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Biomedical subjects

Marko Salmi

Publications and source records attributed to Marko Salmi.

At least 19 recordsLinked to original sources
PubMed2006

Developmental regulation of the adhesive and enzymatic activity of vascular adhesion protein-1 (VAP-1) in humans.

Vascular adhesion protein-1 (VAP-1) is a homodimeric glycoprotein that belongs to a unique subgroup of cell-surface-expressed oxidases. In adults, endothelial VAP-1 supports leukocyte rolling, firm adhesion, and transmigration in both enzyme activity-dependent and enzyme activity-independent manner. Here we studied the induction and function of VAP-1 during human ontogeny. We show that VAP-1 is already found in the smooth muscle at embryonic week 7. There are marked time-dependent switches in VAP-1 expression in the sinusoids of the liver, in the peritubular capillaries of the kidney, in the capillaries of the heart, and in the venules in the lamina propria of the gut. Fetal VAP-1 is dimerized, and it is enzymatically active. VAP-1 in fetal-type venules is able to bind cord blood lymphocytes. Also, adenovirally transfected VAP-1 on human umbilical vein endothelial cells is involved in rolling and firm adhesion of cord blood lymphocytes under conditions of physiologic shear stress. We conclude that VAP-1 is synthesized from early on in human vessels and it is functionally intact already before birth. Thus, VAP-1 may contribute critically to the oxidase activities in utero, and prove important for lymphocyte trafficking during human ontogeny.

Amine Oxidase (Copper-Containing)↗
PubMed2006

Liver steatosis coexists with myocardial insulin resistance and coronary dysfunction in patients with type 2 diabetes.

Nonalcoholic fatty liver (NAFL) is a common comorbidity in patients with type 2 diabetes and links to the risk of coronary syndromes. The aim was to determine the manifestations of metabolic syndrome in different organs in patients with liver steatosis. We studied 55 type 2 diabetic patients with coronary artery disease using positron emission tomography. Myocardial perfusion was measured with [15O]H2O and myocardial and skeletal muscle glucose uptake with 2-deoxy-2-[18F]fluoro-D-glucose during hyperinsulinemic euglycemia. Liver fat content was determined by magnetic resonance proton spectroscopy. Patients were divided on the basis of their median (8%) into two groups with low (4.6 +/- 2.0%) and high (17.4 +/- 8.0%) liver fat content. The groups were well matched for age, BMI, and fasting plasma glucose. In addition to insulin resistance at the whole body level (P = 0.012) and muscle (P = 0.002), the high liver fat group had lower insulin-stimulated myocardial glucose uptake (P = 0.040) and glucose extraction rate (P = 0.0006) compared with the low liver fat group. In multiple regression analysis, liver fat content was the most significant explanatory variable for myocardial insulin resistance. In addition, the high liver fat group had increased concentrations of high sensitivity C-reactive protein, soluble forms of E-selectin, vascular adhesion protein-1, and intercellular adhesion molecule-1 (P < 0.05) and lower coronary flow reserve (P = 0.02) compared with the low liver fat group. In conclusion, in patients with type 2 diabetes and coronary artery disease, liver fat content is a novel independent indicator of myocardial insulin resistance and reduced coronary functional capacity. Further studies will reveal the effect of hepatic fat reduction on myocardial metabolism and coronary function.

Biomarkers↗
PubMed2006

Vascular amine oxidases are needed for leukocyte extravasation into inflamed joints in vivo.

OBJECTIVE: Leukocyte traffic from the blood to the joints is crucial in the pathogenesis of arthritis. A bifunctional endothelial cell-surface glycoprotein, AOC3 (amine oxidase, copper-containing 3; also known as vascular adhesion protein 1), has both adhesive and enzymatic properties. We undertook this study to determine the contribution of AOC3 and its oxidase activity to leukocyte trafficking into inflamed joints in vivo. METHODS: We used gene-modified animals, molecular modeling, an AOC3 enzyme inhibitor, oxidase assays, and arthritis models (adjuvant-induced arthritis [AIA] in rats and anti-type II collagen antibody-induced arthritis in mice) to dissect the importance of AOC3 in vivo. RESULTS: The AOC3 inhibitor fitted well with a covalent binding mode into the active site of the AOC3 crystal structure. It selectively blocked the oxidase activity of AOC3 in enzyme assays. Intraperitoneal and oral administration of the AOC3 inhibitor significantly ameliorated rat AIA. In anti-type II collagen antibody-induced arthritis in mice, the AOC3 inhibitor also improved the outcome of the joint inflammation. The acute semicarbazide-sensitive amine oxidase blockade by the inhibitor had even more pronounced effects than genetic deletion of AOC3. Enzymatic analyses showed that the inhibitor also blocked 2 other structurally very closely related AOCs, but not any of more than 100 other enzymes tested. CONCLUSION: These are the first data to demonstrate that the enzymatic activity of the atypical endothelial adhesion molecule AOC3, and possibly that of other closely related ecto-oxidases, is crucial for leukocyte exit from the vessels in inflamed joints in vivo.

