C5, C9, terminal complement complex, and S-protein, but the staining intensity and the proportion of positive vessels were significantly higher in inflammatory bowel disease than in controls. In addition, there was significantly more C3c reactivity associated with the terminal complement complex-positive submucosal vessels of active inflammatory bowel disease lesions than in histologically normal colon. Vascular C activation may therefore be a continuous process in active inflammatory bowel disease lesions, presumably related to the degree of inflammation and immune complex formation.

In Caucasoids HLA-DQB1 genes encoding amino acids other than aspartic acid at position 57 of the DQ beta chain (non-Asp-57) are associated with susceptibility to develop insulin-dependent diabetes mellitus (IDDM), while resistance is associated with aspartic acid at this residue (Asp-57). Following amplification of genomic DNA by the polymerase chain reaction, the DQB1 alleles of 87 random Norwegian IDDM patients and 187 healthy controls were investigated with 11 different sequence-specific oligonucleotide probes. Of these patients 82% carried DQB1 alleles encoding non-Asp-57 at both of their DQ beta chains, compared to 27% of the controls (relative risk = 12.2, p less than 0.0001). Sixteen percent of the patients (versus 51% of the controls) were heterozygous Asp-57/non-Asp-57. Two percent of the patients (22% of the controls) were apparently Asp-57 homozygous. The results demonstrate that non-Asp-57 DQ beta chains are associated with susceptibility to develop IDDM but also indicate that the protection associated with DQ beta Asp-57 may not be as dominant as reported by others.

Expression of the gamma/delta T-cell receptor (TcR) for antigen on CD3+ intraepithelial lymphocytes (IEL) was studied in situ by two-colour immunofluorescence on jejunal tissue sections from 24 patients with coeliac disease and 17 controls. The proportion of intraepithelial TcR gamma/delta+ cells was significantly increased (P less than 0.002) in untreated (median 20%, range 11-53%) as well as in treated (gluten-free diet) coeliac disease (median 23%, range 16-55%) compared with controls (median 2%, range 0-39%). Although TcR alpha/beta+ IEL dominated both in controls and coeliac disease, T cells expressing the TcR gamma/delta were preferentially located within the epithelium rather than in the lamina propria. Paired staining for TcR gamma/delta and CD8 revealed that most (approximately 90%) intraepithelial TcR gamma/delta+ lymphocytes in coeliac disease were CD8-. A remarkably large fraction (median 67%, range 58-94%) of intraepithelial TcR gamma/delta+ cells expressed the V delta 1/J delta 1-encoded epitope revealed by monoclonal antibody delta TCS1. Our results suggested that increase of the intraepithelial TcR gamma/delta+ CD8- subset of T cells is particularly related to coeliac disease.