Plasma levels of hepatocyte growth factor and placental growth factor predict mortality in a general population: a prospective cohort study.

BACKGROUND: Circulating levels of growth factors involved in leucocyte production and angiogenesis could be indicative of underlying aberrations of tissue homeostasis and therefore be utilized as predictors of risk for all-cause cardiovascular disease (CVD) or cancer mortality. METHODS: Baseline plasma levels of a range of growth factors were measured in two cohorts of the population-based FINRISK study (1997 Discovery cohort, N = 8444, aged 25-74; 2002 Replication cohort, N = 2951, aged 51-74 years) using a multiplexed bead array methodology and ELISA. Participants were followed up by linking them to registry data. RESULTS: In the Discovery cohort (653 deaths; 216 CVD-related, 231 cancer-related), fully adjusted Cox proportional hazard regression models showed that increased plasma hepatocyte growth factor (HGF) and placental growth factor (PlGF) were associated with higher risk of 10-year mortality (HR, 1.29 [95% confidence interval (CI), 1.18-1.41] and HR, 1.23 [95% CI, 1.14-1.32], respectively). In the Replication cohort (259 deaths; 83 CVD-related, 90 cancer-related), baseline HGF levels also predicted all-cause mortality (HR, 1.2 [95% CI, 1.08-1.32]; PlGF data not available). By including HGF levels in a CVD mortality model, 9% of all CVD deaths were correctly reclassified in the Discovery cohort (categorical net reclassification improvement [NRI] for events, P = 4.0 × 10-4 ). Moreover, adding HGF to all-cause and CVD mortality models resulted in an overall clinical NRI of 0.10-0.18 in the Discovery cohort and meta-analyses (P < 0.05 for all tests). CONCLUSION: Blood levels of HGF and PlGF may serve as new biomarkers for predicting increased risk of death in the general population.

Expression of the semicarbazide-sensitive amine oxidase in articular cartilage: its role in terminal differentiation of chondrocytes in rat and human.

OBJECTIVE: Semicarbazide-sensitive amine oxidase (SSAO) catalyzes the oxidation of primary amines into ammonia and reactive species (hydrogen peroxide, aldehydes). It is highly expressed in mammalian tissues, especially in vascular smooth muscle cells and adipocytes, where it plays a role in cell differentiation and glucose transport. The study aims at characterizing the expression and the activity of SSAO in rat and human articular cartilage of the knee, and to investigate its potential role in chondrocyte terminal differentiation. DESIGN: SSAO expression was examined by immunohistochemistry and western blot. Enzyme activity was measured using radiolabeled benzylamine as a substrate. Primary cell cultures of rat chondrocytes were treated for 21 days by a specific SSAO inhibitor, LJP 1586. Terminal chondrocyte differentiation markers were quantified by RT-qPCR. The basal and IL1ß-stimulated glucose transport was monitored by the entrance of (3)[H]2-deoxyglucose in chondrocytes. RESULTS: SSAO was expressed in chondrocytes of rat and human articular cartilage. SSAO expression was significantly enhanced during the hypertrophic differentiation of chondrocytes characterized by an increase in MMP13 and in alkaline phosphatase (ALP) expressions. SSAO inhibition delayed the late stage of chondrocyte differentiation without cell survival alteration and diminished the basal and IL1ß-stimulated glucose transport. Interestingly, SSAO activity was strongly increased in human osteoarthritic cartilage. CONCLUSIONS: SSAO was expressed as an active form in rat and human cartilage. The results suggest the involvement of SSAO in rat chondrocyte terminal differentiation via a modulation of the glucose transport. In man, the increased SSAO activity detected in osteoarthritic patients may trigger hypertrophy and cartilage degeneration.

Human vascular adhesion protein 1 (VAP-1) is a unique sialoglycoprotein that mediates carbohydrate-dependent binding of lymphocytes to endothelial cells.

