Mouse CD146/MCAM is a marker of natural killer cell maturation

CD146/melanoma cell adhesion molecule is an adhesion molecule expressed by endothelial cells and by a small fraction of activated T and B lymphocytes in humans. In order to analyze the pattern of CD146 expression in mouse leukocytes at steady-state conditions, we generated a set of novel rat anti-mouse CD146 monoclonal antibodies. CD146 expression was undetectable on monocytes, dendritic cells, T cells or B cells, but was expressed on about 30% of neutrophils and 60% of NK cells. Within murine lymphocytes, CD146 was defined as a novel NK-specific surface molecule. An increased percentage of CD146+ cells was found in the most mature CD27(-)CD11b+ NK cell subpopulation, which also displays higher expression of Ly49C/I, Ly49D and KLRG1 and lower expression of NKG2A/C/E molecules. CD146+ NK cells were found to be less cytotoxic and produce less IFN-gamma than CD146(-) NK cells upon stimulation with target cells or activating antibodies. These findings define CD146 as a marker of mouse NK cell maturation that may be used as an alternative to the combined use of CD27 and CD11b staining to detect final stages of NK cell maturation.     

Pre-eclampsia is associated with the failure of melanoma cell adhesion molecule (MCAM/CD146) expression by intermediate trophoblast

The purpose of this study was to determine whether the expression of melanoma cell adhesion molecule (MCAM/CD146), like most other endothelial adhesion molecules, is reduced in the pregnancy disorder pre-eclampsia, and whether it is associated with the invasiveness of trophoblast. We used immunohistochemical approaches and analyzed 42 placentas from control and pre-eclamptic patients using an anti-CD146 monoclonal antibody. Our data show that in normal placentas, CD146 staining was specifically detected in intermediate trophoblasts that are most invasive and migratory, but not detected in noninvasive cytotrophoblasts or syncytiotrophoblasts. However in pre-eclampsia, CD146 was no longer present in the intermediate trophoblasts, although it was detectable in the blood vessels. At the same time, we tested CD31/PECAM-1, another endothelial adhesion molecule, and found its negative staining for all kinds of trophoblasts in both normal and pre-eclamptic placentas. The different staining patterns of CD146 in the normal and pre-eclamptic placentas provide the first evidence that in pre-eclampsia, intermediate trophoblasts fail to express CD146, implicating that CD146 plays an important role in trophoblast invasion.     

Relationship of CD146 expression to secretion of interleukin (IL)‐17, IL‐22 and interferon‐γ by CD4+ T cells in patients with inflammatory arthritis

Expression of the adhesion molecule, CD146/MCAM/MelCAM, on T cells has been associated with recent activation, memory subsets and T helper type 17 (Th17) effector function, and is elevated in inflammatory arthritis. Th17 cells have been implicated in the pathogenesis of rheumatoid arthritis (RA) and spondyloarthritides (SpA). Here, we compared the expression of CD146 on CD4⁺ T cells between healthy donors (HD) and patients with RA and SpA [ankylosing spondylitis (AS) or psoriatic arthritis (PsA)] and examined correlations with surface markers and cytokine secretion. Peripheral blood mononuclear cells (PBMC) were obtained from patients and controls, and synovial fluid mononuclear cells (SFMC) from patients. Cytokine production [elicited by phorbol myristate acetate (PMA)/ionomycin] and surface phenotypes were evaluated by flow cytometry. CD146⁺CD4⁺ and interleukin (IL)‐17⁺CD4⁺ T cell frequencies were increased in PBMC of PsA patients, compared with HD, and in SFMC compared with PBMC. CD146⁺CD4⁺ T cells were enriched for secretion of IL‐17 [alone or with IL‐22 or interferon (IFN)‐γ] and for some putative Th17‐associated surface markers (CD161 and CCR6), but not others (CD26 and IL‐23 receptor). CD4⁺ T cells producing IL‐22 or IFN‐γ without IL‐17 were also present in the CD146⁺ subset, although their enrichment was less marked. Moreover, a majority of cells secreting these cytokines lacked CD146. Thus, CD146 is not a sensitive or specific marker of Th17 cells, but rather correlates with heterogeneous cytokine secretion by subsets of CD4⁺ helper T cells.     

