Expression and modulation of the human immunoglobulin A Fc receptor (CD89) and the FcR gamma chain on myeloid cells in blood and tissue.

CD89, the human immunoglobulin A (IgA) Fc receptor (FcR), is a potential target for antibody-based therapeutics, but little is known about its expression and modulation in vivo. In this study, we examined the expression pattern of CD89 and its signalling subunit, the FcR gamma chain, on circulating myeloid cells and in various tissues. Our results showed a wide tissue distribution of CD89+ cells. Thus, CD89+ cells were evident as clusters in tonsils and appendix and scattered in varying numbers in lymph nodes, kidney, liver, intestinal mucosa, bronchoalveolar lavage and peritoneal fluid. Most CD89+ cells were identified as neutrophils with high levels of CD89. A few recently emigrated macrophages (CD14low), weakly positive for CD89, were occasionally found in the tissues and more often in the peritoneal fluid. The level of CD89 on neutrophils in tissues and peripheral blood was similar, whereas on monocytes it was much lower in the tissues than in blood, and it was absent on CD14-/CD68+ intestinal lamina propria macrophages. Conversely, we detected much higher levels of the FcR gamma chain in monocytes than in neutrophils, but the FcR gamma chain was also downregulated in tissue macrophages as well as in in vitro-differentiated monocyte-derived macrophages and dendritic cells. The implications of our current findings on the biological functioning of CD89 are discussed.

Interferon-gamma-secreting T cells localize to the epithelium in coeliac disease.

Increased levels of interferon-gamma (IFN-gamma) transcripts have previously been found in duodenal biopsy specimens from patients with untreated coeliac disease (CD). Such samples and duodenal control mucosa were therefore studied to locate and phenotype cells spontaneously secreting IFN-gamma. Specimens were collected from consecutively recruited patients with untreated (seven), treated (four) or refractory (three) CD and from five histologically normal controls. Morphological and immunohistochemical examinations were performed, and epithelial and lamina propria cell suspensions were prepared from parallel samples. Unstimulated viable cells secreting IFN-gamma were identified and phenotyped with a new fluorescence-activated cell sorter-based assay, and IFN-gamma messenger RNA (mRNA) was analysed in snap-frozen aliquots of the same suspensions. Untreated CD cases had the highest fraction of IFN-gamma+ cells in the epithelial compartment (median 2.6%, range 1.6-6.2%) and, less strikingly, in the lamina propria compartment (1.6%, range 0.3-3.6%), followed by refractory (1.4%, 1.0-1.9%; and 0.3%, 0.0-1.2%) and treated (0.8%, 0.5-0.9%; and 0.7%, 0.2-1.1%) disease and finally the controls (0.5%, 0.3-0.9%; and 0.2%, 0.1-0.7%). IFN-gamma mRNA data supported these findings. IFN-gamma+ intraepithelial lymphocytes were mostly CD3+ and CD8+, whereas many positive lamina propria cells were CD8-. We conclude that isolated T cells spontaneously secreting IFN-gamma localize preferentially in the epithelium of patients with classical and refractory CD.

Heterogeneity of intraepithelial lymphocytes in refractory sprue: potential implications of CD30 expression.

BACKGROUND: Refractory sprue is defined as primary or secondary failure to respond to a gluten free diet in patients with coeliac disease-like enteropathy and may signify cryptic or overt enteropathy associated T cell lymphoma. AIMS: To study in detail jejunal morphology and immunophenotypes in patients with refractory sprue in the search for features that might be useful to predict prognosis. PATIENTS: Seven patients are described, representing all such cases identified in our hospital over a 13 year period. METHODS: Biopsy and/or surgical resection specimens were examined by morphology, immunohistochemistry, including enzymatic and immunofluorescent detection, and molecular biology. RESULTS: All patients had phenotypically abnormal intraepithelial lymphocytes (IELs) that lacked CD8, T cell receptor alpha beta (or gamma delta), and/or expressed CD30 in addition to variable expression of the natural killer cell receptor CD94. A monoclonal T cell population was present in six cases, data from the seventh being inconclusive. Three patients had overt lymphoma with CD30+ tumour tissue intervening between intact mucosa that contained neoplastic IELs. Intriguingly, CD30+ IELs were observed both a long way away from, and in direct continuity with, the tumours in these patients. Such CD30+ cells were hardly detected in patients without tumours, two of which are in good health several years after the initial diagnosis. CONCLUSIONS: Our data suggest that abnormal IELs in patients with refractory sprue are phenotypically heterogeneous. CD30 expression by these cells may indicate a worse prognosis, including the occurrence of overt lymphoma.