The regulated interactions of leukocytes with vascular endothelial cells are crucial in controlling leukocyte traffic between blood and tissues. Vascular adhesion protein-1 (VAP-1) is a novel, human endothelial cell molecule that mediates tissue-selective lymphocyte binding. Two species (90 and 170 kD) of VAP-1 exist in lymphoid tissues. Glycosidase digestions revealed that the mature 170-kD form of VAP-1 expressed on the lumenal surfaces of vessels is a heavily sialylated glycoprotein. The sialic acids are indispensable for the function of VAP-1, since the desialylated form of VAP-1 no longer mediates lymphocyte binding. We also show that L-selectin is not required for binding of activated lymphocytes to VAP-1 under conditions of shear stress. The 90-kD form of VAP-1 was only seen in an organ culture model, and may represent a monomeric or proteolytic form of the larger species. These data indicate that L-selectin negative lymphocytes can bind to tonsillar venules via the VAP- 1-mediated pathway. Moreover, our findings extend the role of carbohydrate-mediated binding in lymphocyte-endothelial cell interactions beyond the known selectins. In conclusion, VAP-1 naturally exists as a 170-kD sialoglycoprotein that uses sialic acid residues to interact with its counter-receptors on lymphocytes under nonstatic conditions.

Induction and function of vascular adhesion protein-1 at sites of inflammation.

Emigration of leukocytes from the blood into the tissues is critical in controlling lymphocyte patrolling in different lymphatic organs and in leukocyte accumulation at sites of inflammation. During the first stage of the extravasation process, leukocytes bind to the endothelial lining of vessels. At the molecular level, several adhesion molecules on leukocytes and endothelial cells function as receptor-ligand pairs in mediating this dynamic interaction. Recently, we have identified a novel human endothelial cell molecule, vascular adhesion protein 1 (VAP-1), that mediates lymphocyte binding (Salmi, M., and S. Jalkanen. 1992. Science [Wash. DC] 257:1407). VAP-1 was initially characterized by mAb 1B2 which inhibits lymphocyte adhesion to high endothelial venules (HEV) and to purified VAP-1 protein. Here we report the location and function of VAP-1 in normal and inflamed tissues in humans. VAP-1 is abundant in HEV of lymphatic organs belonging to the peripheral lymph node system, but considerably less is expressed in vessels of mucosa-associated lymphatic tissues. A subset of venules in most normal nonlymphatic tissues like skin, brain, kidney, liver, and heart is also VAP-1 positive. In addition to vessels, VAP-1 is distributed on a few other cell types, most notably in dendritic-like cells of germinal centers. At sites of inflammation, such as in inflammatory bowel diseases and chronic dermatoses, expression of VAP-1 is clearly increased. The induced VAP-1 is functional, since mAb 1B2 inhibits lymphocyte binding to inflamed lamina propria venules by approximately 60%. Thus VAP-1 is an endothelial adhesion molecule that under normal conditions is expressed mainly in HEV of lymphatic tissues. However, expression of functional VAP-1 in vivo is upregulated during an inflammatory reaction at other sites as well. Inducibility of VAP-1 suggests that it may play a significant role, not only in recirculation of lymphocytes, but also in controlling entry of leukocytes into sites of inflammation.

Dual binding capacity of mucosal immunoblasts to mucosal and synovial endothelium in humans: dissection of the molecular mechanisms.

