The expression of T cell related costimulatory molecules in multiple myeloma

Presentation of tumour antigen by malignant cells not expressing costimulatory molecules is considered to be a major cause of the failure of the host's immune response against tumours. This study has determined the expression of the B7 family of costimulatory molecules on malignant plasma cells and the expression of the counter receptor molecules, CD28 and CD152 (CTLA-4), on T cells of patients with multiple myeloma. CD28 expression was present on most CD4 cells but was lower on CD8 cells especially from those patients who also showed evidence of expanded T cell clones (median 40%. z=2.4; p<0.02). CD152 expression was increased in 50% (9/18) of patients with myeloma. CD80 (B7-1) expression was present on the plasma cells of only 1 of 27 samples but CD86 (B7-2) expression within the normal range was present on the plasma cells of 14 of 27 samples. Primitive plasma cells (CD38++ CD45++) had a higher expression of CD86 (median 78%) than mature plasma cells (CD38++ CD45-) (median 19%, z=3.7; p<0.01). Thus patients with expanded T cell clones have a downregulated T cell CD28 expression and lack B7-1 expression on their malignant plasma cells. These results are consistent with the concept that engagement of the T cell receptor by tumour antigen on B7-1 deficient malignant plasma cells would result in T cell anergy rather than productive immunity.     

Variability in cytokine and cell adhesion molecule staining in arthroscopic synovial biopsies: quantification using color video image analysis

OBJECTIVE: To investigate the variability in immunostaining for cytokines and cell adhesion molecules using multiple arthroscopically directed synovial biopsies from within a rheumatoid knee joint, quantitated by color video image analysis. METHODS: Needle arthroscopic biopsies were taken from multiple sites (4-7 sites) around a knee joint in 8 patients with rheumatoid arthritis (RA). In 5 patients, immunoperoxidase staining for the cytokines tumor necrosis factor alpha (TNF-alpha), interleukin 8 (IL-8), and IL-1beta as well as the IL-1 receptor antagonist protein (IL-1ra) was performed. In 3 patients, immunoperoxidase staining for the cell adhesion molecules E-selectin (CD62E), P-selectin (CD62P), intercellular adhesion molecule 1 (ICAM-1, CD54), and platelet endothelial cell adhesion molecule (PECAM, CD31) was performed. Immunostaining was quantified using color video image analysis. RESULTS: The overall probability of paired biopsies from the same RA knee joint being significantly different from each other due to sampling variation was at most 22% for cytokine staining (usually less than 10%). There were no significant differences between intrabiopsy and interbiopsy variability for cell adhesion molecule staining of the sublining and vessels. CONCLUSION: The variability in cytokine and cell adhesion molecule staining within any single biopsy usually reflects the variability between biopsies taken from different sites in the same rheumatoid joint when the immunostaining is quantified using color video image analysis. Therefore, only a small number of synovial biopsies are required to accurately determine the cytokine and cell adhesion molecule expression in a single joint.     

Recent developments in peroxisome biology

Membrane molecules such as CD36 (OKM5), intercellular adhesion molecule-1 (ICAM-1, CD54), gamma interferon-induced protein 10 (gamma-IP10) and IL-1 are induced and/or upregulated in psoriatic epidermis. These molecules have important accessory, trafficking or signalling functions in the immune system and also play a role in the pathophysiology of psoriasis. The relevance of adhesion molecules, CD36 and epidermal IL-1 in psoriasis was studied in vitro in the autologous mixed epidermal cell - T lymphocyte reaction (MECLR). Their level of expression was quantitated in epidermal cell suspensions (ECS) from patients with psoriasis and their function was assessed by blocking with specific mAbs and antisera or by depleting CD36+ cells from the ECS prior to the MECLR. ECS from psoriatic lesions contained increased numbers of CD36+ (23 +/- 12%), ICAM-1(+) (31 +/- 14%) and IL-1(+) (57 +/- 21%) cells. The autologous MECLR was inhibited in samples from all patients by mAb to CD2 (LFA-2), CD11a (LFA-1alpha), CD18 (LFA-1beta), ICAM-1, CD58 (LFA-3) and an antiserum to IL-1beta. Thus, adhesion molecules facilitate inflammation in psoriasis not only via adhesion and recruitment of T lymphocyte in psoriatic lesions, but also via activation of T cells. Furthermore CD36 molecules on psoriatic epidermal cells do not costimulate autologous T lymphocytes in psoriasis. The observed costimulatory function of IL-1beta in the MECLR emphasizes its relevance in psoriasis.     

