Generation of mucosal cytotoxic T cells against soluble protein by tissue-specific environmental and costimulatory signals

We compared peripheral and mucosal primary CD8 T cell responses to inflammatory and noninflammatory forms of antigen in a T cell-adoptive transfer system. Immunization with the soluble antigen, ovalbumin (ova), administered i.p. or orally without adjuvant, activated nonmucosal CD8 T cells but did not induce cytotoxic activity. However, after activation, the transferred cells entered the intestinal mucosa and became potent antigen-specific killers. Thus, exogenous intact soluble protein entered the major histocompatibility complex class I antigen presentation pathway and induced mucosal cytotoxic T lymphocytes. Moreover, distinct costimulatory requirements for activation of peripheral versus mucosal T cells were noted in that the CD28 ligand, B7-1, was critical for activated mucosal T cell generation but not for activation of peripheral CD8 T cells. The costimulator, B7-2, was required for optimum activation of both populations. Infection with a new recombinant vesicular stomatitis virus encoding ovalbumin induced lytic activity in mucosal as well as peripheral sites, demonstrating an adjuvant effect of inflammatory mediators produced during virus infection. Generation of antiviral cytotoxic T lymphocytes was also costimulation-dependent. The results indicated that induction of peripheral tolerance via antigen administration may not extend to mucosal sites because of distinct costimulatory and inflammatory signals in the mucosa.     

Tropomyosin isoforms in intestinal mucosa: production of autoantibodies to tropomyosin isoforms in ulcerative colitis

BACKGROUND & AIMS: Autoantibodies against tropomyosins (TMs) have been reported in ulcerative colitis (UC). In this study the hTM isoforms (hTM1-5) present in intestinal epithelial cells and in smooth muscle were investigated, and the immunoreactivity against hTMs by immunoglobulin G (IgG) produced in vitro by colonic mucosal lymphocytes (LPMCs) from patients with UC, Crohn's disease (CD), and controls was examined. METHODS: TMs were extracted from colonic and jejunal epithelial cells and smooth muscle, and hTM isoforms were identified using isoform-specific monoclonal antibodies by enzyme-linked immunosorbent assay and transblot analysis. The immunoreactivity of IgG produced by colonic LPMCs was analyzed against the recombinant hTM isoforms. RESULTS: The major hTM isoforms present in colonic and jejunal epithelial cells are hTM5 and hTM4, whereas intestinal smooth muscle contains the hTM1-3 isoforms. The IgG synthesized in vitro by LPMCs from UC (n = 19) recognized hTM5 and hTM1, more significantly (P < 0.04 to <0.001) when compared with CD (n = 12) and controls (n = 17). However, IgG produced by LPMCs from CD did not show such anti-hTM reactivity. Mucosal anti-hTM IgG mainly belonged to the IgG1 subclass. CONCLUSIONS: Intestinal epithelial cells and smooth muscle have distinct hTM isoforms. Patients with UC, and not CD, show mucosal autoantibody response against hTM isoforms, particularly hTM5 and hTM1.     

p126 (CDw101), a costimulatory molecule preferentially expressed on mucosal T lymphocytes

Intestinal mucosal lymphocytes are defined by their anatomic location within the epithelium (intraepithelial lymphocytes), the interstitium between the epithelial basement membrane and the underlying muscularis mucosa (lamina propria lymphocytes), or in organized lymphoid tissues (Peyer's patches). Although intestinal intraepithelial lymphocytes have a distinct localization, their function has not been determined. To define cell surface proteins that are involved in intestinal intraepithelial lymphocyte localization or function, cultured human mucosal lymphocytes were used as immunogens to develop mAbs that react predominantly with this cell population in an immunohistochemical screening assay. Three mAbs were selected that subsequently were found by biochemical analysis to identify a 200-kDa homodimeric polypeptide on 88 to 98% of CD3+ mucosal lymphocytes but only 18 +/- 13% of PBLs. Expression on granulocytes and monocytes was also observed. This polypeptide has been termed p126 based on its SDS-PAGE-determined M(r) under reducing conditions. Cleveland digest maps demonstrated similarity between the p126 and CDw101 polypeptides. Determined amino acid sequence analysis of the purified p126 polypeptide revealed that it is the protein product of the recently identified V7 gene, which has structural similarities to members of the Ig gene superfamily. Two of the anti-p126 mAbs were costimulatory with suboptimal concentrations of anti-CD3 mAb inducing proliferation of cultured intestinal intraepithelial lymphocytes. Thus, we conclude that p126 is CDw101 encoded by a gene that predicts a seven-Ig domain chain-like structure. It has restricted expression predominantly on mucosal T lymphocytes and appears to have a costimulatory function of special relevance for CD28- T cells and for mucosal lymphocytes.     