Lymphocytes continuously migrate throughout the body in search of antigens. Virgin lymphocytes recirculate freely between the blood and different lymphatic organs, whereas immunoblasts extravasate preferentially into sites similar to those where they initially responded to antigen. Tissue-specific extravasation of lymphocytes is largely controlled by distinct lymphocyte surface receptors that mediate lymphocyte binding to high endothelial venules (HEV). In the present study, the molecular mechanisms determining the specificity of human mucosal (lamina propria) lymphocyte binding to different endothelial recognition systems were analyzed. Mucosal immunoblasts adhered five times better than small mucosal lymphocytes to mucosal HEV. Importantly, mucosal immunoblasts also bound to synovial HEV almost as efficiently as to mucosal HEV, but they did not adhere to peripheral lymph node HEV. To study the impact of different homing-associated molecules in this dual endothelial binding, we used a gut-derived T cell line and freshly isolated mucosal immunoblasts. Both cell types expressed integrins alpha 4, beta 1, beta 7, and lymphocyte function associated antigen 1 (LFA-1), and were CD44 positive, but practically L-selectin negative. Binding of mucosal immunoblasts to mucosal HEV was almost completely abolished by pretreatment with anti-beta 7 monoclonal antibodies, but it was independent of alpha 4/beta 1 function. In contrast, alpha 4/beta 1 partially mediated immunoblast adherence to synovial HEV, whereas alpha 4/beta 7 had only a minor role in adherence of blasts at this site. CD44 and LFA-1 contributed to HEV-binding both in mucosa and synovium. Taken together, this is the first report that demonstrates a critical role for alpha 4/beta 7 in the binding of gut lymphocytes to mucosal venules in humans. Moreover, a hitherto unknown interaction between mucosal effector cells and synovial endothelial cells was shown to be only partially mediated by the currently known homing receptors. The dual endothelial binding capacity of mucosal blasts may help to explain the pathogenesis of reactive arthritis not uncommonly associated with inflammatory and infectious bowel disease.

CD73 is involved in lymphocyte binding to the endothelium: characterization of lymphocyte-vascular adhesion protein 2 identifies it as CD73.

We have recently described a monoclonal antibody (mAb) 4G4 recognizing a 70-kD molecule constitutively expressed on human endothelial cells and on subpopulations of lymphocytes. We showed that this molecule, which we named lymphocyte-vascular adhesion protein 2 (L-VAP-2), mediates lymphocyte adhesion to cultured endothelial cells. Protein sequencing of tryptic peptides from immunoaffinity-purified L-VAP-2 revealed sequence identity between L-VAP-2 and CD73 (ecto-5'-nucleotidase, E.C.3.1.3.5), and COS cells transfected with a CD73 cDNA were positively stained with the mAb 4G4, which recognizes L-VAP-2. mAb 4G4 was also able to partially inhibit the ecto-5'-nucleotidase activity of peripheral blood lymphocytes. Moreover, cross-precipitation studies performed with mAb 4G4 and a CD73 workshop mAb 1E9 showed that these two antibodies recognize the same molecule. Since the tissue distribution and biochemical characteristics of the two molecules are also similar, we conclude that L-VAP-2 and CD73 are the same glycoprotein. Adhesion experiments showed significantly increased binding of freshly isolated lymphocytes to COS cells transfected with a CD73 cDNA, as compared to mock-transfected COS cells, and binding of lymphocytes to CD73-expressing COS cells was inhibited by the presence of mAb 4G4 in the adhesion assay. CD73 is a glycosyl phosphatidylinositol-linked molecule previously shown to have a cosignalling role in T lymphocyte proliferation. Our data suggest that it also has a function in mediating lymphocyte adhesion to the endothelium.

Vascular adhesion protein 1 (VAP-1) mediates lymphocyte subtype-specific, selectin-independent recognition of vascular endothelium in human lymph nodes.