Non-MHC-restricted cell-mediated lysis of human oligodendrocytes in vitro: relation with CD56 expression

Oligodendrocytes and their myelin membranes are the apparent target of the autoimmune response that characterizes the human adult central nervous system-demyelinating disease multiple sclerosis. Human oligodendrocytes do not express MHC class II molecules, a requirement for MHC-restricted injury mediated by myelin-reactive CD4+ T cells, the cell type implicated in initiating the disease process. In this study we observed that human adult central nervous system-derived oligodendrocytes can be susceptible to non-MHC-restricted lysis mediated by myelin basic protein-reactive CD4+ T cell lines. Cytotoxicity was significantly greater (37 +/- 4 vs 7 +/- 3%) with cell lines in which a high proportion of cells (mean, 28 +/- 6%) expressed CD56 compared with cytotoxicity mediated by low CD56 cell lines (8 +/- 2%). High CD56 cell lines, when rested in IL-2, lost cytotoxic activity and had reduced expression of CD56 (mean, 5 +/- 2%). CD4+ T cells isolated from short term (3-day) anti-CD3/IL-2-activated mononuclear cell cultures did not express CD56 and were not cytotoxic to oligodendrocytes unless lectin was added. In contrast, an enriched population of non-T cells extracted from the same activated MNC cultures expressed CD56 as well as other NK cell-associated surface molecules and was cytotoxic. These results indicate the potential susceptibility of human oligodendrocytes to non-MHC-restricted injury mediated by both Ag-reactive and nonspecific cellular immune effector cells, with CD56 expression being a common feature of the effector cells.     

Effect of rifampin on CD1b expression and double-negative T cell responses against mycobacteria-derived glycolipid antigen

Non-classical antigen-presentation by CD1 molecules expressed on cytokine-activated monocytes (CAM), and cell-mediated responses supported by double-negative (DN) and by CD8+ responder alphabeta T cells, are involved in host resistance against mycobacterial infections. The CD1b protein is responsible for presentation of non-peptide, lipid antigens to T cells. In this context, a pivotal role is played by induction of CD1b protein on the membrane of human monocytes activated by GM-CSF alone, and more efficiently by GM-CSF combined with IL-4. Rifampin (RFP), a drug which is extensively utilized for chemoprophylaxis or treatment of Mycobacterium tuberculosis, is known to reduce a number of B, or T cell-dependent responses. Therefore we undertook immunopharmacological studies on RFP, to determine the effects of this agent on human macrophage function, relative to antigen presentation by CD1b molecules and on DN T cell cytolytic function. The results showed that: (a) graded concentration of RFP (2 or 10 microg/ml) induced a significant increase of CD1b expression, in CAM as evaluated by FACS analysis; (b) RFP increased significantly the specific mAb binding to CD1b on CAM surface; (c) treatment of effector cells with RFP did not reduce DN T cell-mediated cytolysis against lymphoblastoid cells transfected with CD1b cDNA (C1R.b6 cells), pulsed with M. tuberculosis. These results suggest that RFP could be of potential value in improving mycobacterial antigen presentation without impairing responder T cell function.     

Characterization of monoclonal antibodies that recognize canine CD34

Using a polyclonal antiserum against canine CD34, we previously found that CD34 is expressed on canine bone marrow progenitor cells in a manner analogous to that found in humans. To further characterize CD34+ cells and to facilitate preclinical canine stem cell transplant studies, monoclonal antibodies (MoAbs) were raised to CD34. A panel of 10 MoAbs was generated that reacted with recombinant CD34 and with CD34+ cell lines and failed to react with CD34- cell lines. Binding properties of five purified MoAbs were determined by BIAcore analysis and flow cytometric staining, and several MoAbs showed high affinity for CD34. Two antibodies, 1H6 and 2E9, were further characterized, and in flow cytometry studies typically 1% to 3% of stained bone marrow cells were CD34+. Purified CD34+ bone marrow cells were 1.8- to 55-fold enriched for colony-forming unit-granulocyte-macrophage and for long-term culture initiating cells as compared with bone marrow mononuclear cells, whereas CD34- cells were depleted of progenitors. Three autologous transplants were performed with CD34+ cell fractions enriched by immunomagnetic separation. After marrow ablative total body irradiation (920 cGy), prompt hematopoietic recovery was seen with transplanted cell doses of 相似文献   