Age-related changes in CCR9+ circulating lymphocytes: are CCR9+ naive T cells recent thymic emigrants?

The chemokine receptor CCR9 is reported to be predominantly expressed by thymocytes as well as by circulating gut-homing and resident T cells in the small intestinal mucosa. Its ligand thymus-expressed chemokine (TECK) is produced by thymic and small intestinal epithelium. Here we report that the proportion of circulating CCR9+ naive T cells (mostly CD4+) declines with age, from approximately 15% of all T cells at birth to around 1% in adults. The proportion of CCR9+ T cells lacking the classical gut-homing receptor alpha4beta7, was much higher in children than in adults. Therefore, circulating CD3+CCR9+CD45RA+ cells have most likely left the thymus quite recently. This notion was supported by the small number of CCR9+ naive T cells which was present shortly after thymectomy. Establishing a phenotypic marker for recent thymic emigrants might provide a powerful tool in the clinical assessment and follow-up after cancer chemotherapy, hematopoietic stem cell transplantation, and during antiretroviral treatment of human immunodeficiency virus (HIV)-infected patients.

Immunoglobulin A cell distribution in the human small intestine: phenotypic and functional characteristics.

We compared B-cell phenotypes in Peyer's patches and solitary lymphoid follicles (organized gut-associated lymphoid tissue, GALT) with those in jejunal or ileal lamina propria. In situ, immunostaining showed that small B cells of naive [surface immunoglobulin D-positive (sIgD+) CD27-] and memory (sIgD+/- CD27+) phenotypes occurred almost exclusively in GALT, whereas the lamina propria contained only scattered sIgA+ CD27+ memory cells. In contrast, B-cell blasts and plasma cells negative for CD20 and often also for CD19 but with strong expression of CD38, CD27 and cytoplasmic IgA (cIgA), dominated in the lamina propria but were scarce in GALT. By flow cytometry, the proportion of dispersed CD19+ B lymphocytes varied from 4 to 42% among jejunal mucosal samples; between 5 and 50% of these were sIgD+, suggesting a variable contamination with GALT cells. B-cell blasts and plasma cells, identified by their large size and strong expression of CD38, were regularly found (25-35% of the total mononuclear cell population). Distinction between B-cell blasts and mature plasma cells was made by the presence or absence of human leucocyte antigen (HLA) class II molecules, CD45RA, CD19 and surface immunoglobulin. No CD19+ B cells outside GALT expressed CD5, but a very small portion of the lamina propria B-cell blasts were positive for CD28. Dispersed sIgA+ lamina propria cells expressed low levels of CD40, proliferated on CD40 ligation and constitutively secreted IgA in vitro. We concluded that the lamina propria B-cell compartment consists mainly of B-cell blasts and plasma cells but also has scattered, small sIgA+ cells that can proliferate in response to CD40 ligation and may therefore function as local memory cells for recall antigens.

The B-cell system of human mucosae and exocrine glands.