Interactions between lymphocyte surface receptors and their ligands on vascular endothelial cells regulate the exit of lymphocytes from the circulation. Distinct subsets of mononuclear cells bind to high endothelial venules (HEVs) in different lymphoid organs to a different extent, but the molecular mechanisms behind this selectivity have remained poorly characterized. Here we show that vascular adhesion protein-1 (VAP-1) mediates subtype-specific binding of CD8-positive T cells and natural killer cells to human endothelium. VAP-1-dependent, oligosaccharide-dependent peripheral lymph node (PLN) HEV adhesion under shear was independent of L-selectin, P-selectin glycoprotein ligand 1, and alpha4 integrins, the known lymphocyte receptors involved in the initial recognition of endothelial cells. PLN HEV adhesion was also critically dependent on peripheral lymph node vascular addressins (PNAds), but lymphocyte L-selectin was absolutely required for PNAd binding. Most lymphocytes relied on both PNAd and VAP-1 in HEV binding. The overlapping function of L-selectin ligands and VAP-1 in PLN introduces a new control point into the lymphocyte extravasation process. Finally, intravital microscopy revealed that VAP-1 is involved in initial interactions between human lymphocytes and endothelial cells in inflamed rabbit mesenterial venules in vivo. In conclusion, VAP-1 is a novel contact-initiating ligand that discriminates between different subpopulations of mononuclear cells and is an appealing target for selective modulation of adhesion of CD8- and CD16-positive effector cells.

Cloning of vascular adhesion protein 1 reveals a novel multifunctional adhesion molecule.

Vascular adhesion protein 1 (VAP-1) is a human endothelial sialoglycoprotein whose cell surface expression is induced under inflammatory conditions. It has been shown previously to participate in lymphocyte recirculation by mediating the binding of lymphocytes to peripheral lymph node vascular endothelial cells in an L-selectin-independent fashion. We report here that the VAP-1 cDNA encodes a type II transmembrane protein of 84.6 kD with a single transmembrane domain located at the NH2-terminal end of the molecule and six potential N-glycosylation sites in the extracellular domain. In vivo, the protein exists predominantly as a homodimer of 170-180 kD. Ax endothelial cells transfected with a VAP-1 cDNA express VAP-1 on their cell surface and bind lymphocytes, and the binding can be partially inhibited with anti-VAP-1 mAbs. VAP-1 has no similarity to any currently known adhesion molecules, but has significant identity to the copper-containing amine oxidase family and has a monoamine oxidase activity. We propose that VAP-1 is a novel type of adhesion molecule with dual function. With the appropriate glycosylation and in the correct inflammatory setting, its expression on the lumenal endothelial cell surface allows it to mediate lymphocyte adhesion and to function as an adhesion receptor involved in lymphocyte recirculation. Its primary function in other locations where it is expressed, such as smooth muscle, may depend on its inherent monoamine oxidase activity.

Mannose receptor is a novel ligand for L-selectin and mediates lymphocyte binding to lymphatic endothelium.

Continuous lymphocyte recirculation between blood and lymphoid tissues forms a basis for the function of the immune system. Lymphocyte entrance from the blood into the tissues has been thoroughly characterized, but mechanisms controlling lymphocyte exit from the lymphoid tissues via efferent lymphatics have remained virtually unknown. In this work we have identified mannose receptor (MR) on human lymphatic endothelium and demonstrate its involvement in binding of lymphocytes to lymphatic vessels. We also show that the binding requires L-selectin, and L-selectin and MR form a receptor-ligand pair. On the other hand, L-selectin binds to peripheral lymph node addressins (PNAds) on high endothelial venules (HEVs) that are sites where lymphocytes enter the lymphatic organs. Interestingly, MR is absent from HEVs and PNAds from lymphatic endothelium. Thus, lymphocyte L-selectin uses distinct ligand molecules to mediate binding at sites of lymphocyte entrance and exit within lymph nodes. Taken together, interaction between L-selectin and MR is the first molecularly defined mechanism mediating lymphocyte binding to lymphatic endothelium.

Human T cell clones express functional homing receptors required for normal lymphocyte trafficking.

To function efficiently in vivo, lymphocytes must circulate from the blood into lymphoid tissues and other sites of immune reaction. Herein, we show that human cytotoxic and helper T cell clones and lines, maintained in vitro with IL-2, express the functional capacity to recognize and bind to high endothelial venules (HEV), a capacity essential for lymphocyte exit from the blood, and hence for normal lymphocyte trafficking. The expression of functional homing receptors distinguishes human T cell clones from their murine counterparts, which uniformly lack receptors for HEV and are unable to migrate normally from the blood in vivo. The results raise the possibility that human T cell clones may be more effective in mediating in vivo immune responses than is suggested by murine models.