Immunohistochemical detection of tumor cells in the bone marrow of breast cancer patients

BACKGROUND: Contamination of bone marrow and peripheral blood stem cells with tumor cells is a problem that may be encountered when autologous hematopoietic stem cell transplantation is conducted concurrently with high-dose chemotherapy. METHODS: Using monoclonal antibodies to a variety of tumors, the detection of tumor cells in the bone marrow of breast cancer patients was studied by immunohistochemistry. RESULTS: KL-1 and CAM5.2 were strongly reactive with breast cancer cells, but not with normal bone marrow cells. The reactivity of the tumor cells with EMA was not strong, and DF-3 and 115D8 yielded only slightly positive reactions. These latter antibodies also exhibited some reactivity to normal bone marrow cells. When tumor cells were admixed with normal cells, the sensitivity of CAM5.2 and EMA permitted the detection of one cell in 10(4), but with KL-1, the detection of one in 10(5) cells was possible. When immunohistochemical staining was used in testing 40 patients with advanced or recurrent breast cancer, positive reactions were obtained in four of 27 patients (14.8%) with KL-1, four of 26 (15.4%) with CAM5.2, and nine of 37 (23.7%) with KL-1 + CAM5.2, figures similar to those reported by others who studied stage IV patients. CONCLUSIONS: Immunohistochemical staining with KL-1 and CAM5.2 is therefore considered to be a useful technique for detecting contamination by tumor cells.     

The effects of a community-based pulmonary rehabilitation programme on exercise tolerance and quality of life: a randomized controlled trial

The molecules B7.1 and B7.2 deliver costimulatory signals of critical importance to naive T cells, and may thus be involved in abrogation of oral tolerance in IBD. Functional disparity apparently exists among antigen-presenting cells in vivo. We wanted to examine if differential B7 expression occurs on mucosal macrophage subsets. Cryosections of bowel specimens from patients with IBD and normal controls were subjected to immunofluorescence and immunoperoxidase staining. In normal mucosa, selective subepithelial accumulation of B7.2+ cells was found. In inflamed IBD mucosa, however, subsets appeared consisting of both B7.2(hi) and B7.1(hi) cells as well as CD14(hi) macrophages. Notably, outside lymphoid aggregates the prominent fraction of recently recruited CD14(hi) macrophages comprised most (approximately 80%) of the B7.1(hi) cells, whereas most (approximately 70%) B7.2(hi) cells were identified as resident mucosal macrophages (CD14(lo) or CD14-). Differential expression of B7.1 and B7.2 on two functionally different subsets of intestinal macrophages implies separate immunoregulatory roles for the two molecules. This finding is in keeping with recent experimental data demonstrating that monocyte-derived cells are crucial for immune responses at mucosal surfaces. Preferential B7.1 up-regulation might be critical in breaking the immunological tolerance to luminal antigens in IBD, but it cannot be excluded that it is a secondary pathogenic event.     

Public health issues in aquaculture

Immune surveillance of skin cancer involves the stimulation of effector T cells by tumor-derived antigens and antigen-presenting cells (APCs). An effective APC must not only display processed antigen in the context of MHC molecules but also express co-stimulatory molecules that are required to fully activate T cells. One of the most common cutaneous neoplasms is basal cell carcinoma. To investigate expression of the co-stimulatory molecules CD80 (B7-1) and CD86 (B7-2) on tumor-associated dendritic cells (TADCs), cryosections from basal cell carcinomas were immunostained. In basal cell carcinomas, only 1 to 2% of intratumor and 5 to 10% of peritumor APCs expressed CD80 or CD86. In contrast, biopsies of immunological/inflammatory dermatoses revealed that 38 to 73% of APCs expressed CD80 and CD86. To further evaluate their phenotype and function, TADCs were isolated from tissue samples of basal cell carcinomas; they were non-adherent to plastic, displayed a typical dendritic morphology, and expressed high levels of major histocompatibility class II molecules on their surface. When TADCs were compared with dendritic cells from blood for presentation of superantigens (staphylococcal enterotoxins A and B) to resting autologous T cells, TADCs were consistently weaker stimulators of T cell proliferation than blood dendritic cells. When analyzed by flow cytometry, TADCs expressed high levels of HLA-DR, but only 5 to 10% co-expressed CD80 or CD86. A 3-day culture in granulocyte/macrophage colony-stimulating factor-containing medium partially reconstituted the TADC expression of CD80 and CD86 as well as their immunostimulatory capacity. Thus, in this common skin cancer, although there are prominent collections of HLA-DR-positive APCs in and around tumor cells, the TADCs are deficient in important co-stimulatory molecules as well as being weak stimulators of T cell proliferation. The paucity of co-stimulatory molecule expression and functional activity of TADCs may explain why the local T lymphocytic infiltrate fails to become fully activated to eradicate adjacent tumor cells. From a clinical perspective, these findings suggest a novel immunotherapeutic strategy targeting T cell co-stimulatory molecules on professional APCs in cutaneous oncology.     

