Semliki Forest virus infection leads to increased expression of adhesion molecules on splenic T-cells and on brain vascular endothelium

Semliki Forest virus A7 (SFV-A7) is a neurotropic alphavirus that leads to an asymptomatic encephalitis in adult immunocompetent mice. We studied the expression of leukocyte and endothelial cell adhesion molecules in the spleen and in the central nervous system (CNS) during SFV-A7 infection. Kinetics of the expression of LFA-1 alpha/CD11a, LFA-1 beta/CD18, Mac-1/CD11b, VLA-4/CD49d, ICAM-1/CD54 and L-selectin/CD62L was determined on splenic CD4+ and CD8+ T-cells and macrophages by flow cytometry. Time course of the expression of these antigens and VCAM-1/CD106 as well as viral antigens in the CNS was studied by immunoperoxidase staining. In the spleen, a sustained increase in LFA-1-expression and a temporary increase at day 7 in the expression of VLA-4, Mac-1 and ICAM-1 were detected on CD8+ T-cells. L-selection was down-regulated on CD4+ cells. Adhesion molecules on macrophages remained unchanged. In the CNS, expression of Mac-1+, VLA-4+ and LFA-1+ cells increased in parallel with the kinetics of the expression of their ligands ICAM-1 and VCAM-1 on brain vessels. Upregulation of adhesion of molecules peaked between days 5-8 and was most prominent in the cerebellar and brain stem white matter where viral antigens were most abundant. We conclude that the adhesion molecules profile of splenic T cells is altered during SFV-A7 infection which may influence their homing into the CNS. Macrophages are probably recruited non-specifically as a consequence of activation of the brain vascular endothelium in the inflamed areas of the brain.     

Increase in adhesion molecules on CD4+ cells and CD4+ cell subsets in synovial fluid from patients with rheumatoid arthritis

OBJECTIVE: To elucidate the role of adhesion molecules in the pathogenesis of rheumatoid arthritis (RA). METHODS: We evaluated their expression and that of an activation marker on CD4+ cell populations and CD4+ cell subsets in specimens of peripheral blood (PB) and synovial fluid (SF) obtained from 10 patients with RA and 7 with osteoarthritis (OA). A 2 or 3-color immunofluorescent method was used for analysis. RESULTS: The SF from both groups of patients showed a greater density of adhesion molecules including LFA-1 alpha, LFA-1 beta, CD2, VLA-4 alpha and VLA-5 alpha on CD4+ cells, and a higher percentage of CD4+HLA-DR+ cells compared with their PB. IN PB-CD4+ cell subsets from the arthritic and healthy subjects, the CD4+CD45RO+ cell population showed an increased expression of adhesion molecules compared with CD4+CD45RA+ cell population. The expression of adhesion molecules on circulating CD4+ cell population and CD4+ cell subsets from the patients with RA and OA was comparable to that from healthy subjects. SF from both groups of patients showed a higher percentage of CD4+CD45RO+ cells and a lower percentage of CD4+CD45RA+ cells. In SF-CD4+ cell subsets from patients with RA, the CD4+CD45RO+ cell population had an increased expression of VLA-4 alpha compared to the CD4+CD45RA+ cell population; however, there was no significant difference in other adhesion molecule expression and the percentage of HLA-DR+ cells between the 2 cell subsets. Furthermore, the expression of VLA-4 alpha on the CD4+CD45RO+ cell population in SF from patients with RA was significantly higher than that in matched PB. In CD4+CD45RA+ cell population from both groups of patients, SF showed an enhanced expression of adhesion molecules and an increased percentage of HLA-DR+ cells compared with matched PB. CONCLUSION: Our results suggest that increased expression of adhesion molecules and increased percentage of HLA-DR+ cells on CD4+ cells in SF may be responsible for cellular interactions between these cells and synovial cells or extracellular matrix.     