Lymphocyte CD44 binds the COOH-terminal heparin-binding domain of fibronectin.

The lymphocyte-high endothelial venule (HEV) cell interaction is an essential element of the immune system, as it controls lymphocyte recirculation between blood and lymphoid organs in the body. This interaction involves an 85-95-kD class of lymphocyte surface glycoprotein(s), CD44. A subset of lymphocyte CD44 molecules is modified by covalent linkage to chondroitin sulfate (Jalkanen, S., M. Jalkanen, R. Bargatze, M. Tammi, and E. C. Butcher. 1988. J. Immunol. 141:1615-1623). In this work, we show that removal of chondroitin sulfate by chondroitinase treatment of lymphocytes or incubation of HEV with chondroitin sulfate does not significantly inhibit lymphocyte binding to HEV, suggesting that chondroitin sulfate is not involved in endothelial cell recognition of lymphocytes. Affinity-purified CD44 antigen was, on the other hand, observed to bind native Type I collagen fibrils, laminin, and fibronectin, but not gelatin. Binding to fibronectin was studied more closely, and it was found to be mediated through the chondroitin sulfate-containing form of the molecule. The binding site on fibronectin was the COOH-terminal heparin binding domain, because (a) the COOH-terminal heparin-binding fragment of fibronectin-bound isolated CD44 antigen; (b) chondroitin sulfate inhibited this binding; and (c) finally, the ectodomain of another cell surface proteoglycan, syndecan, which is known to bind the COOH-terminal heparin binding domain of fibronectin (Saunders, S., and M. Bernfield. 1988. J. Cell Biol. 106: 423-430), inhibited binding of CD44 both to intact fibronectin and to its heparin binding domain. Moreover, inhibition studies showed that binding of a lymphoblastoid cell line, KCA, to heparin binding peptides from COOH-terminal heparin binding fragment of fibronectin was mediated via CD44. These findings suggest that recirculating lymphocytes use the CD44 class of molecules not only for binding to HEV at the site of lymphocyte entry to lymphoid organs as reported earlier but also within the lymphatic tissue where CD44, especially the subset modified by chondroitin sulfate, is used for interaction with extracellular matrix molecules such as fibronectin.

Regulated expression of exon v6 containing isoforms of CD44 in man: downregulation during malignant transformation of tumors of squamocellular origin.

CD44 is a family of glycoproteins involved in cell-cell and cell-matrix interactions. In addition to the major 90-kD form present on most hematopoietic cells, larger 140-230 kD forms are found on keratinocytes and carcinoma cell lines. These bigger isoforms of CD44 arise by alternative splicing that results in insertion of one or more of the "variant" exons into the extracellular part of the 90-kD constant form of the molecule. In rat, v6 (variant exon v6) containing form of CD44 confers metastatic potential to carcinoma cells, and therefore, it is of interest to study the distribution of this isoform in humans. We raised antibodies against a synthetic peptide containing a sequence encoded by the exon v6. A mAb thus obtained (designated Var3.1) strongly reacted with the plasma membranes of squamous cells in upper layers of skin and tonsil surface epithelia. Weaker staining was seen in germinal centers, vascular endothelia and enterocytes. Exon v6 containing forms of CD44 (CD44v6) were absent from tissue leukocytes and connective tissue components. In comparison, Hermes-3 epitope (on the constant part) containing forms of CD44 were preferentially localized in basal layers of epithelia, present on the surface on most leukocytes and connective tissue cells, and undetectable on the luminal surface of high endothelial venules. In benign neoplasms, epithelial cells stained with mAb Var3.1 like in normal tissues. In contrast, immunostaining of 30 squamous carcinoma specimens (both primary and metastatic lesions) revealed that malignant transformation resulted in downregulation or disappearance of Var3.1 epitope, but in majority of cases, not in diminished synthesis of the Hermes-3 epitope. Biochemical analyses showed that mAb Var3.1 recognized two major forms of CD44 (220 and 300 kD). In conclusion, epitopes on exon v6 and constant part of CD44 are differentially synthesized and regulated during normal and malignant growth of cells in man.