Modulation of intercellular adhesion molecule 1 (ICAM-1) expression on the human mast-cell line (HMC)-1 by inflammatory mediators

The effect of inflammatory mediators on the expression of several surface adhesion molecules on the human mast-cell line (HMC)-1 was studied. By flow cytometry, it could be shown that among several surface adhesion molecules (ICAM-UCDS4, VLA-4/CD49d, Mac-UCD11b, LFA-1/CD11a, LFA-2/CD2, LFA-3/CDS8, VCAM-1), only the constitutively expressed immunoglobulin family member intercellular adhesion molecule-1 (ICAM-1) is modulated by proinflammatory cytokines on HMC-1 mast cells. Stimulation with tumor necrosis factor-a (TNF-α) and interferon-γ (IFN-γ) resulted, in addition to interleukin-(lL-)4, in selective upregulation of ICAM-1 expression. Costimulation of either IL-4 or IFN-γ with TNF-α further increased the ICAM-1 expression as compared to the stimuli alone. In contrast, stem-cell factor (SCF), granulocyte/macrophage colonystimulating factor (GM-CSF), IL-10, IL-8, monocyte chemotactic and activating factor (MCAF), and the complement split product C5a failed to modulate the expression of any adhesion molecule examined. The levels of cytoplasmic free calcium in HMC-1 mast cells were not altered by cross-linking surface ICAM-1, suggesting linkage of other intracellular signaling pathways. This cytokine-induced upregulation of ICAM-1 expression might reveal a putative regulatory mechanism of mast-cell interaction with effector cells bearing the counterparts of ICAM-1 (CD54), the molecules Mac-1 (CD11b/CD18) and leukosialin (CD43), and the principal ligand LFA-1 (CD11a/CD18).     

The stromal cell antigen CD248 (endosialin) is expressed on naive CD8+ human T cells and regulates proliferation

CD248 (endosialin) is a transmembrane glycoprotein that is dynamically expressed on pericytes and fibroblasts during tissue development, tumour neovascularization and inflammation. Its role in tissue remodelling is associated with increased stromal cell proliferation and migration. We show that CD248 is also uniquely expressed by human, but not mouse (C57BL/6), CD8(+) naive T cells. CD248 is found only on CD8(+) CCR7(+) CD11a(low) naive T cells and on CD8 single-positive T cells in the thymus. Transfection of the CD248 negative T-cell line MOLT-4 with CD248 cDNA surprisingly reduced cell proliferation. Knock-down of CD248 on naive CD8 T cells increased cell proliferation. These data demonstrate opposing functions for CD248 on haematopoietic (CD8(+)) versus stromal cells and suggests that CD248 helps to maintain naive CD8(+) human T cells in a quiescent state.     

活化白细胞黏附分子及其与肿瘤侵袭转移的关系

ALCAM主要表达在白细胞和胸腺上皮细胞,属于免疫球蛋白基因超家族的成员,是淋巴细胞抗原CD6的配基。ALCAM/CD166在多种肿瘤细胞表面高表达,参与肿瘤的发生发展。已通过体外试验和动物模型证实ALCAM/CD166对多种肿瘤生长和转移相关的肿瘤细胞特性具有调节作用;在人类肿瘤,ALCAM/CD166与乳腺癌、前列腺癌、食道癌、结肠癌及恶性黑色素瘤等肿瘤患者的生存率及预后密切相关。     

Changes in expression of the cell adhesion molecule PECAM-1 (CD31) during differentiation of human leukemic cell lines

Abstract: PECAM-1, a member of the immunoglobulin gene superfamily, is widely distributed on cells of the vascular system, and mediates cellular interactions through both homotypic and heterotypic adhesive mechanisms. Previous studies have demonstrated that PECAM-1 is initially expressed at high levels on CD34⁺ multipotential progenitors in the bone marrow, but is subsequently downregulated in more committed precursors of all lineages. Interestingly, although PECAM-1 expression is high on circulating monocytes and neutrophils, little is known about the upregulation of PECAM-1 expression during terminal myelomonocytic differentiation. We have further characterized this process by examining PECAM-1 expression during chemically-induced differentiation of the U937, HL-60 and HEL cell lines. Quantitative Western blot analysis of cellular lysates indicated that PECAM-1 expression could be upregulated in U937 and HL-60 cells by phorbol esters or dimethyl sulfoxide. Northern blot analysis showed that PECAM-1 mRNA levels appeared to increase in parallel with that of PECAM-1 protein. We also observed a marked difference in the apparent molecular mass of PECAM-1 that was lineage-specific, both in differentiated leukemic cell lines and in their corresponding leukocyte population. Immunofluorescence localization indicated that the cellular distribution of PECAM-1 in U937 and HL-60 cells was similar to that of their normal circulating counterparts, and that the pattern of distribution again displayed lineage fidelity. The ability to induce the expression of PECAM-1 molecules having different glycosylation and surface expression patterns may prove useful for further elucidation of the role of PECAM-1 in hematopoiesis, as well as studies of the cell lineage-specific modulation of PECAM-1 function.     