Fas antigen (CD95) expression in peripheral blood progenitor cells from patients with leukaemia and lymphoma

Fas antigen (CD95) is a cell surface receptor belonging to the tumour necrosis factor/nerve growth factor superfamily and is able to induce apoptosis when triggered by its' natural ligand or an anti-Fas antibody. Fas expression is low on CD34+ bone marrow (BM) progenitor cells, but is increased by various cytokines in vitro. We investigated Fas expression on CD34+ cells from 39 peripheral blood progenitor cell (PBPC) harvests and from 5 normal BM harvests by dual colour flow cytometry to determine if Fas expression was altered during mobilisation. By including calibrated microbeads during flow cytometry, we quantified the number of Fas antigen molecules per cell. A low percentage of PBPC (22%) and normal BM (23%) CD34+ cells expressed Fas antigen. Fas expression varied on CD34+ cells from different diseases and the highest expression was found in ALL (52%). There was a significant three fold increase in the number of Fas molecules/cell expressed on CD34+ cells (PBPC 6,230 molecules/cell, BM 2,236; p = 0.0003). This level of expression was considerably less than that for CD3/CD19 lymphocytes (33,095 molecules/cell) and CD14 monocytes (47,467 molecules/cell) in the PBPC harvest. In conclusion, mobilisation including the use of growth of factors, has minimal effect on CD34 progenitor cell Fas expression.     

Managing hypertensive urgencies in primary care

The c-kit gene product (CD117) is known to be expressed by a variety of normal human tissue cell types, including breast epithelium, germ cells, melanocytes, immature myeloid cells, and mast cells. To further characterize the expression of this antigen, 117 normal human tissues and 576 human tumors were studied by paraffin section immunohistochemistry. Varying degrees of CD117 expression were identified in various normal cells and in 53% of all tumors studied. In most cases (42% of total), CD117 expression was weak. Expression was most common in mast cell disease (100%), testicular germ cell tumors (100%), endometrial carcinomas (100%), papillary and follicular thyroid carcinomas (100%), small cell carcinomas (91%), malignant melanomas (90%), and ovarian epithelial carcinomas (87%). Strong immunoreactivity was only identified in cases of mast cell disease (11 of 11 cases), serous ovarian carcinoma (3 of 16), malignant melanoma (2 of 40), small cell lung carcinoma (one of seven), and adenoid cystic carcinoma (one of one). Although the pattern of reactivity was primarily cytoplasmic, a membrane staining pattern was seen in a subset of cases, and strong membrane staining was identified in normal mast cells and all cases of mast cell disease. The lack of tumor specificity of weak expression of this antigen limits its diagnostic utility in most cases. However, the strong membrane reactivity for CD117 identified in mast cells may be useful in the diagnosis of mast cell disorders.     

Low expression of CD40 and B7 on macrophages infiltrating UV-exposed human skin; role in IL-2Ralpha-T cell activation

After UV exposure of skin, epidermal Langerhans cells (LC) are depleted, whereas CD11b+CD36 CD1a- monocytes/macrophages (UV-Mphi) infiltrate. Different immunological outcomes in vivo are mediated by LC (sensitization) and UV-Mphi (tolerance) which may be related to the distinct T cell activation states that these antigen-presenting cells (APC) induce. We previously demonstrated that CD4+ T lymphocytes activated by UV-Mphi are, in contrast to LC-activated T cells, IL-2Ralpha deficient, and we hypothesize that this differential T cell activation is related to differences in co-stimulatory molecules between UV-Mphi and LC. Using four-color flow cytometry, we found a reduced capacity to up-regulate expression of the important co-stimulatory molecules CD40, B7-1 and B7-2 by UV-Mphi relative to LC. This alteration in co-stimulatory molecule expression was selective, because UV-Mphi express equal levels of ICAM-1 and ICAM-3, and increased levels of LFA-1, relative to LC. After bidirectional signaling with T cells during alloantigen presentation, UV-Mphi still exhibited less CD40 and B7-1 than LC. Addition of IFN-gamma induced CD40 and B7-1 expression on UV-Mphi and restored IL-2Ralpha expression on UV-Mphi-activated T cells but had no effect on IL-2Ralpha on resting or LC-activated T cells. The restoration of IL-2Ralpha expression on UV-Mphi-activated T cells by IFN-gamma was inhibited (67 %, p = 0.005) by addition of neutralizing anti-CD40. Therefore, differences in co-stimulatory molecule expression, in particular CD40, on UV-Mphi and LC are critical in determining the distinct T cell activation induced by these APC.     