The mucosae and exocrine glands harbour the largest activated B-cell system of the body, amounting to some 80-90% of all immunoglobulin (Ig)-producing cells. The major product of these immunocytes is polymeric (p)IgA (mainly dimers) with associated J chain. Both pIgA and pentameric IgM contain a binding site for the polymeric Ig receptor (pIgR), or secretory component (SC), which is a requirement for their active external transport through secretory epithelia. The pIgR/SC binding site depends on covalent incorporation of the J chain into the quaternary structure of the polymers when they are produced by the local immunocytes. This important differentiation characteristic appears to be sufficient functional justification for the J chain to be expressed also by most B cells terminating at secretory effector sites with IgD or IgG production; they probably represent a "spin-off" from sequential downstream CH switching on its way to pIgA expression, thus apparently reflecting a maturational stage of effector B-cell clones compatible with homing to these sites. Observations in IgA-deficient individuals suggest that the magnitude of this homing is fairly well maintained even when the differentiation pathway to IgA is blocked. Certain microenvironmental elements such as specific cytokines and dendritic cells appear to be required for induction of IgA synthesis, but it remains virtually unknown why this isotype normally is such a dominating product of local immunocytes and why they have such a high level of J chain expression. Also, despite the recent identification of some important requirements in terms of adhesion molecules (e.g. integrin alpha 4 beta 7 and MAdCAM-1) that explain the "gut-seeking" properties of enterically induced B cells, the origin of regionalized homing of B cells to secretory effector sites outside the gut remains elusive. Moreover, little is known about immune regulation underlying the striking disparity of both the class (IgD, IgM) and subclass (IgA1, IgA2, IgG1, IgG2) production patterns shown by local immunocytes in various regions of the body, although the topical microbiota and other environmental stimuli might be important. Rational design of local vaccines will depend on better knowledge of both inductive and migratory properties of human mucosal B cells.

Severe liver failure in exertional heat stroke.

Exertional heat stroke usually occurs in warm climates. Increased serum levels of liver enzymes is a common finding in this condition, whereas liver failure is a more rare event that carries a poor prognosis. Liver transplantation has been recommended as treatment in cases of severe liver failure, but no long-term survival after this procedure in exertional heat stroke has been described. We report the case of a 31-year-old man who had a heat stroke after running 5 km at 21 degrees C. He developed severe liver damage, with serum alanine aminotransferase (ALAT) activities increasing to 16,410 U/l (reference values, 10-50 U/l) after 48 h, concomitantly with a pronounced coagulation disturbance, with Normotest (NT) decreasing to 12% (international normalized ratio (INR) = 4.2) (reference values, 70%-130% for NT and 0.8-1.2 for INR). A liver biopsy on the 5th day after the incident showed extensive liver cell necrosis. The patient was referred to be considered for liver transplantation but recovered completely on conservative treatment. We conclude that exertional heat stroke is a diagnostic possibility also in temperate climates and that severe liver failure may ensue. The liver injury is reversible, and the indications for liver transplantation in this situation have not been clarified.

Regional specialization in the mucosal immune system: primed cells do not always home along the same track.

According to the current paradigm of lymphocyte trafficking, primed B and T cells extravasate in the intestinal lamina propria chiefly by means of the mucosal homing receptor alpha4beta7, which interacts with the vascular addressin MAdCAM-1. However, as discussed here, this mechanism cannot explain the preferential homing of B cells with a high level of J-chain expression to mucosal effector sites outside the gut.

Regional specialization in the mucosal immune system: what happens in the microcompartments?

Mucosal immunity is an important arm of the immune system because it operates in tissues involved in everyday infectious defence as well as in tolerance against innocuous environmental and dietary antigens. Here, Per Brandtzaeg and colleagues discuss compartmentalized regulation of mucosal B cells and mechanisms that might explain the strikingly regionalized effector disparity of the human mucosal immune system.

Phenotypic distribution of T cells in human nasal mucosa differs from that in the gut.