Evolutionary conservation of tissue-specific lymphocyte-endothelial cell recognition mechanisms involved in lymphocyte homing.

Tissue-specific interactions with specialized high endothelial venules (HEV) direct the homing of lymphocytes from the blood into peripheral lymph nodes, mucosal lymphoid organs, and tissue sites of chronic inflammation. These interactions have been demonstrated in all mammalian species examined and thus appear highly conserved. To assess the degree of evolutionary divergence in lymphocyte-HEV recognition mechanisms, we have studied the ability of lymphocytes to interact with HEV across species barriers. By using an in vitro assay of lymphocyte binding to HEV in frozen sections of lymphoid tissues, we confirm that mouse, guinea pig, and human lymphocytes bind to xenogeneic as well as homologous HEV. In addition, we show that mouse and human lymphoid cell lines that bind selectively to either peripheral lymph node or mucosal vessels (Peyer's patches, appendix) in homologous lymphoid tissues exhibit the same organ specificity in binding to xenogeneic HEV. Furthermore, monoclonal antibodies that recognize lymphocyte "homing receptors" and block homologous lymphocyte binding to peripheral lymph node or to mucosal HEV, also inhibit lymphocyte interactions with xenogeneic HEV in a tissue-specific fashion. Similarly, anti-HEV antibodies against organ-specific mouse high endothelial cell "addressins" involved in lymphocyte homing to peripheral lymph node or mucosal lymphoid organs, not only block the adhesion of mouse lymphocytes but also of human lymphocytes to target mouse HEV. The results illustrate a remarkable degree of functional conservation of elements mediating these cell-cell recognition events involved in organ-specific lymphocyte homing.

Lymphocyte recognition of high endothelium: antibodies to distinct epitopes of an 85-95-kD glycoprotein antigen differentially inhibit lymphocyte binding to lymph node, mucosal, or synovial endothelial cells.

The tissue-specific homing of lymphocytes is directed by specialized high endothelial venules (HEV). At least three functionally independent lymphocyte/HEV recognition systems exist, controlling the extravasation of circulating lymphocytes into peripheral lymph nodes, mucosal lymphoid tissues (Peyer's patches or appendix), and the synovium of inflamed joints. We report here that antibodies capable of inhibiting human lymphocyte binding to one or more HEV types recognize a common 85-95-kD lymphocyte surface glycoprotein antigen, defined by the non-blocking monoclonal antibody, Hermes-1. We demonstrate that MEL-14, a monoclonal antibody against putative lymph node "homing receptors" in the mouse, functionally inhibits human lymphocyte binding to lymph node HEV but not to mucosal or synovial HEV, and cross-reacts with the 85-95-kD Hermes-1 antigen. Furthermore, we show that Hermes-3, a novel antibody produced by immunization with Hermes-1 antigen isolated from a mucosal HEV-specific cell line, selectively blocks lymphocyte binding to mucosal HEV. Such tissue specificity of inhibition suggests that MEL-14 and Hermes-3 block the function of specific lymphocyte recognition elements for lymph node and mucosal HEV, respectively. Recognition of synovial HEV also involves the 85-95-kD Hermes-1 antigen, in that a polyclonal antiserum produced against the isolated antigen blocks all three classes of lymphocyte-HEV interaction. From these studies, it is likely that the Hermes-1-defined 85-95-kD glycoprotein class either comprises a family of related but functionally independent receptors for HEV, or associates both physically and functionally with such receptors. The findings imply that related molecular mechanisms are involved in several functionally independent cell-cell recognition events that direct lymphocyte traffic.

Differential regulation and function of CD73, a glycosyl-phosphatidylinositol-linked 70-kD adhesion molecule, on lymphocytes and endothelial cells.