Spontaneous and ligand-induced trafficking of CXC-chemokine receptor 4

A chimeric protein consisting of CXC-chemokine receptor 4 (CXCR4) and the green fluorescent protein (GFP) was used for studying receptor localization and trafficking in real time in stably transduced HeLa, U-937, CEM, and NIH/3T3 cells. CXCR4-GFP was fully active as a co-receptor in mediating human immunodeficiency virus (HIV) entry. Both CXCR4 and CXCR4-GFP were found to undergo significant spontaneous endocytosis. Only 51.5 +/- 7.8% of receptor molecules were found on the plasma membrane in CD4-positive cells, 43.9 +/- 8.5% were found in CD4-negative HeLa cells, 75.6 +/- 9.7% were found in U-937 cells, 72.5 +/- 7.9 were found in CEM cells, and almost none were found in in NIH/3T3 cells. Stromal cell-derived factor-1alpha induced rapid endocytosis of cell surface receptor molecules. A significant part of CXCR4 was targeted to lysosomes upon binding of the ligands, and recycling of internalized CXCR4 was not efficient. Only about 30% of receptor molecules recycled back to the cell surface in HeLa cells, 5% recycled in U937, and 10% recycled in CEM cells, suggesting that the protective effect of chemokines against HIV infection can be attributed not only to competition for binding but also to depletion of the co-receptor molecules from the cell surface. Envelope glycoprotein gp120 of syncytia-inducing/lymphocyte tropic HIV-1 strains induced rapid internalization of CXCR4 in both CD4-negative and CD4-positive cells, suggesting that gp120 is a high affinity ligand of CXCR4.     

Correlation of very late activation integrin and CD44 expression with extrarenal invasion and metastasis of renal cell carcinomas

Cell adhesion molecules mediate cell-cell and cell-matrix interactions, and they are thought to play an important role in tumor invasion and metastasis. Altered expression of integrins and CD44 in renal cell carcinoma has been recently demonstrated, but an association with invasive or metastatic behavior has not been reported. We examined very late activation (VLA) integrin and CD44 expression in 37 renal cell carcinomas and correlated adhesion molecule expression with multiple histological and clinical parameters. Most tumors exhibited positive staining for VLA3 (81%). Approximately one third of the tumors stained positively for VLA6 and CD44, and fewer (27%) were positive for VLA2. Only a few tumors were positive for VLA4 (8%) and VLA5 (14%). Most of the tumors exhibiting positive staining showed a combination of membranous and cytoplasmic staining patterns. Low-grade tumors positive for VLA6 showed a tendency for basilar staining of the tumor cells, whereas high-grade tumors exhibited diffuse cytoplasmic staining. All tumors exhibiting weak or strong positive staining for VLA4 or VLA5 showed extrarenal invasion or were known to have developed metastases at the time of nephrectomy. All tumors strongly positive for VLA2 or CD44 showed invasion beyond the renal capsule or metastases. In contrast to a previous study, no association was observed between positive staining and tumor grade. Nor were tumor size, architectural pattern, cell type, or DNA ploidy found to be associated with particular staining patterns. Although many of the invasive tumors showed no difference in VLA integrin or CD44 expression compared with tumors confined to the kidney, increased expression in some of them suggests that these cell adhesion molecules may contribute to the invasive or metastatic phenotype.     