Methylprednisolone reduces adhesion molecules in blood and cerebrospinal fluid in patients with MS

OBJECTIVE: To analyze the expression of adhesion molecules on mononuclear cells from blood and CSF of patients with exacerbations of MS before and after megadose IV methylprednisolone (MP). BACKGROUND: Adhesion molecules regulate transmigration of lymphocytes and monocytes/macrophages to the CNS and have an important role in the pathogenesis of MS. METHODS: The expression of very late activation antigen 4 (VLA-4) and vascular cell adhesion molecule 1, lymphocyte function-associated antigen 1 (LFA-1), and intercellular adhesion molecule 1 (ICAM-1) was analyzed immunocytologically on lymphocytes and monocytes from blood and CSF of 23 patients and 11 healthy control subjects. The results were correlated with the Expanded Disability Status Scale and in half of the patients with the number of T2-weighted MS plaques and brain atrophy analyzed by MRI. RESULTS: After treatment, the mean proportions of VLA-4, LFA-1, and ICAM-1 on blood lymphocytes (p < 0.0003, p < 0.00001, p < 0.01) and monocytes (p < 0.0001, p < 0.0002, p < 0.007) of 23 patients decreased. The expression of these adhesion proteins was also diminished on CSF leukocytes. However, even after treatment, the levels of VLA-4 and LFA-1 on lymphocytes from blood of MS patients remained higher than in the control subjects. The level of VLA-4 and LFA-1 on blood lymphocytes (r=0.67, p=0.023) and VLA-4 on monocytes (r=0.61, p=0.047) correlated with the number of T2-weighted lesions. CONCLUSIONS: Megadose MP may suppress brain inflammation by reducing the expression of adhesion molecules on mononuclear cells from blood and CSF of MS patients. The inhibition of cellular trafficking in MS by MP offers an important means of altering the autoimmune response in MS.     

Adhesion molecule expression on eosinophils in idiopathic eosinophilic pneumonia

Idiopathic eosinophilic pneumonia (IEP) is characterized by the accumulation of eosinophils in the alveolar spaces and the interstitium of the lung, frequently accompanied by peripheral eosinophilia. To clarify the roles of adhesion molecules of eosinophils in the pathogenesis of eosinophilic pneumonia, we analysed their expression by eosinophil and T-lymphocyte populations in peripheral blood and bronchoalveolar lavage fluid (BALF) obtained from 11 patients with eosinophilic pneumonia, using flow cytometric methods. Cell differentials in BALF showed increased numbers of eosinophils, the increase correlating with the number of activated T-lymphocytes in BALF. The expressions of CD11a (lymphocyte function-associated antigen-1 (LFA-1)), CD11b (Mac-1), CD18, CD49d (very late activation antigen-4 (VLA-4)), and CD62L (L-selectin) by eosinophils in BALF were all lower than those of eosinophils in peripheral blood. In contrast, CD54 (intercellular adhesion molecule-1 (ICAM-1)) was expressed by eosinophils in BALF, but not by those in peripheral blood. These results indicate that intercellular adhesion molecule-1 expression by eosinophils in bronchoalveolar lavage fluid but not in peripheral blood may be induced by locally activated T-cells or macrophages and may be important in the pathogenesis of idiopathic eosinophilic pneumonia.     

Fluorescein-labeled dextran concentration is increased in BAL fluid after ANTU-induced edema in rats

We identified the cell cycle status of CD34(+) cells of steady-state bone marrow (BM) and peripheral blood (PB) obtained from healthy volunteers, and those of apherasis PB samples collected from healthy donors who had been administered granulocyte colony-stimulating factor (G-CSF). More than 10% of CD34(+) cells in BM were in S+G2/M phase. In contrast, regardless of whether G-CSF treatment was performed, less than 2% of CD34(+) cells in PB were cycling. BM CD34(+) cells showed greater VLA-4 expression and adherence to stromal cells than PB CD34(+) cells. In addition, when cycling and dormant BM CD34(+) cells were analyzed separately, the cells in S+G2/M phase expressed more VLA-4 and adhered to the stromal cell monolayer more efficiently than the cells in G0/G1 phase. Furthermore, this adhesion of CD34(+) cells to the stromal cell layer was almost completely inhibited by anti-VLA-4 antibody. Taken together, these results suggest that CD34(+) progenitors in G0/G1 phase of the cell cycle differ from those in S+G2/M phase in adhesiveness mediated by VLA-4 in the hematopoietic microenvironment.     