Simultaneous cross-linking of CD6 and CD28 induces cell proliferation in resting T cells

In the present study, we showed that simultaneous ligation of the monoclonal antibodies (mAb) against CD6 and CD28 induces T-cell proliferation in purified resting T lymphocytes in the absence of T-cell receptor (TCR) occupancy. No cell proliferation was observed when the mAb were cross-linked alone or used simultaneously in the soluble form. T-cell proliferation mediated through CD6/CD28 is accompanied by the up-regulation of interleukin-2 (IL-2) mRNA and expression of IL-2 receptors on the cell surface. In the presence of IL-2-neutralizing mAb the proliferative response of the T cell induced through CD6/CD28 was inhibited dose dependently. Cross-linking mAb to CD6 and CD28 alone or together did not down-regulate the CD3/TCR complex. T-cell proliferation mediated through CD6/CD28 was only partially blocked by the immunosuppressive drug, cyclosporin A (CsA), whereas anti-CD28-induced T-cell proliferation in the presence of the phorbol ester, 12-O-tetradecanoylphorbol-13-acetate (TPA), was unaffected. In sharp contrast T-cell proliferation mediated by anti-CD6 in the presence of TPA was efficiently blocked by CsA. In addition, two protein kinase C (PKC) inhibitors, GF 109203X and H-7 dose-dependently inhibited T-cell proliferation mediated through CD6/CD28, suggesting that PKC activation may be involved. Furthermore, there was a marked differential dose-dependent inhibitory effect of the PKC inhibitors on T-cell proliferation mediated by the co-ligation of anti-CD6 or anti-CD28 in the presence of anti-CD3, with the former being more sensitive to PKC inhibition. Taken collectively, our results suggest that T-cell activation can occur through an antigen-independent pathway by cross-linking the accessory molecules, CD6 and CD28, and that these two cell surface antigens may have distinct signalling pathways.     

A comparison of the antigen-presenting capabilities of class II MHC-expressing human lung epithelial and endothelial cells

Human lung alveolar epithelial cells constitutively express class II major histocompatibility complex (MHC). Human lung microvascular endothelial and small airway epithelial cells can be induced to express class II MHC by stimulation with the pro-inflammatory cytokine interferon-gamma. The levels of class II MHC on lung epithelial and endothelial cells were comparable to those seen on an Epstein-Barr virus (EBV)-transformed B-cell line. However, the costimulatory molecules B7-1 and B7-2 were not expressed. The ability of the class II MHC expressing human lung parenchymal cells to present alloantigen to CD4+ T lymphocytes was investigated. Freshly isolated human alveolar epithelial cells (type II pneumocytes) and monolayers of interferon-gamma-stimulated small airway epithelial and lung microvascular endothelial cells were co-cultured with allogeneic CD4+ T lymphocytes and proliferation determined by [3H]thymidine incorporation. A clear difference was observed between effects of the epithelial and endothelial cells on CD4+ T-lymphocyte activation. Alveolar and small airway epithelial cells failed to stimulate the proliferation of allogeneic CD4+ T lymphocytes whereas lung microvascular endothelial cells did stimulate proliferation. This difference could not be explained by the levels of class II MHC or the lack of B7-1 and B7-2 solely. Microvascular endothelial cells, and not alveolar or small airway epithelial cells, possess B7-independent costimulatory pathways.     