T-cell infiltration of primary CNS lymphoma

We stained 13 primary CNS lymphomas (PCNSLs) (six from patients with AIDS, seven from immunocompetent patients) with a panel of antibodies to T cells (pan T cell [CD3], T helper cell [CD4], T suppressor cell [CD8], delta/delta cell [CD4-8-]), B cells (CD20), hematopoietic cells (T200), and NK cell (CD56). We estimated the percentage of tumor cells staining with each antibody. All tumors were B-cell lymphomas. The non-AIDS tumors showed a significant infiltration with CD3+ cells (mean of 10.82% of total cells). The AIDS patients' tumors showed a smaller percentage of CD3+ infiltrating cells (mean, 4.88% of total cells) (p<0.01). CD4+ cells were 9.11% of the total hematopoietic cells in the non-AIDS patients and 3.13% in AIDS patients (p<0.01). AIDS patients showed some CD8+ cells (0.3%), which was significantly higher than in immunocompetent patients (0%) (p<0.05). Very few tumor cells stained with the NK cell and delta/delta cell markers. Both immunocompetent and AIDS patients with PCNSL exhibit significant CD3+ and CD4+ cell infiltration of their tumors; this infiltration is significantly lower in AIDS patients. AIDS patients show a minor CD8+ cell infiltration of their tumors. These results on PCNSL are different from systemic lymphomas, which show a higher CD4 and CD8 cell infiltration, and may offer insights into the more aggressive nature of AIDS-related PCNSL.     

Maternal and fetal plasma concentrations of endothelin, lipidhydroperoxides, glutathione peroxidase and fibronectin in relation to abnormal umbilical artery velocimetry

The topoisomerase II alpha (topo II alpha) enzyme is the target for several chemotherapeutic agents, including etoposide, teniposide, mitoxantrone, and doxorubicin (topo II poisons). The enzyme also is a marker of cell proliferation. Most cases of Hodgkin's disease (HD) are responsive to combination chemotherapy regimes that include topo II poisons such as doxorubicin. Immunoperoxidase methods for detection of the topo II alpha isoenzyme are now available for use in formalin-fixed, paraffin-embedded tissues, which may provide information about the proliferative capacity and possible sensitivity of tumors to drugs that target topo II. We used a specific antibody to analyze subsets of HD for topo II alpha staining patterns. Formalin-fixed blocks from 49 cases of HD, including 20 nodular sclerosis (NS), 14 mixed-cellularity (MC), and 15 lymphocyte-predominant (LP) subtypes, were analyzed by dual staining for topo II in combination with monoclonal antibodies against Reed-Sternberg (RS) cells consisting of CD15 for the NS and MC subtypes and CD20 for LP lymphocytic and histiocytic (L & H) cells. The number of morphologically appropriate cells coexpressing the RS or L & H marker and topo II alpha was quantitated. Positive nuclear staining for topo II alpha in RS or L & H cells was seen in 100% of cases, irrespective of subtype. Coexpression of CD15 and topo II alpha was seen in 58.4% of the RS cells or mononuclear variants in NSHD cases and 68.4% in MCHD cases. No significant difference in the percentage of neoplastic cells expressing topo II alpha was found between NS and MC subtypes. Cases of LPHD showed coexpression of CD20 and topo II alpha in 84.4% of the L & H cells, a significant increase over the level of tumor cell coexpression seen in NSHD and MCHD (P < .001). Only one case was found to have a low (< 25% of tumor cell coexpression) level of topo II alpha expression. Immunohistochemical detection of a high level of topo II alpha expression in HD, irrespective of subtype, suggests a molecular explanation for the excellent response of most HD to standard combination chemotherapy, which can include topo II poisons. The LP subtype has a higher expression of topo II alpha in the neoplastic cell population than do NS or MC subtypes, perhaps indicating increased sensitivity of these tumors to topo II poisons. It may be possible to identify subsets of HD that are more or less sensitive to conventional chemotherapeutic regimes, which would help in the selection of appropriate treatment.     

Expression of the receptor for urokinase-type plasminogen activator in normal and neoplastic blood cells and hematopoietic tissue