Cytokines and immune regulation in thyroid autoimmunity

We studied the role of cytokines and immune regulation in thyroid tissue from patients with Graves' disease. Immunohistochemistry showed that the thyroid glands are characterized by an aberrant expression of HLA class II antigens on thyrocytes, generation of new blood vessels and infiltration of mononuclear cells. We demonstrated that CD4+ memory cells were more frequent in thyroid glands from Graves' patients than were CD4+ naive cells. The intrathyroidal T cells demonstrated an enhanced expression of the adhesion molecules LFA-1, CD2, VLA-4 and VLA-5, and vascular endothelial cells of capillaries and thyrocytes in thyroid glands reacted with anti-ICAM-1 monoclonal antibody. The adhesion molecules and HLA antigens on both vascular endothelial cells and thyrocytes were regulated by inflammatory cytokines. These results suggest that circulating lymphocytes migrate into thyroid tissues and that memory T cells are retained in the thyroid tissues by cellular interactions with thyrocytes or with extracellular matrix.     

Involvement of lymphocyte function-associated antigen-1 and intercellular adhesion molecule-1 in osteoclastogenesis: a possible role in direct interaction between osteoclast precursors

In our search for molecules involved in the process of osteoclast differentiation, we examined the surface phenotypes of the preosteoclast-like cells and osteoclast-like multinucleated cells (MNCs) formed in bone marrow cultures, using monoclonal antibodies recognizing different antigen molecules expressed on hematopoietic cells. Among these cell surface antigens, lymphocyte function-associated antigen-1 (LFA-1) and intercellular adhesion molecule-1 (ICAM-1) were highly expressed on mononuclear cells in the cultures for forming preosteoclast-like mononuclear cells. The double detection of these two antigen molecules with osteoclast-specific antigen and with calcitonin receptor, using a fluorescence-activated cell sorter or autoradiography technique, revealed that LFA-1 and ICAM-1 were expressed on the preosteoclasts. The expression of ICAM-1 was detected on both preosteoclasts and osteoclast-like MNCs, whereas the expression of LFA-1 was restricted to preosteoclasts. We designed a peptide with the sequence of the binding site of ICAM-1 against the ligand LFA-1. In the whole bone marrow culture system for forming osteoclast-like MNCs, a significant inhibition of MNC formation was observed by the addition of this peptide. These results strongly suggest the involvement of an LFA-1/ICAM-1-interaction in osteoclastogenesis.     

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.     

O-glucuronidation, a newly identified metabolic pathway for topotecan and N-desmethyl topotecan