Phenotypic and functional studies are required to understand the immunoregulatory role of mucosal T cells. Information about T cells in the human upper respiratory tract is limited and conflicting. Therefore, we phenotyped T cells in nasal mucosa by means of multicolor in situ immunofluorescence. In normal mucosa, most CD3+ intraepithelial lymphocytes (IELs) and lamina propria lymphocytes (LPLs) (> 90%) expressed T-cell receptor (TCR)alpha/beta, and only approximately 5% expressed TCRgamma/delta. Although most IELs in the surface epithelium were CD8+ (64%), many expressed CD4 (30%) and the CD4 phenotype dominated (55%) only slightly in the lamina propria. This result was strikingly different from that obtained for comparable compartments in histologically normal jejunal mucosa, where IELs consisted of 83% CD8+ and LPLs of 73% CD4(+) T cells. Nasal CD3+ IELs and LPLs were mainly CD45RO+CD45RA- and usually expressed CD7. The integrin alphaEbeta7 was, as expected, more common on IELs than on LPLs (78 versus 20%). In conclusion, nasal T cells show several similarities to those of the normal jejunum but some notable differences exist, especially a relative increase in CD4+ T cells in the epithelium and a decrease in the lamina propria. It should be explored whether this disparity, together with an increased expression of epithelial adhesion molecules, might contribute to local immunological overstimulation and partly explain the relatively high frequency of airway allergy.

Major histocompatibility complex class II-dependent antigen presentation by human intestinal endothelial cells.

BACKGROUND & AIMS: In the normal gut, human intestinal microvascular endothelial cells (HIMECs) express major histocompatibility complex (MHC) class II molecules. Enhanced expression is found in chronic inflammation. We examined the cytokine regulation of MHC class II molecules and the associated invariant chain (Ii) in HIMECs and investigated whether such cells can process and present a complex protein antigen to T cells. METHODS: Enzyme-linked immunosorbent assay, flow cytometry, immunoelectron microscopy, as well as T-cell activation assay with HIMECs and HLA-DR-restricted T-cell clones were employed. RESULTS: In unstimulated HIMEC monolayers, HLA-DR, -DP, and -DQ and Ii were undetectable at the protein level, but interferon gamma (IFN-gamma) (100 U/mL) induced expression that peaked for DR after 2-3 days, for DP after 4-6 days, for DQ after 10-12 days, and for Ii after 2-3 days. Tumor necrosis factor alpha had no effect alone but enhanced class II expression in combination with IFN-gamma, most notably for DQ and DP. HLA-DR3-restricted and Mycobacterium tuberculosis heat shock 65-kilodalton-specific T-cell clones were activated to produce IFN-gamma in response to relevant antigen presented by IFN-gamma-treated HIMECs. This response was inhibited by blocking monoclonal antibody to HLA-DR and by chloroquine when compared to professional antigen-presenting cells, HIMECs activated T-cell clones quite efficiently. CONCLUSIONS: These data suggest that microvascular endothelial cells can present complex protein antigens in the human gut.

Cellular and molecular mechanisms for induction of mucosal immunity.

The epithelial glycoprotein called secretory component (SC) is quantitatively the most important receptor of the immune system because it is responsible for external transport of locally produced polymeric IgA (pIgA) to generate remarkably large amounts of secretory IgA. Antibodies of this type constitute the major mediators of specific humoral immunity. Transmembrane SC belongs to the Ig supergene family and functions as a common pIg receptor, also translocating pentameric IgM externally to form secretory IgM. The B cells responsible for mucosal production of Ig polymers are initially stimulated in organized mucosa-associated lympho-epithelial structures, particularly the Peyer's patches in the distal small intestine; from these inductive sites they migrate ("home") as memory cells to exocrine tissues all over the body. Mucous membranes are thus furnished with secretory antibodies in an integrated way, ensuring a variety of specificities at every secretory effector site. There is currently great interest in exploiting this integrated or "common" mucosal immune system for oral vaccination against pathogenic infectious agents and also to induce tolerance in T cell-mediated auto-immune diseases. However, much remains to be learned about mechanisms for antigen uptake and processing necessary to elicit stimulatory or suppressive mucosal immune responses in humans. Moreover, evidence is emerging for the existence of considerable regionalization with regard to functional links between inductive sites and effector sites of mucosal immunity.

A leiomyoma of the iris documented by immunohistochemistry and electron microscopy.