Amine Oxidase (Copper-Containing)↗
PubMed2005

Molecular identification of PAL-E, a widely used endothelial-cell marker.

The pathologische anatomie Leiden-endothelium (PAL-E) antibody has been used for almost 20 years as a specific marker for vascular endothelial cells. Due to the fact that this antibody works only in very limited applications, the molecular identity of PAL-E has remained unknown. In this work, we demonstrate by double stainings, cross-immunoprecipitations, and transfectants that the PAL-E antigen is identical with a protein designated PV-1 (plasmalemmal vesicle 1) or FELS (fenestrated endothelial-linked structure protein) and is not vimentin, as reported earlier. As the expression of this molecule is by no means restricted to fenestrated endothelium, we suggest the use of the name PLVAP for this protein. Molecular identification of PLVAP should help in the production of new tools for the identification of vascular as opposed to lymphatic endothelium and to elucidate the function of this protein.

Animals↗
PubMed2005

Functional analysis of Peutz-Jeghers mutations reveals that the LKB1 C-terminal region exerts a crucial role in regulating both the AMPK pathway and the cell polarity.

Germline mutations of the LKB1 gene are responsible for the cancer-prone Peutz-Jeghers syndrome (PJS). LKB1 encodes a serine-threonine kinase that acts as a regulator of cell cycle, metabolism and cell polarity. The majority of PJS missense mutations abolish LKB1 enzymatic activity and thereby impair all functions assigned to LKB1. Here, we have investigated the functional consequences of recurrent missense mutations identified in PJS and in sporadic tumors which map in the LKB1 C-terminal non-catalytic region. We report that these C-terminal mutations neither disrupt LKB1 kinase activity nor interfere with LKB1-induced growth arrest. However, these naturally occuring mutations lessened LKB1-mediated activation of the AMP-activated protein kinase (AMPK) and impaired downstream signaling. Furthermore, C-terminal mutations compromise LKB1 ability to establish and maintain polarity of both intestinal epithelial cells and migrating astrocytes. Consistent with these findings, mutational analysis reveals that the LKB1 tail exerts an essential function in the control of cell polarity. Overall, our results ascribe a crucial regulatory role to the LKB1 C-terminal region. Our findings further indicate that LKB1 tumor suppressor activity is likely to depend on the regulation of AMPK signaling and cell polarization.

AMP-Activated Protein Kinase Kinases↗
PubMed2005

Function-blocking antibodies to human vascular adhesion protein-1: a potential anti-inflammatory therapy.

Human vascular adhesion protein-1 (VAP-1) is a homodimeric 170-kDa sialoglycoprotein that is expressed on the surface of endothelial cells and functions as a semicarbazide-sensitive amine oxidase and as an adhesion molecule. Blockade of VAP-1 has been shown to reduce leukocyte adhesion and transmigration in in vivo and in vitro models, suggesting that VAP-1 is a potential target for anti-inflammatory therapy. In this study we have constructed mouse-human chimeric antibodies by genetic engineering in order to circumvent the potential problems involved in using murine antibodies in man. Our chimeric anti-VAP-1 antibodies, which were designed to lack Fc-dependent effector functions, bound specifically to cell surface-expressed recombinant human VAP-1 and recognized VAP-1 in different cell types in tonsil. Furthermore, the chimeric antibodies prevented leukocyte adhesion and transmigration in vitro and in vivo. Hence, these chimeric antibodies have the potential to be used as a new anti-inflammatory therapy.

Amine Oxidase (Copper-Containing)↗
PubMed2005

Absence of the endothelial oxidase AOC3 leads to abnormal leukocyte traffic in vivo.

Leukocyte migration from the blood to tissues is a prerequisite for normal immune responses. We produced mice deficient in an endothelial cell-surface oxidase (amine oxidase, copper containing-3 [AOC3], also known as vascular adhesion protein-1 [VAP-1]) and found that this enzyme is needed for leukocyte extravasation in vivo. Real-time imaging shows that AOC3 mediates slow rolling, firm adhesion, and transmigration of leukocytes in vessels at inflammatory sites and lymphoid tissues. Absence of AOC3 results in reduced lymphocyte homing into lymphoid organs and in attenuated inflammatory response in peritonitis. These data alter the paradigm of leukocyte extravasation cascade by providing the first physiological proof for the concept that endothelial cell surface enzymes regulate the development of inflammatory reactions in vivo and suggest that this enzyme should be useful as an anti-inflammatory target.