BACKGROUND AND OBJECTIVE: CD34+ hematopoietic progenitor cells (HPCs) constitute a heterogeneous population both in size and in immunological features. Lack of CD38, HLA-DR and lineage committed antigens as well as the co-expression of Thy-1 (CDw90) and c-kit receptor (CD117), are able to identify the so-called stem cells. A flow cytometric study was carried out to investigate the co-expression of Thy-1 and c-kit receptors, both members of Ig superfamily adhesion molecules, involved in cell to cell and cell to stroma interactions, on bone marrow (BM), mobilized peripheral blood (PB) and human umbilical cord blood (HUCB) CD34+ HPCs. DESIGN AND METHODS: Lysed whole blood from 15 BM, 25 mobilized PB and 25 HUCB samples were used to perform a five-dimensional flow cytometric evaluation of both CDw90 and CD117 on CD34+ cells. RESULTS: Few CD34+ cells co-expressed Thy-1 antigen in all three compartments (BM: 11.2 +/- 7.2%; PB: 6.2 +/- 3.6%; HUCB: 6 +/- 2.9%; BM vs PB < 0.04; BM vs HUCB < 0.008; PB vs HUCB ns). c-kit receptor was detected on the majority of CD34+ HPCs, particularly in HUCB (HUCB: 80.7 +/- 8.2%; BM: 72.3 +/- 13.1%; PB: 64.2 +/- 17%; HUCB vs BM < 0.03; HUCB vs PB < 0.0001; BM vs PB ns). CD34+Thy-1+ and CD34+c-kit+ HPCs generally displayed HLA-DR antigen, as expression of early cell commitment. However, the most immature CD34+Thy-1+HLA-DR- (HUCB: 1 +/- 0.6%; BM: 0.4 +/- 03%; PB: 0.7 +/- 0.5%; HUCB vs BM < 0.0001; BM vs PB < 0.04; HUCB vs PB ns) and CD34+c-kit+HLA-DR- HPCs (HUCB: 6.5 +/- 4.4%; BM: 6.3 +/- 4.8%; PB: 2.2 +/- 1.8%; HUCB vs BM ns; BM vs PB < 0.0001; HUCB vs PB < 0.0001) were mainly detected in HUCB. Finally, the greatest percentage of CD34+Thy-1+c-kit+ cells was found in BM (6.9 +/- 4.1%) followed by leukapheretic samples (4.4% +/- 2.7) and then by HUCB (3.7 +/- 1.2%; BM vs PB ns; BM vs HUCB < 0.001; HUCB vs PB ns). INTERPRETATION AND CONCLUSIONS: Since the blood release or HPCs is probably due to a perturbation of the adhesive interactions between these cells and the marrow stroma, the different pattern of Thy-1 and c-kit receptor expression on CD34+ HPCs found in the three hemopoietic compartments evaluated can lead to new knowledge about the mobilization kinetics in which the Ig superfamily adhesion molecules are involved.     

Adhesion of leukocytes to endothelial cells in atherosclerosis]

The adherence of blood monocytes to the endothelium, followed by transmigration beneath the endothelium, are initiating events in the formation of foam cells, promoting atherogenesis. We showed that adhesion molecules on leukocytes were up- or down-regulated in atherosclerosis, when binding of monoclonal antibodies was measured by indirect immunofluorescence with flow cytometry. Expression of PE-CAM-1 (CD31) on monocytes and LFA-1 (CD11a) on lymphocytes was increased with age. Expression of PECAM-1 in monocytes was also up-regulated in patients with coronary artery disease. Being unchanged on aging, expression of HAR (CD44) on polymorphonuclear leukocytes and monocytes was increased in patients with coronary artery disease. On the other hand, expression of L-selectin (CD62L) on polymorphonuclear leukocytes, and LFA-1, CR3 (CD11b) and VLA-4 (CD49d) on monocytes was decreased. These findings may show the mechanism of increased chemotaxis of monocytes beneath the endothelium during the incipient stage of atherosclerosis.     

HCA, an immunoglobulin-like adhesion molecule present on the earliest human hematopoietic precursor cells, is also expressed by stromal cells in blood-forming tissues