PURPOSE: To report an iris leiomyoma documented in accordance with today's stricter diagnostic criteria. METHODS: Light microscopy, immunohistochemistry and electron microscopy were performed on the 3 mm ball-like, greyish-white vascularized tumour of the iris, close to the sphincter pupillae. RESULTS: Light microscopy of the extirpated neoplasm showed the characteristic appearance of a leiomyoma with densely packed spindle-shaped cells, with oval nuclei and granular cytoplasm. On electron microscopic examination the tumour exhibited the characteristic features of a smooth muscle neoplasm. The immunohistochemistry was consistent with a myogenic tumour because the tumour cells were positive for smooth muscle actin and desmin, but negative for S-100 and melanin. CONCLUSION: The case illustrated the necessity of performing ancillary procedures such as electron microscopy and immunohistochemistry to substantiate a correct, although rare diagnosis.

Mucosal immunity--a major adaptive defence mechanism.

The epithelial glycoprotein called secretory component (SC) is quantitatively the most important receptor of the immune system because it is responsible for external transport of locally produced polymeric IgA (pIgA) to generate remarkably large amounts of secretory IgA. Antibodies of this type constitute the major mediators of specific humoral immunity. Transmembrane SC belongs to the Ig supergene family and functions as a common pIg receptor, also translocating pentameric IgM externally to form secretory IgM. The B cells responsible for mucosal pIg production are initially stimulated in organized mucosa-associated lymphoepithelial structures, particularly the Peyer's patches in the distal small intestine; from these inductive site they migrate as memory cells to exocrine tissues all over the body. Mucous membranes are thus furnished with secretory antibodies in an integrated way, ensuring a variety of specificities at every secretory effector site. There is currently great interest in exploiting this integrated or "common" mucosal immune system for oral vaccination against pathogenic infectious agents and also to induce tolerance in T cell-mediated autoimmune diseases. However, much remains to be learned about mechanisms for antigen uptake and processing necessary to elicit stimulatory or suppressive mucosal immune responses. Moreover, evidence is emerging for the existence of considerable regionalization with regard to functional links between inductive sites and effecter sites of mucosal immunity.

Phenotypes of B and T cells in human intestinal and mesenteric lymph.

BACKGROUND & AIMS: Cells in lymph draining the human gut have not been characterized previously. The aim of this study was to phenotype B and T cells present in microlymphatics of Peyer's pathces and in mesenteric lymph. METHODS: The studies were conducted by multicolor immunohistochemistry, flow cytometry, and immunocytochemistry. RESULTS: In decreasing order of frequency, microlymphatics in Peyer's patches contained naive T (CD3+CD45RA+ alpha 4 beta 7low) and B (sIgD+CD20+ alpha 4 beta 7low) lymphocytes, memory T (CD45RO+ alpha 4 beta 7+) and B (sIgD-CD20+ alpha 4 beta 7+) lymphocytes, and B-cell blasts (CD19+CD38high alpha 4 beta 7high). Naive cells were usually positive for L-selectin, memory cells were either positive or negative, and B-cell blasts were usually negative. Mesenteric lymph contained naive T (approximately 60%) and B (approximately 25%) lymphocytes, memory T and B lymphocytes (approximately 10%), and B-cell blasts (approximately 2%). Cytospins confirmed these results and showed, in addition, that B-cell blasts contained cytoplasmic immunoglobulin (Ig) A, IgM, or IgG in overall proportions of 5:1: < 0.5. CONCLUSIONS: Our results are similar to the phenotypes previously described in animal thoracic or mesenteric lymph. A fraction of the B cells stimulated in Peyer's patches are near terminal differentiation (contain cytoplasmic Ig) before they enter peripheral blood. Many memory cells, and most if not all B-cell blasts entering lymph show an adhesion molecule profile (alpha 4 beta 7high L-selectin low) in keeping with the presumed phenotype of lymphoid cells destined for mucosal effector sites such as the gut lamina propria.

Topographic distribution of homing receptors on B and T cells in human gut-associated lymphoid tissue: relation of L-selectin and integrin alpha 4 beta 7 to naive and memory phenotypes.

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.