CD8+CD103+ T cells analogous to intestinal intraepithelial lymphocytes infiltrate the pancreas in chronic pancreatitis

OBJECTIVE: Chronic pancreatitis is a painful chronic inflammatory disease of the exocrine pancreas that is associated with the replacement of functional parenchyma by extended fibrosis and with a massive infiltration of T lymphocytes. However, to date further characterization of infiltrating T cells in chronic pancreatitis has not been undertaken. METHODS: Using the novel method of multiepitope imaging with fluorochrome-tagged specific monoclonal antibodies, which allows the simultaneous localization and characterization of T cells in tissues, we analyzed the distribution and phenotypes of T cells infiltrating the pancreas in chronic pancreatitis. RESULTS: The mean CD4:CD8 ratio in 10 cases of chronic pancreatitis was 2.4:1. In order of decreasing frequency, the following markers were observed: CD45RO, CD18, TCRgammadelta, and CD103. The lymphocytes, especially of the CD4+ subset, were found mainly in the fibrous stroma, but T cells were also observed periductally. A T-cell subset bearing the phenotype CD8+CD103+, analogous to intestinal intraepithelial lymphocytes, was found intracalating between the cells of the ductal epithelium. CONCLUSIONS: Phenotyping of the T lymphocytes in chronic pancreatitis supports the concept of the involvement of cell-mediated cytotoxicity in the pathogenesis of this disease. In addition, intraepithelial lymphocytes were found interspersed between the ductal epithelial cells, pointing to a role of this T-cell subset as a first-line defense against deleterious epithelial events in chronic pancreatitis.     

Activation of T lymphocytes by syngeneic murine intestinal smooth muscle cells

BACKGROUND & AIMS: Intestinal smooth muscle cells (ISMCs) express major histocompatibility complex II (MCH II) and intercellular adhesion molecule 1 (ICAM-1) after exposure to interferon gamma (IFN-gamma). T lymphocytes invade the intestinal musculature during Crohn's disease or pseudoobstruction. The aim of this study was to determine whether ISMCs activate syngeneic T cells via MHC II and ICAM-1. METHODS: Cultured murine ISMCs were exposed to IFN-gamma for 72 hours and analyzed for Mac-1 (CD11B CD18) antigen, MHC II, and ICAM-1 expression using enzyme-linked immunosorbent assay and fluorescence-activated cell sorter scan. T lymphocytes from mesenteric lymph nodes of ovalbumin-sensitized mice were examined for their ability to proliferate after coculture with IFN-gamma-pretreated and ovalbumin-pretreated ISMCs using [3H]thymidine incorporation. RESULTS: ISMCs expressed smooth muscle alpha-actin before and after IFN-gamma exposure. No macrophages were identified in these cultures. Exposure to IFN-gamma and ovalbumin for 72 hours induced MHC II and ICAM-1 expression; these treated ISMCs induced T-cell proliferation, whereas untreated ISMCs did not. T-cell proliferation was markedly enhanced by adding interleukin 2 and was blocked by antibodies against MHC II and ICAM-1. CONCLUSIONS: ISMCs activate T lymphocytes in an MHC II-linked manner and thus possess the ability to modulate immune function in the gut.     

Signal transduction and glycophosphatidylinositol-linked proteins (lyn, lck, CD4, CD45, G proteins, and CD55) selectively localize in Triton-insoluble plasma membrane domains of human leukemic cell lines and normal granulocytes

Src-family nonreceptor protein tyrosine kinases (NRPTK) are associated with cell surface receptors in large detergent-resistant complexes: in epithelial cells, yes is selectively located in vesicle structures containing caveolin ("caveolae"). These formations are typically also endowed with glycophosphatidylinositol (GPI)-anchored proteins. In the present study, we observed lck, lyn, src, hck, CD4, CD45, G proteins, and CD55 (decay-accelerating factor) expression in the buoyant low-density Triton-insoluble (LDTI) fraction of selected leukemic cell lines and granulocytes. We provide a detailed analysis of the two most highly expressed NRPTK, p53/p56lyn and p56lck, which are involved in the transduction of signals for proliferation and differentiation of monocytes/B lymphocytes and T lymphocytes, respectively. We show that lyn is selectively recovered in LDTI complexes isolated from human leukemic cell lines (promyelocytic [HL-60], erythroid [K562] and B-lymphoid [697]) and from normal human granulocytes, and that lck is recovered from LDTI fractions of leukemic T- and B-lymphoid cell lines (CEM, 697). In LDTI fractions of leukemic cells, lck and lyn are enriched 100-fold as compared with the total cell lysates. Analysis of these fractions by electron microscopy shows the presence of 70- to 200-nm vesicles: lyn and lck are homogenously distributed in the vesicles, as revealed by an immunogold labeling procedure. These novel results propose a role for these vesicles in signal transduction mechanisms of normal and neoplastic hematopoietic cells. In support of this hypothesis, we further observed that molecules participating in B- and T-cell receptor activation cofractionate in the LDTI fractions, CD45/lyn (B cells) and CD45/lck/CD4 (T cells).     