We have previously shown that the HCA/ALCAM (CD166) glycoprotein, a member of the immunoglobulin family that mediates both homophilic and heterophilic cell-cell adhesion, via the CD6 ligand, is expressed at the surface of all of the most primitive CD38(-/lo), Thy-1(+), rho123(lo), CD34(+) hematopoietic cells in human fetal liver and fetal and adult bone marrow. In the present report we show that HCA is also expressed by subsets of stromal cells in the primary hematopoietic sites that sequentially develop in the human embryo and fetus, ie, the paraaortic mesoderm, liver, thymus, and bone marrow. Adult bone marrow stromal cells established in vitro, including those derived from Stro-1(+) progenitors and cells from immortalized cell lines, express HCA. In contrast, no HCA expression could be detected in peripheral lymphoid tissues, fetal spleen, and lymph nodes. HCA membrane molecules purified from marrow stromal cells interact with intact marrow stromal cells, CD34(+) CD38(-) hematopoietic precursors, and CD3(+) CD6(+) peripheral blood lymphocytes. Finally, low but significant levels of CD6 are here for the first time detected at the surface of CD34(+) rho123(med/lo) progenitors in the bone marrow and in mobilized blood from healthy individuals. Altogether, these results indicate that the HCA/ALCAM surface molecule is involved in homophilic or heterophilic (with CD6) adhesive interactions between early hematopoietic progenitors and associated stromal cells in primary blood-forming organs.     

Suppression of experimental autoimmune encephalomyelitis with a TNF binding protein (TNFbp) correlates with down-regulation of VCAM-1/VLA-4

The effect of a novel TNF binding protein (TNFbp), a polyethylene glycol-linked form of the type I soluble receptor of TNF, on the expression of adhesion molecules has been investigated with a passive transfer model of experimental autoimmune encephalomyelitis (EAE) in SJL/J mice. The expression of L-selectin, VLA-4 and LFA-1 on spleen cells of EAE animals treated with TNFbp or saline was examined by FACS analysis. The expression of VCAM-1 and ICAM-1 was investigated by immunochemistry in spinal cord tissue of SJL/J mice with EAE. In animals sensitized for EAE and treated with TNFbp, the expression of VCAM-1 in the central nervous system as well as VLA-4 on spleen cells was clearly diminished. Reduction in VCAM-1 staining and VLA-4 expression corresponded to inhibition of inflammation in the spinal cord and to prevention of clinical signs of EAE. The results have also shown that myelin basic protein responses as well as non-antigen-specific responses were not diminished in animals treated with TNFbp. The findings suggest that TNFbp might prevent EAE development by modulating the expression of VCAM-1 and VLA-4.     

L-selectin crosslinking induces integrin-dependent adhesion: evidence for a signaling pathway involving PTK but not PKC

L-selectin mediates the initial contact of leukocytes with the endothelium prior to extravasation. Here we demonstrate that L-selectin engagement can induce rapid and avid integrin-dependent T cell adhesion to recombinant immobilized cell adhesion molecules (CAMs) including ICAM-1, ICAM-3, and VCAM-1, as well as to the extracellular matrix protein fibronectin (FN). L-selectin-induced adhesion to these integrin ligands shares characteristics with CD3 mAb- or phorbol ester-induced adhesion in requiring metabolic energy, tyrosine kinase and ligand-stimulated Ca2+ channel activity. However, L-selectin-induced adhesion is distinct from that induced by phorbol ester or CD3 crosslinking in being relatively independent of protein kinase C (PKC) activity and actin polymerization. Consistent with the higher levels of L-selectin expression on CD45RA+ (naive) cells, L-selectin crosslinking induces a greater proportion of naive relative to memory cell binding to CAMs and FN. In contrast, exposure to phorbol ester or CD3 crosslinking is more effective in inducing CD45RO+ (memory) cell adhesion. Thus, in addition to its role in leukocyte capture and rolling on the endothelium, L-selectin may contribute to beta 1 and beta 2 integrin-dependent binding and arrest.     