Prevention of adjuvant arthritis by the W3/25 anti-CD4 monoclonal antibody is associated with a decrease of blood CD4(+)CD45RC(high) T cells

Imbalance between Th1 and Th2 functions is considered to play a key role in the induction and development of several autoimmune diseases, and the correction of that imbalance has led to effective therapies of some experimental pathologies. To examine whether CD4(+)CD45RC(high) (Th1-like) and CD4(+)CD45RC(low) (Th2-like) lymphocytes play a role in the pathogenesis of adjuvant arthritis (AA) and in its prevention by anti-CD4 antibody, CD45RC expression on CD4(+) T cells was determined in arthritic rats and in animals treated with an anti-CD4 MoAb (W3/25) during the latency period of AA. The phenotype of regional lymph node lymphocytes from arthritic rats in the active phase of the disease was determined by flow cytometry. Peripheral blood lymphocytes from rats treated with W3/25 MoAb were also analysed for 2 weeks after immunotherapy finished. IgG2a and IgG1 isotypes of sera antibodies against the AA-inducing mycobacteria, considered to be associated with Th1 and Th2 responses, respectively, were also determined by ELISA techniques. Fourteen days after arthritis induction, regional lymph nodes presented an increase in CD4+CD45RC(high) T cell proportion. Preventive immunotherapy with W3/25 MoAb inhibited the external signs of arthritis and produced a specific decrease in blood CD4(+)CD45RC(high) T cells and a diminution of antibodies against mycobacteria, more marked for IgG2a than for IgG1 isotype. These results indicate a possible role of CD4(+)CD45RC(high) T lymphocytes in the pathogenesis of AA, and suggest that the success of anti-CD4 treatment is due to a specific effect on CD4(+)CD45RC(high) T subset that could be associated with a decrease in Th1 activity.     

Platelet-endothelial cell adhesion molecule-1 (CD31) redistributes from the endothelial junction and is not required for the transendothelial migration of melanoma cells

We have examined the role of platelet-endothelial cell adhesion molecule-1 (PECAM-1/CD31) during the transendothelial migration of melanoma cells using a novel in vitro system. Comparable studies have suggested the involvement of PECAM-1 in leukocyte transendothelial migration. Such studies have been confirmed using in vivo models of inflammation. These studies prompted us to examine the role of PECAM-1 in tumor cell transendothelial migration. Anti-PECAM-1 monoclonal antibodies, known to block leukocyte transendothelial migration, were tested in co-cultures of human melanoma cells seeded on a monolayer of human lung microvascular endothelial cells. None of these antibodies inhibited the transmigration of melanoma cells. Moreover, confocal microscopy revealed the dissolution of the PECAM-1 adhesion complexes in the endothelial junctions associated with melanoma cells and the lack of PECAM-1 in heterotypic contacts between transmigrating melanoma cells and adjacent endothelial cells. These data, therefore, indicate that PECAM-1 is not required for the transendothelial migration of melanoma cells. This revised version was published online in July 2006 with corrections to the Cover Date.     

Increased adhesion of human monocytes to IL-4-stimulated human venous endothelial cells via CD11/CD18, and very late antigen-4 (VLA-4)/vascular cell adhesion molecule-1 (VCAM-1)-dependent mechanisms.

Expression of adhesion molecules on endothelial cells (EC) can be up-regulated or induced by cytokines. The aim of the present study was to investigate the effect of IL-4 on both the expression of adhesion molecules on EC and monocyte adhesion to EC. Flow cytometric analysis showed that VCAM-1 expression on EC was up-regulated after stimulation with IL-4 for 24 h, whereas the expression of E-selectin (formerly called endothelial leucocyte adhesion molecule-1 (ELAM-1)) was not enhanced, and that of intercellular adhesion molecule-1 (ICAM-1) only slightly. The adhesion of monocytes to EC increased to maximum values upon stimulation of EC with IL-4 for 24 h. Coating of monocytes with MoAb against the integrin beta 2-subunit (CD18) significantly inhibited their adhesion to IL-4-stimulated EC; maximal inhibition was found when monocytes were coated with anti-CD18 MoAb in combination with MoAb against CD49d (the alpha-chain of VLA-4), whereas no inhibition was found when monocytes were coated only with MoAb against CD49d. Monocyte adhesion was not significantly inhibited when IL-4-stimulated EC were coated with MoAbs against ICAM-1 or VCAM-1 alone or in combination. Adhesion of monocytes was inhibited to a greater extent when in addition to coating of monocytes with MoAb against CD18 the EC were coated with MoAb against VCAM-1. From these results we conclude that monocytes bind to IL-4-stimulated EC via interaction of CD11/CD18 molecules on the monocytes with an as yet unknown endothelial ligand, and interaction of VLA-4 on monocytes with VCAM-1 on EC.     