CD73, otherwise known as ecto-5'-nucleotidase, is a glycosyl-phosphatidylinositol-linked 70-kD molecule expressed on different cell types, including vascular endothelial cells (EC) and certain subtypes of lymphocytes. There is strong evidence for lymphocyte CD73 having a role in several immunological phenomena such as lymphocyte activation, proliferation, and adhesion to endothelium, but the physiological role of CD73 in other cell types is less clear. To compare the biological characteristics of CD73 in different cell types, we have studied the structure, function, and surface modulation of CD73 on lymphocytes and EC. CD73 molecules on lymphocytes are shed from the cell surface as a consequence of triggering with an anti-CD73 mAb, mimicking ligand binding. In contrast, triggering of endothelial CD73 does not have any effect on its expression. Lymphocyte CD73 is susceptible to phosphatidylinositol phospholipase, whereas only a small portion of CD73 on EC could be removed by this enzyme. Furthermore, CD73 on EC was unable to deliver a tyrosine phosphorylation inducing signal upon mAb triggering, whereas triggering of lymphocyte CD73 can induce tyrosine phosphorylation. Despite the functional differences, CD73 molecules on lymphocytes and EC were practically identical structurally, when studied at the protein, mRNA, and cDNA level. Thus, CD73 is an interesting example of a molecule which lacks structural variants but yet has a wide diversity of biological functions. We suggest that the ligand-induced shedding of lymphocyte CD73 represents an important and novel means of controlling lymphocyte-EC interactions.

A 90-kilodalton endothelial cell molecule mediating lymphocyte binding in humans.

Interactions between leukocyte surface receptors and their ligands on vascular endothelial cells control lymphocyte traffic between the blood and various lymphoid organs, as well as extravasation of leukocytes into sites of inflammation. A heretofore undescribed 90-kilodalton human endothelial cell adhesion molecule (VAP-1) defined by a monoclonal antibody 1B2 is described. The expression pattern, molecular mass, functional properties, and an amino-terminal amino acid sequence define VAP-1 as an endothelial ligand for lymphocytes. VAP-1 helps to elucidate the complex heterotypic cell interactions that direct tissue-selective lymphocyte migration in man.

A distinct endothelial cell recognition system that controls lymphocyte traffic into inflamed synovium.

Lymphocytes are essential mediators of normal tissue inflammatory reactions and of pathologic tissue damage in, for example, rheumatoid arthritis and other autoimmune diseases. In a study of the mechanisms controlling lymphocyte entry into sites of inflammation from the blood, the function and specificity of lymphocyte-endothelial interactions were examined in inflamed joint tissue (synovium) from patients with rheumatoid arthritis. Synovial high endothelial venules (HEV) supported the binding of normal peripheral blood lymphocytes in vitro. The characteristics of this binding, which were similar to those of lymphocyte-HEV interactions controlling lymphocyte migration into organized lymphoid tissues, included a requirement for calcium ions, a dependence on metabolic activity, and a preferential adherence of circulating lymphocytes as opposed to immature thymocytes. However, the binding of lymphocytes to synovial HEV was not inhibited by a monoclonal antibody to lymphocyte receptors for lymph node HEV, and synovial HEV failed to bind either lymph node HEV-specific or mucosal HEV-specific B lymphoblastoid cells. The results suggest that a lymphocyte-endothelial cell recognition system that is distinct from such systems in organized lymphoid tissues directs the extravasation of normal lymphocytes as well as pathologically important effector cells into inflamed synovium.

Vascular adhesion protein-1 in human ischaemic stroke.