Multiparameter phenotype mapping of normal and post-chemotherapy B lymphopoiesis in pediatric bone marrow

We studied the differentiation profiles of B cell precursors (BCP) in normal and post-chemotherapy pediatric bone marrow (BM) using multiparameter flow cytometry. The goal of our study was to draw a comprehensive phenotypic map of the three major maturational BCP stages in BM. By correlating lineage-associated markers, CD45RA, and several adhesion molecules, the stage-specific patterns were found to differ in certain details from previously published concepts. Among the earliest BCP, a subset of CD34+ CD10(lo) precursors was repeatedly observed in addition to the well characterized CD34+ CD10(hi) CD19+ majority of cells. Only two-thirds of these CD34+ CD10(lo) cells expressed CD19. However, uniformity of phenotypic features, absence of T lineage markers, and the regeneration kinetics after chemotherapy suggest the B lineage affiliation of the CD34+ CD10(lo) precursors in general. In the more mature BCP, expression of CD10, CD20, cytoplasmic and surface mu chains (c mu and s mu) was observed to overlap more than previously recognized. We found that CD20 and c mu appear early during B cell ontogeny (already on CD34+ BCP), and that CD10 is lost late, following the onset of s mu expression. Differences between normal and post-chemotherapy BM specimens regarding the phenotypic appearance of BCP were exclusively due to differences in the subset composition, as post-chemotherapy samples showed a preponderance of immature stages. Our observations may build a framework for comparing leukemic cells with their normal counterparts to define possible leukemia-associated aberrations useful for residual disease studies.     

Are circulating neutrophils intravascularly activated in patients with anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitides?

Vascular injury in vasculitis may be due to activation of circulating neutrophils resulting in their increased adhesiveness to locally activated endothelium (Shwartzman phenomenon). Previously, we demonstrated up-regulation of endothelial intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in biopsies from patients with ANCA-associated vasculitis. In the present study, we investigated the expression of adhesion molecules (CD11b, ICAM-1, VLA-4, L-selectin) and activation markers (CD66b, CD64, CD63) on circulating neutrophils from patients with ANCA-associated vasculitis in comparison with their expression on cells from healthy volunteers and patients with sepsis. We related these findings to parameters of disease activity. Surface marker expression was determined by using a non-activating whole blood flow cytometric assay. The expression of activation markers, but not the expression of adhesion molecules, was increased on neutrophils from patients with active vasculitis. The expression of CD63 and CD66b on neutrophils correlated with disease activity as determined by the Birmingham Vasculitis Activity Score (BVAS). In contrast to patients with active vasculitis, patients with sepsis showed up-regulation of all markers, including adhesion molecules, suggesting that circulating neutrophils are fully activated in sepsis. We conclude that in ANCA-associated vasculitis, circulating neutrophils are not fully activated, since they do not express increased levels of adhesion molecules as sepsis or in the Shwartzman reaction. These findings are compatible with the concept that in vivo vascular damage in ANCA-associated vasculitides does not occur due to a Shwarzman-like reaction but only after ANCA-induced neutrophil activation at the endothelial cell surface.     

Effects of intravenous methylprednisolone therapy on leukocyte and soluble adhesion molecule expression in MS

Intravenous methylprednisolone (IVMP) may inhibit inflammatory cell recruitment to active MS lesions by effects on leukocyte or endothelial cell adhesion molecule expression. We investigated 15 MS patients in relapse receiving a 5-day course of IVMP (500 mg/day) and 15 normal subjects. Patients' blood samples were obtained pretreatment, at 6 and 24 hours after the first dose, and 48 hours after completion of therapy. Levels of L-selectin, leukocyte functional antigen 1 (LFA-1), Mac-1, and very late activation antigen 4 (VLA-4) expression were determined on alphabeta and gammadelta T cells and monocytes by dual-color immunofluorescent flow cytometry. Serum levels of soluble (s) L-selectin, sE-selectin, soluble intercellular adhesion molecule 1 (sICAM-1) and soluble vascular cell adhesion molecule 1 (sVCAM-1) were measured by ELISA. There was a marked decrease in the T-cell and monocyte counts at 6 hours after therapy, with recovery to baseline at 24 to 48 hours. Adhesion molecule expression was normal on circulating T cells and monocytes in active MS. IVMP resulted in significant changes in the percent adhesion molecule expression on monocytes: increased L-selectin expression at 24 hours, decreased Mac-1 expression at 6 hours, and decreased VLA-4 expression at 6 hours and 24 hours following treatment. T-cell adhesion molecule expression was unaffected by the therapy. Serum sE-selectin was reduced at 6 hours and 24 hours following treatment. IVMP alters the distribution and kinetics of monocyte adhesion molecule expression and endothelial cell release of E-selectin, which may limit monocyte recruitment to areas of tissue destruction in MS.     