Expression of the receptor for the urokinase type plasminogen activator (uPAR) has been studied by flow cytometry and immunohistology in normal blood and bone marrow cells, in vitro activated lymphoid cells, and tissue samples from reactive lymph nodes (n = 6), thymus (n = 2) and malignant lymphomas (n = 82), or leukemias (n = 32). HL-60 myeloid precursor cells and CD34-positive normal stem cells also were analyzed. In the normal cells, staining was confined to monocytes, macrophages, neutrophils, and myeloid precursors. No labelling was seen of normal or activated lymphoid cells. Purified CD34-positive hematopoietic progenitors were uPAR negative, but expressed uPAR during differentiation in short-term liquid culture stimulated in vitro by recombinant interleukin (IL)-1, IL-3, IL-6, granulocyte-macrophage colony stimulating factor (CSF), granulocyte-CSF, and stem cell factor. Enhanced uPAR expression was also seen in HL-60 cells after induction of differentiation with dimethyl sulfoxide or 1 alpha,25-dihydroxyvitamin D3. In lymphomas and leukemias, the staining pattern was similar to that seen in the normal cells with labelling of monocytic and myeloid that seen in the normal cells with labelling of monocytic and myeloid malignancies, but not of the neoplastic cells in B-cell or T-cell lymphomas or Hodgkin's disease. In conclusion, uPAR is a differentiation marker for myeloid and monocytic cells, and may act to facilitate migration of these cells in normal and pathologic conditions by cell-associated plasminogen activation. Whether expression of uPAR in myeloid and monocytic malignancies relates to their growth and behavior will be an important topic for investigations in the future.     

Modulation of CD72 by ligation of B cell receptor complex molecules on CD5+ B cells

B cells expressing CD5 also carry its ligand, CD72. As an approach to understanding the role of CD5 and CD72 on B cells, we have examined the association of CD72 with CD5 and slgM by modulation/co-modulation and capping/co-capping following ligation of these surface molecules with specific antibodies. Modulation and co-modulation were measured after 24 h, whilst capping was measured after 1 h. CD5 and slgM co-modulated each other, CD72 co-modulated with slgM and CD5, but anti-CD72 did not affect either slgM or CD5. CD5 and slgM co-capped each other, whilst CD72 failed to co-cap with either slgM or CD5. The CD5-induced co-modulation of CD72 was partially blocked by specific protein tyrosine kinase inhibitor, but not the slgM-induced co-modulation, Protein kinase C (PKC) inhibitors abrogated the anti-mu- but not the anti-CD5-triggered modulation of CD72, whereas PKC activators prevented the CD5- but not the slgM-induced 24 h modulation of CD72. None of these drugs was able to modify the anti-CD72-induced modulation of CD72. Our data suggest that CD5 is physically associated with slgM in the B cell receptor complex but not with CD72. Furthermore, from the effect of drugs on modulation, there appears to be different associations of CD72 with slgM and CD5. These two pathways differed in some respects, consistent with a co-stimulatory function of CD72 and CD5 in B cell activation.     

Infinity and the limits of the unconscious

Macrophages play an important role in the intestinal mucosal immune system. However, they are a poorly defined cell population. We therefore determined their phenotype in normal colonic mucosa. Macrophages were isolated from colonic biopsies and surgical specimens by collagenase digestion. Colonic macrophages were positively sorted by anti-CD33 magnetic beads. Flow cytometric triple fluorescence analysis was applied to study CD14, CD16, CD33, CD44, CD11b, CD11c, CD64, HLA-DR, CD80, CD86 and CD3/CD19 expression. CD33 was evaluated as a positive marker for intestinal macrophages. CD33+ cells isolated from normal colonic mucosa showed co-expression of the established intracellular macrophage marker CD68 in FACS analysis. CD33+ cells were capable of phagocytosis. Isolation of this cell population by magnetic anti-CD33 beads and culture resulted in a 4.2-40-fold increase in IL-1beta and 4.5-44-fold increase in tumour necrosis factor-alpha (TNF-alpha) secretion compared with unsorted lamina propria mononuclear cells (LPMC). Of the CD33+ cells, 90.9 +/- 6.9% (mean +/- s.d.) were CD44+. However, macrophages from colonic mucosa showed only a low expression of CD14 (10.5 +/- 3.8%), CD16 (10.1 +/- 3.9%), HLA-DR (27.3 +/- 9.2%), CD11b (17.4 +/- 6.8%), CD11c (17.8 +/- 10.4%). Furthermore, expression of CD80 (9.2 +/- 4.2%) and CD86 (15.1 +/- 7.3%) was low, suggesting a low ability of normal intestinal macrophages to activate T cells and T cell-mediated immune responses. We conclude that CD33 is useful for the isolation and flow cytometric characterization of colonic macrophages. These cells exhibit a single phenotype in normal mucosa (CD33++, CD44++, CD14-, CD16-, CD11b-, CD11c-, HLA-DRlow, CD80-, CD86-) lacking lipopolysaccharide (LPS) receptor and costimulatory molecules.