AIMS: Leukocyte extravasation exacerbates tissue injury after ischaemic stroke. Vascular adhesion protein-1 (VAP-1) is an endothelial adhesion molecule with the potential capacity to guide transmigration of inflammatory cells into ischaemic brain. Moreover, VAP-1 could worsen ischaemic brain injury due to its function as a semicarbazide-sensitive amine oxidase (SSAO) producing toxic metabolites from primary amines. The purpose of this study was to elucidate these aspects of VAP-1-function in the pathogenesis of human ischaemic stroke. METHODS: We studied VAP-1 expression in infarcted and control brains post mortem using immunohistochemistry. Levels of soluble VAP-1 (sVAP-1) in the serum of patients with acute stroke and in control sera were determined using enzyme-linked immunosorbent assay. RESULTS: In the acute phase of ischaemic stroke, the frequency of VAP-1-stained vessels was strongly diminished in the ipsilateral hemisphere but in the contralateral hemisphere it was comparable with the expression in the control brains. In the serum of acute stroke patients with a symptom duration <6 h the level of sVAP-1 was significantly increased (652 +/- 224 ng/ml; mean +/- SD) when compared with an age- and sex-matched control group (542 +/- 104 ng/ml; P < 0.05). CONCLUSIONS: As both cell surface and sVAP-1 possess vasculopathy-promoting SSAO enzymatic activity, our results imply that by inducing SSAO-derived toxic metabolites, VAP-1 might aggravate ischaemic vascular changes. The subsequent release of sVAP-1 into circulation could be further examined as a potential marker of early ischaemic vasculopathy.

Inhibition of semicarbazide-sensitive amine oxidases decreases lymphocyte infiltration in the early phases of rat liver allograft rejection.

Vascular adhesion protein-1 (VAP-1) has been shown to mediate lymphocyte adhesion to endothelia at sites of inflammation in vitro and in vivo. VAP-1 is also an ectoenzyme with semicarbazide-sensitive amine oxidase (SSAO) activity. In this study we investigated whether inhibition of SSAO influences the inflammatory infiltration in acute rat liver allograft rejection. BN recipients of DA liver allografts were treated with 50 mg/kg/d semicarbazide, an inhibitor of SSAO, or similar volumes of saline. 10 rats/group were followed for graft survival, and 10 rats/group were sacrificed on day 7 post-transplantation for histology and T-lymphocyte isolation. The area percentage of portal inflammatory infiltrates in the grafts was assessed from digital photomicrographs. The proportion of CD4-, CD8- and IL2-receptor positive lymphocytes in the graft was quantified with flow cytometry. On day 7, semicarbazide treatment significantly decreased the inflammatory infiltrate area in the grafts. CD4-, CD8- and IL2-receptor positive cells were equally affected. However, animal survival was not affected. Blockade of the enzymatic activity of VAP-1 has a significant effect on lymphocyte infiltration early in acute liver rejection. Later, activation of other adhesion pathways can by-pass the blockade caused by VAP-inhibition.

Homing of mucosal leukocytes to joints. Distinct endothelial ligands in synovium mediate leukocyte-subtype specific adhesion.

Inflammation and infection of the gut can be followed by reactive arthritis at a distant joint. Leukocyte recruitment into synovium is essential for this process, but nothing is known about the endothelial adhesion molecules in synovial membrane which direct the homing of activated, gut-derived leukocytes to joints. Here we analyzed the expression of the known endothelial adhesion molecules in inflamed synovium and their function in binding of mucosal leukocytes. Intercellular adhesion molecule-1 (ICAM-1/CD54) and vascular adhesion protein-1 (VAP-1) were most prominently expressed in synovial vessels. All other adhesion molecules were found at lower levels in inflamed synovia, except mucosal addressin which was absent. Binding of macrophages isolated from lamina propria of the gut to synovial endothelium was almost entirely P-selectin-dependent. In contrast, small intestinal lymphocytes and immunoblasts both relied mainly on VAP-1 in recognition of synovial vessels. Thus, endothelial P-selectin and VAP-1 mediate binding of mucosal effector cells to synovium in a leukocyte subtype-selective manner. Antiadhesive therapy against these inducible molecules should ablate the pathogenetic cascade leading to inappropriate homing of leukocytes to joints in arthritis.