Biomechanics of human common carotid artery and design of novel hybrid textile compliant vascular grafts

Entorhinal cortex lesion (ECL) leads to anterograde degeneration of perforant path axons and is known to induce a rapid and intense reaction of astrocytes and microglial cells in the deafferented dentate gyrus. Phagocytosis of degenerating axons involves the establishment and maintenance of cell-matrix and cell-cell interactions by activated glial cells. It was thus our aim to investigate whether the process of axon phagocytosis is accompanied by the expression of adhesion molecules on activated microglial cells or reactive astrocytes, as such molecules mediate bot cell-matrix and cell-cell interactions. We found that the integrin adhesion molecules leukocyte function antigen-1 (LFA-1), very late antigen-4 (VLA-4), and the ligand for LFA-1, intercellular adhesion molecule-1 (ICAM-1), were expressed on microglial cells accumulating in the outer molecular layer of the deafferented dentate gyrus. This upregulation of adhesion molecule expression on microglial cells showing morphological criteria of activation occurred rapidly following ECL, reached its peak at 3 days post lesion (dpl), and gradually returned to control levels after 9 dpl. Astrocytes were never labeled by antibodies directed against these adhesion molecules. Prelabeling of the perforant path with a fluorescent tracer and subsequent ECL led to phagocytosis of fluorescent-labeled axonal debris by cells that were located in the outer molecular layer and showed typical microglial morphology. Double-fluorescence labeling demonstrated that microglial cells engaged in the phagocytosis of axonal debris expressed LFA-1, VLA-4, and the LFA-1-ligand ICAM-1. In conclusion, our results demonstrate that anterograde degeneration of perforant path axons results in adhesion molecule expression on activated microglial cells engaged in axon phagocytosis. The expression of such molecules could represent a mechanism that retains activated microglia in areas of axonal degeneration and perhaps enables the interaction of microglial cells with each other or with other immunocompetent cells.     

Mobilization and homing of peripheral blood progenitors is related to reversible downregulation of alpha4 beta1 integrin expression and function

Despite the wide use of mobilized peripheral blood (PB) progenitor cells (PBPC) for clinical transplantation the mechanism(s) underlying their mobilization and subsequent engraftment are still unknown. We compared the adhesive phenotype of CD34(+) colony-forming cells (CFC) in bone marrow (BM) and PB of normal donors before and after administration of granulocyte colony-stimulating factor (G-CSF) for 5 d. G-CSF-mobilized PB CFC cells adhered significantly less to BM stroma, fibronectin, and to the alpha4 beta1 binding fibronectin peptide, CS1, because of decreased expression of the alpha4 integrin. Since incubation of BM CD34(+) cells for 4 d with G-CSF at concentrations found in serum of G-CSF- treated individuals did not affect alpha4-dependent adhesion, G-CSF may not be directly responsible for the decreased alpha4-mediated adhesion of PB CFC. Culture of G-CSF-mobilized PB CD34(+) cells with cytokines at concentrations found in BM stromal cultures upregulated alpha4 expression and restored adhesion of mobilized PB CFC to stroma, fibronectin, and CS1. Adhesion of cultured, mobilized PB CFC to stroma and CS1 could not be further upregulated by the beta1 activating antibody, 8A2. This indicates acquisition of a maximally activated alpha4 beta1 integrin once PB CFC have been removed from the in vivo mobilizing milieu. Thus, decreased alpha4 expression on CD34(+) CFC in PB may be responsible for the aberrant circulation of mobilized PB CD34(+) cells. Reexpression of a maximally activated alpha4 beta1 integrin on mobilized PB CFC removed from the mobilizing in vivo milieu may contribute to the early engraftment of mobilized PBPC.     