Intraepithelial lymphocytes and their recognition of non-classical MHC molecules

Recent studies of the TCR alpha and beta chains expressed by normal human IELs suggest that these intestinal lymphocytes are directed at a limited set of antigens, presumably on intestinal epithelial cells in view of their anatomic location. The direct sequence analysis of these cells has indicated that they are oligoclonal and cannot, therefore, be responding to the complex mixture of antigens which are present in the lumen. The abundant expression of the CD8 accessory molecule by the IELs, in addition, indicates that these putative intestinal epithelial cell antigens are presented by MHC class I or I-like molecules. The expression of CD8 also suggests that these cells function biologically in part as cytolytic T lymphocytes which is consistent with a variety of functional studies. Taken together with their expression of the CD45RO isoform, these phenotypic and functional observations suggest that iIELs are cytolytic, memory cells which are responsive to an extremely limited number of antigens bound to major histocompatibility complex (MHC) class I or class I-like molecules. Several non-polymorphic MHC class I-like molecules such as Qa, the thymus leukemia antigen (TL) and CD1 in the mouse and CD1 in human represent important candidate ligands for these oligoclonal iIELs. TL and CD1 are expressed specifically by murine intestinal epithelial cells. In humans, CD1d is constitutively expressed by intestinal epithelial cells. In addition, we have isolated iIEL T cell clones which specifically recognize members of the CD1 gene family when expressed on a transfected B cell line that lacks HLA-A and B and have shown that the proliferation of peripheral blood T cells to intestinal epithelial cells is CD1d dependent. Thus, the evidence to date strongly implicate the nonpolymorphic, class Ib molecules as novel restriction elements for unique populations of lymphocytes within the intestinal epithelium.     

In situ characterization of gingival mononuclear cells in rapidly progressive periodontitis

Rapidly progressive periodontitis (RPP) has been suggested as a distinct clinical entity within the spectrum of early onset periodontitis. Immunological mechanisms have been considered in the pathogenesis of RPP. This study was designed to evaluate the distribution and phenotypic properties of the lymphocyte populations within the affected gingival tissue of patients with RPP. Biopsies were obtained from 16 patients between 22 and 33 years of age. The tissue samples were processed for both histopathological and immunohistochemical examinations. Gingival tissue T lymphocytes (CD3+), helper T cells (CD4+), suppressor-cytotoxic T cells (CD8+), and cells positive for HLA-DR antigen were identified using monoclonal antibodies with an immunoperoxidase technique. Intracytoplasmic immunoglobulin-containing cells were also stained immunohistochemically with polyclonal antibodies. CD3+ cells were mainly located beneath the pocket epithelium. CD4+ and CD8+ cells were evenly distributed within this T-cell infiltrate with a CD4+/CD8+ ratio of 1:12. Numerous HLA-DR+ cells were also observed in the lymphocytic infiltrates. The majority of mononuclear cells located throughout the stroma were IgG+ plasma cells. Our results indicate that RPP patients present an IgG-bearing plasma cell dominated lesion with equal participation of both T-cell subpopulations. These findings suggest that activation and proliferation of B-cells play an important role in the pathogenesis of periodontal diseases.     