Characterization of ''adult-type'' mast cells derived from human bone marrow CD34+ cells cultured in the presence of stem cell factor and interleukin-6. Interleukin-4 is not required for constitutive expression of CD54, FcεRIα and chymase, and CD13 expression is reduced during differentiation

BACKGROUND: In vitro-derived human mast cells exhibit different properties, depending in part on the source of progenitor cells. Most investigations have used fetal liver, cord blood or peripheral blood. Few have used adult bone marrow. OBJECTIVE: Human mast cells derived in vitro from the CD34(+) progenitors in bone marrow and cord blood that had been cultured with recombinant human stem cell factor (rhSCF) and recombinant human interleukin-6 (rhIL-6) were compared. METHODS AND RESULTS: After 12 weeks of culture, nearly all of the cells were mast cells, and nearly all of these had cytoplasmic granules containing both tryptase and chymase (MCTC type), stained metachromatically with acidic toluidine blue, and expressed CD117 on the cell surface. Both tryptase protein and mRNA were detected by two weeks of culture. Chymase mRNA and protein were detected at 4 weeks but not at 2 weeks of culture. By 12 weeks, chymase content per cell, measured by ELISA, was significantly higher (P < 0.05) in human bone marrow-derived mast cells (HBMMC) (5.6 +/- 0.9 pg) than in cord blood-derived mast cells (CBMC) (2.4 +/- 0.9 pg), whereas histamine and tryptase levels were not significantly different. Of the cluster designations tested, CD29, CD49d, CD51 and CD61 were strongly expressed on HBMMC. CD54 and Fc epsilon RI alpha also were expressed constitutively. Approximately half of CD34-sorted cells at day 0 were CD13(+) and this diminished as mast cell maturation occurred. Electron microscopy revealed that 12-week-old HBMMC had many secretory granules that contained spherical electron dense cores surrounded by electron lucent space, consistent with previous reports of immature MCTC cells developing in vivo. CONCLUSIONS: CD34(+) progenitors of human bone marrow are a rich source of mast cell progenitors capable of expressing granule and surface markers of mature mast cells in the presence of rhSCF and rhIL-6.     

CD20 monoclonal antibodies decrease interleukin-4-stimulated expression of the low-affinity receptor for IgE (FcϵRII/CD23) in human B cells by increasing the extent of its cleavage

CD20 monoclonal antibody (mAb) B1 is known to inhibit B cell proliferation. We show that B1 reduced both anti-μ + interleukin-4 (IL-4)-induced DNA synthesis and the concomitant expression of CD23 at the surface of human tonsillar B cells. B1 mAb had no effect on CD23 mRNA levels. The disappearance of CD23 molecule from the surface correlates with an increase of soluble CD23 fragments in the culture medium, indicating that CD20 mAb B1 stimulated the cleavage of the molecule. B1 also inhibits IgE production by peripheral blood mononuclear cells cultured in the presence of IL-4. Suppression of IgE synthesis and enhancement of CD23 cleavage are concomitant but appear not to be functionally related.     

Tumour necrosis factor-alpha augments the expression of Fc IgE receptor (Fc epsilon RII/CD23) on human monocytic cell lines and down-regulates interleukin-4-driven Fc epsilon RII expression on monocytes.

We investigated the expression of the low affinity Fc IgE receptor (Fc epsilon RII/CD23) on the human monocytic cell lines U937, THP-1, Mono-Mac-6, and cultured human peripheral blood monocytes under stimulation with human tumour necrosis factor-alpha (TNF-alpha) and other cytokines. Fc epsilon RII was demonstrated by flow cytometry analysis employing the anti-Fc epsilon RII monoclonal antibody 3-5. TNF-alpha alone had a weak but significant stimulating effect on the Fc epsilon RII expression on the cell lines U937 and THP-1, and very modestly on Mono-Mac-6 cells. TNF-alpha strongly synergized with interferon-gamma (IFN-gamma) and interleukin-4 (IL-4). IFN-alpha per se was ineffectual, but was able to increase the TNF-alpha effect. Furthermore, the action of TNF-alpha was slightly augmented by human IL-6. Similar effects were noted with TNF-beta alone or in combination with other cytokines. Interestingly, on human monocytes TNF-alpha weakly reduced the basal level of Fc epsilon RII, and markedly diminished the IL-4-induced Fc epsilon RII expression. Our results indicate that several cytokines may interact in a cytokine network to modulate Fc epsilon RII expression on monocytic cell lines. On human blood monocytes, TNF-alpha, like IFN-gamma or IL-6, counteracts the IL-4-induced Fc epsilon RII expression. These data suggest different regulatory pathways of Fc epsilon RII expression on blood monocytes and myelomonocytic cell lines.