Cell adhesion molecules--update

Cell adhesion molecules are glycoproteins expressed on the cell surface and play an important role in inflammatory as well as neoplastic diseases. There are four main groups: the integrin family, the immunoglobulin superfamily, selectins, and cadherins. The integrin family has eight subfamilies, designated as beta 1 through beta 8. The most widely studied subfamilies are beta 1 (CD29, very late activation [VLA] members), beta 2 (leukocyte integrins such as CD11a/CD18, CD11b/CD18, CD11c/CD18, and alpha d beta 2), beta 3 (CD61, cytoadhesions), and beta 7 (alpha 4 beta 7 and alpha E beta 7). The immunoglobulin superfamily includes leukocyte function antigen-2 (LFA-2 or CD2), leukocyte function antigen-3 (LFA-3 or CD58), intercellular adhesion molecules (ICAMs), vascular adhesion molecule-1 (VCAM-1), platelet-endothelial cell adhesion molecule-1 (PE-CAM-1), and mucosal addressin cell adhesion molecule-1 (MAdCAM-1). The selectin family includes E-selectin (CD62E), P-selectin (CD62P), and L-selectin (CD62L). Cadherins are major cell-cell adhesion molecules and include epithelial (E), placental (P), and neural (N) subclasses. The binding sites (ligands/receptors) are different for each of these cell adhesion molecules (e.g., ICAM binds to CD11/CD18; VCAM-1 binds to VLA-4). The specific cell adhesion molecules and their ligands that may be involved in pathologic conditions and potential therapeutic strategies by modulating the expression of these molecules will be discussed.     

Differences in glomerular leukocyte infiltration between IgA nephropathy and membranoproliferative glomerulonephritis

BACKGROUND: An important aspect in glomerular nephritic processes is the enhanced influx of leukocytes into the glomerulus. METHODS: To investigate the mechanisms of intraglomerular leukocyte infiltration in IgA nephropathy (IgA-N) and membranoproliferative glomerulonephritis type I (MPGN-I), we immunohistochemically examined the intraglomerular expression of leukocyte function-associated antigen-1 (LFA-1, CD11a/CD18), macrophage-1 (Mac-1, CD11b/CD18) and intercellular adhesion molecule-1 (ICAM-1, CD54) together with glomerular deposition of C3c and fibrinogen. RESULTS: In IgA-N (n=42), LFA-1+ cells were distributed mainly in glomeruli with intense expression of ICAM-1, and there was a positive correlation (P<0.001) between the number of LFA-1+ cells and the degree of ICAM-1 expression. Mac-1+ cells had no correlation with glomerular C3c deposition, but had a significant correlation with fibrinogen deposition (P<0.05). The number of LFA-1+ cells was significantly greater than of Mac-1+ cells (P<0.05). The number of LFA-1+ cells was strongly correlated with that of CD68+ cells (P<0.00001). In MPGN-I (n= 43), on the contrary, Mac-1+ cells correlated only with C3c deposition (P<0.001), and they were observed mainly in peripheral loops of glomerular capillaries where C3c was deposited with a similar distribution. However, there was no relationship between LFA-1+ cells and ICAM-1 expression. The number of Mac-1+ cells was greater than that of LFA-1+ cells (P<0.0001), and most Mac-1+ cells were identical to CD15+ cells. CONCLUSION: These results indicate the possibility that different mechanisms may cause glomerular leukocyte infiltration in various forms of human glomerulonephritis. The LFA-1/ICAM-1 pathway may play an important role in glomerular leukocyte infiltration in IgA-N, while the Mac-1/complement pathway may be important in MPGN-I. The former may promote mainly the infiltration of CD68+ cells, and the latter may promote that of CD15+ cells. In addition, Mac-1+ cells may act as fibrinogen and complement receptors in IgA-N and MPGN-I, respectively.