Vascular density in melanoma xenografts correlates with vascular permeability factor expression but not with metastatic potential

Previous studies have shown that human airway epithelial cells (AEC) can stimulate allogeneic peripheral blood T-lymphocyte (PBT) proliferation. We now sought to determine which AEC surface molecule/T-cell coreceptors are involved in this process. AEC-induced PBT proliferation was inhibited by 25 microM genestein or herbamycin A (0.9 and 1.8 microM), both tyrosine kinase inhibitors. Anti-phosphotyrosine immunoblots performed on PBT lysates after coculture with AEC demonstrated phosphorylation of 56kD and 60kD bands. To determine whether CD3 associated p59fyn, or CD4 and CD8 associated p56lck phosphotyrosine kinases (PTK) were involved, we assayed kinase activity in lymphocyte lysates immunoprecipitated with anti-p56lck and p59fyn mAbs. PBT cells or murine T-cell line transfectants expressing human CD4 (3G4) or human CD8alpha (3G8) were cocultured with AEC or A549, an alveolar-like cell line lacking class II Ag expression. After A549 or AEC coculture, p56lck activity in PB T-cells peaked at 2 min whereas p59fyn kinase activity continued to rise at 8 min. AEC (expressing class II Ags) stimulate PTK activity in both 3G8 and 3G4 cells. A549 stimulated p56lck in 3G8, but not in 3G4 cells. This activation of p56lck was not blocked by preincubation of A549 with anti-class I or anti-CD1d mAbs. An antibody generated in our laboratory, which recognizes an epithelial specific surface molecule (mAb L12) and which blocks AEC driven PBT proliferation, was shown to block PTK activity of peripheral blood T-cell lysates, though not of 3G8 lysates. These studies suggest that AEC are capable of stimulating CD4 and CD8 associated lck and CD3 associated fyn kinases through class II dependent and independent pathways.     

Glutamine deprivation induces apoptosis in intestinal epithelial cells

BACKGROUND: Glutamine is the most abundant amino acid in the blood, and its deprivation leads to gut mucosal atrophy. The small intestinal mucosa is maintained by a balance between cell proliferation and cell death by apoptosis. We reported that glutamine is required for nitrogen-stimulated proliferation in intestinal epithelial cells. We do not know whether glutamine regulates apoptosis in the gut. The purpose of this study is to determine whether glutamine deprivation induces apoptosis in rat intestinal epithelial (RIE-1) cells and to compare the effect of glutamine starvation with that of methionine and cysteine (Met/Cys) starvation. METHODS: RIE-1 cells were deprived of either glutamine or Met/Cys for 24 hours. Cell numbers were determined by cell counting and tetrazolium enzymatic assay. Apoptosis was quantified by Annexin V assay and confirmed by DNA gel electrophoresis and Hoecsht nuclear staining. RESULTS: Deprivation of glutamine or Met/Cys resulted in decreased cell numbers. However, only the glutamine-deprived group showed significant induction of apoptosis with increased Annexin V staining, DNA laddering, and nuclear condensation. CONCLUSIONS: This study provides biochemical and morphologic evidence that glutamine deprivation induces apoptosis in rat intestinal epithelial cells. In contrast, Met/Cys starvation suppresses cell number without induction of apoptosis. These results suggest that glutamine serves as a specific survival factor in enterocytes.     

Myeloma bone marrow plasma cells: evidence for their capacity as antigen-presenting cells

Myeloma plasma cells constitute 10% to 90% of the total bone marrow cell count in patients with multiple myeloma (MM). These cells express a variety of cell surface markers, such as HLA-ABC and HLA-DR, and surface antigens that are necessary for professional antigen-presenting cells, including adhesion and costimulatory molecules. In this study, we examined the expression of major histocompatability complex (MHC) and costimulatory molecules on CD38(bright,++) plasma cells in bone marrow aspirates from eight MM patients. Small percentages of plasma cells expressed weak but detectable levels of HLA-DR, HLA-DQ, CD40, CD80, and CD86, which could be upregulated by interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha. CD38++ plasma cell and CD38(dim,+) cells were sorted from freshly isolated bone marrow mononuclear cells and tested for their capacity to act as antigen-presenting cells. Indeed, both CD38++ plasma cells and CD38+ cells were able to stimulate allogeneic T cells and present the soluble antigens purified protein derivative and tetanus toxoid to autologous T cells. Recognition of the antigens led to T-cell proliferation and secretion of IFN-gamma and was MHC class-I and -II restricted. Antigen processing and presentation by CD38++ and CD38+ cells were abolished by treatment of the cells with chloroquine. Hence, our study provides for the first time evidence that myeloma plasma cells may act as antigen-presenting cells. Further studies are warranted to examine in detail the molecules required for inducing T-cell stimulation.     

The regulation and functional consequence of proinflammatory cytokine binding on human intestinal epithelial cells

Products of an activated immune system may affect cells within the immune system as well as nonlymphoid cells in the local environment. Given the immunologically activated state of the intestinal tract, it is conceivable that locally produced cytokines could regulate epithelial cell function. To assess whether epithelial cells are targets for particular cytokines, we initiated studies on the binding of a panel of proinflammatory cytokines in freshly isolated epithelial cells from normal and inflammatory bowel disease (IBD) patients as well as in cell lines. Isolated intestinal epithelial cells (IEC) were stained with phycoerythrin-conjugated or biotinylated cytokines to determine the expression and density of receptors for IL-1beta, IL-6, granulocyte-macrophage CSF (GM-CSF), and TNF-alpha. Receptors for IL-1beta, IL-6, and GM-CSF were readily detectable in all epithelial cell preparations at levels equal to (GM-CSFR) or lower than those seen on monocytes. However TNFalpha-R were not detectable on freshly isolated IECs. Receptor density was greater in surface vs crypt epithelial cells, but no significant differences were seen between normal and IBD epithelial cells. Expression of IL-1R and IL-6R was enhanced by LPS and IFN-gamma. Functionally, IL-1beta enhanced proliferation of the IEC cell line, DLD1, whereas GM-CSF treatment of de-differentiated crypt-like DLD1 and HT29 cells resulted in enhanced expression of ICAM-1. Furthermore, TNF-alpha treatment enhanced the secretion of IL-8 and GRO-alpha in HT29 cells, but not in freshly isolated IEC cultures. The differential binding and function of proinflammatory cytokines on IEC support the hypothesis that these cytokines may be involved in normal physiological processes as well as in regulating mucosal immune responses.     

Tacrine inhibits nicotinic secretory and current responses in adrenal chromaffin cells

Previous studies have demonstrated an infiltration of monocytes and increased levels of granulocyte-macrophage colony-stimulating factor (GM-CSF) in the asthmatic lung. To study the possible effects of this cytokine upon the differentiation and function of these newly recruited monocytes, we have developed a model in which monocytes isolated from human peripheral blood were differentiated into macrophages in serum in the presence or absence of GM-CSF. After 7 days, the macrophages increased in size and granularity, had increased phagocytic activity, and expressed various adhesion molecules, CD14 and major histocompatibility complex (MHC) class II. The effects of GM-CSF on antigen presentation by cultured macrophages on the antigen-specific proliferative response of CD4+ T cells to Dermatophagoides pteronyssinus or purified protein derivative of tuberculin and the mitogen phytohaemagglutinin was determined. CD4+ T-cell proliferation was reduced when either antigen was presented by macrophages cultured in serum alone, compared with the values obtained with freshly isolated monocytes. However, CD4+ cell proliferation was comparable to that observed with monocytes when antigen was presented by macrophages which had been pre-cultured with 50 U/ml GM-CSF. CD4+ T-cell proliferation to phytohaemagglutinin was similar when all three populations were used as accessory cells. High numbers of macrophages partially suppressed CD4+ T-cell proliferation in response to antigen presented by monocytes, but there was no significant difference between macrophages cultured in the presence or absence of GM-CSF. This data suggests that GM-CSF directs monocyte differentiation into macrophages with an antigen-presenting, rather than a suppressive, phenotype. Elevated levels of GM-CSF in the asthmatic lung may therefore maintain recently recruited monocytes in an inflammatory and T-cell activating state.     

Demonstration of functional CD40 in B-lineage acute lymphoblastic leukemia cells in response to T-cell CD40 ligand

Because activated T cells were previously shown to induce proliferation of human normal B-cell precursors (BCP) via the CD40 pathway, we investigated the effects of T cells on leukemic blasts isolated from patients with B-lineage acute lymphoblastic leukemia (BCP-ALL). An anti-CD3 activated human CD4+ T-cell clone was found to induce significant call proliferation in four of nine BCP-ALL samples analyzed. In one of these cases, the T-cell effect was clearly dependent on interaction between CD40 and its ligand. Accordingly, a more thorough analysis was performed on this particular leukemia (case 461, adult early pre-B-ALL, mBCR+, Philadelphia+, i(9q)+). Thus, autologous CD4+ T cells isolated from the patient were also able to induce CD40-dependent proliferation of the leukemic blasts. Analysis of the phenotype after coculture showed that, among the CD19+ cells, a proportion gradually lost expression of CD10 and CD34, whereas some cells acquired CD23. In addition, and in contrast with normal BCP, activated T cells promoted maturation of a subset of the case 461 leukemic cells into surface IgM+ cells. The leukemic origin of the cycling and the maturing cells was assessed by the presence of i(9q), a chromosomal abnormality associated with this leukemia and evidenced by fluorescence in situ hybridization. Taken together, these results show that leukemic BCP can be activated via CD40 but that not all cases display detectable stimulation in response to T cells despite their expression of CD40. In addition, the present data suggest that CD4+ T cells could potentially play a role in the physiology of BCP-ALL.     

Increased expression of costimulatory molecules on peripheral blood monocytes in patients with Crohn's disease

BACKGROUND: Activation of T lymphocytes and monocytes/macrophages has been implicated in the pathogenesis of Crohn's disease (CD). Costimulatory molecules play important roles in optimal T-cell activation. METHODS: With flow cytometric analysis we have investigated the expression of the costimulatory molecules B7-1 (CD80), B7-2 (CD86), and CD18 and the intercellular adhesion molecule-1 (ICAM-1) on peripheral blood monocytes and the expression of the activation markers HLA-DR and IL-2R (CD25) on peripheral blood T lymphocytes from 31 CD patients, 17 ulcerative colitis (UC) patients, and 10 healthy controls. RESULTS: In CD patients the percentage of activated T cells (CD3+ HLA-DR+ and CD3+ IL-2R+) was significantly increased compared with those of controls and UC patients (P < 0.05). Most monocytes from all three groups expressed B7-2, CD18, and ICAM-1 molecules (all > 79%), but only a few positive cells expressed B7-1 molecules (< 5%). No significant differences were detected in the percentage positivity of all costimulatory molecules tested among CD, UC, and controls. The mean fluorescence intensity (MFI) of B7-1 in all three groups was very weak and not significantly different. However, in CD patients there was a significantly increased MFI of B7-2, CD18, and ICAM-1 molecules compared with UC and controls (P < 0.05). On the other hand, both the percentage positivity and MFI of HLA-DR molecules on monocytes of UC patients were significantly lower than those of CD patients and controls (P < 0.05). CONCLUSIONS: Expression of the costimulatory molecules B7-2, CD18, and ICAM-1 on peripheral blood monocytes of CD patients is increased. In CD patients activation of peripheral T lymphocytes may correlate with increased expression of these costimulatory molecules on peripheral blood monocytes.     

A mucosal intranet: intestinal epithelial cells down-regulate intraepithelial, but not peripheral, T lymphocytes

Epithelial cells and lymphocytes, including gammadelta and alphabeta T cells, in the gastrointestinal tract epithelium represent a major host defense intranet that is incompletely understood. Cell-to-cell interactions between intraepithelial lymphocytes (IELs) and intestinal epithelial cells (IECs) comprise this intranet, and we have assessed the role of IECs in the regulation of gammadelta and alphabeta T cell responses. When highly purified CD3+ IEL T cells were stimulated via the TCR-CD3 complex, high proliferative responses and cytokine synthesis were induced. However, the addition of viable IECs or purified IEC membranes (mIEC) down-regulated T cell proliferative and cytokine responses. Further, the inhibitory effect of mIEC was not restored by antibodies to TGF-beta, CD1d, E-cadherin, or MHC class I or II. This inhibitory effect was noted for both gammadelta and alphabeta T cell subsets from IELs, and mRNA levels were reduced for both Th1 (IL-2 and IFN-gamma) and Th2 (IL-4 and IL-5) cytokines in gammadelta and alphabeta IELs. In contrast, a purified membrane fraction obtained from thymocytes did not inhibit IEL proliferative responses. Further, mIEC did not inhibit splenic alphabeta T cell proliferative responses. These findings show that cell-to-cell interactions between intraepithelial gammadelta and alphabeta T cells and IECs occur via cell surface molecules, suggesting an intranet to prevent potential inflammatory responses at the intestinal mucosal surface.