Amine Oxidase (Copper-Containing)↗
PubMed2005

Rules of recruitment for Th1 and Th2 lymphocytes in inflamed liver: a role for alpha-4 integrin and vascular adhesion protein-1.

The mechanisms that mediate the recruitment of Th1 and Th2 lymphocytes in vivo are poorly understood. We demonstrate that the mechanisms by which exogenously produced CD4(+) Th1 and Th2 cells roll and adhere in Con A-inflamed liver microcirculation differ dramatically: Th1 cells use alpha(4)beta(1)-integrin and Th2 cells use the vascular adhesion protein (VAP)-1. P-selectin plays no detectable role in Th1 or Th2 cell trafficking in liver microcirculation. Cellular recruitment in the liver sinusoids has previously been shown to be independent of many known adhesion molecules, leading to the suggestion that recruitment in these structures is mediated by physical trapping. While this may still be true for neutrophils, Th1 and Th2 cells use alpha(4)-integrin and VAP-1, respectively, to adhere within the liver sinusoids.

Amine Oxidase (Copper-Containing)↗
PubMed2005

Vascular adhesion protein-1 is involved in both acute and chronic inflammation in the mouse.

Vascular adhesion protein-1 (VAP-1) is an endothelial molecule that possesses both adhesive and enzymatic properties in vitro. So far, however, elucidation of its in vivo function has suffered from the lack of function-blocking reagents that are suitable for use in animal models. In this work we produced monoclonal antibodies against murine VAP-1 and characterized them using in vitro binding assays. We then examined whether the antibodies could prevent leukocyte migration in in vivo inflammation models, including two acute models (peritonitis induced with proteose peptone and interleukin-1 and air pouch inflammation enhanced by CCL21) and one chronic model (autoimmune diabetes in nonobese diabetic mice). Antibodies 7-88 and 7-106 inhibited migration of granulocytes and monocytes in both acute models of inflammation. Strikingly, antibody 7-88 significantly prevented diabetes in a subset of nonobese diabetic mice. The results show for the first time that in mouse models of inflammation, VAP-1 mediates leukocyte trafficking to sites of inflammation and thus is a potential target for anti-inflammatory therapies.

Acute Disease↗
PubMed2005

Cell-surface enzymes in control of leukocyte trafficking.

Leukocyte trafficking between the blood and the tissues is pivotal for normal immune responses. Cell-adhesion molecules (such as selectins and leukocyte integrins) and chemoattractants (such as chemokines) have well-established roles in supporting leukocyte exit from the blood. Emerging data now show that, for both leukocytes and endothelial cells, enzymatic reactions that are catalysed by cell-surface-expressed enzymes with catalytic domains outside the plasma membrane (known as ectoenzymes) also make crucial contributions to this process. Ectoenzymes can function physically as adhesion receptors and can regulate the recruitment of cells through their catalytic activities. Here, we provide new insights into how ectoenzymes--including nucleotidases, cyclases, ADP-ribosyltransferases, peptidases, proteases and oxidases--guide leukocyte traffic.

Animals↗
PubMed2005

Crystal structure of the human vascular adhesion protein-1: unique structural features with functional implications.

The expression of human vascular adhesion protein-1 (hVAP-1) is induced at sites of inflammation where extravasation of lymphocytes from blood to the peripheral tissue occurs. We have solved the X-ray structure of hVAP-1, a human copper amine oxidase (CAO), which is distinguished from other CAOs in being membrane-bound. The dimer structure reveals some intriguing features that may have fundamental roles in the adhesive and enzymatic functions of hVAP-1, especially regarding the role of hVAP-1 in inflammation, lymphocyte attachment, and signaling. Firstly, Leu469 at the substrate channel may play a key role in controlling the substrate entry; depending on its conformation, it either blocks or gives access to the active site. Secondly, sugar units are clearly observed at two of the six predicted N-glycosylation sites. Moreover, mutagenesis analysis showed that all of the predicted sites were glycosylated in the protein used for crystallization. Thirdly, the existence of a solvent-exposed RGD motif at the entrance to each active site in hVAP-1 suggests that it may have a functional role.

Amine Oxidase (Copper-Containing)↗
PubMed2005

Lymphocyte homing to the gut: attraction, adhesion, and commitment.

Lymphocytes continuously migrate from the blood into the intestine. Naive lymphocytes leave the blood through high endothelial venules in Peyer's patches. During the multistep extravasation cascade, they sequentially roll on, firmly adhere to, and transmigrate through the endothelial layer using multiple adhesion molecules and chemotactic signals. In the organized lymphoid tissues of the gut, lymphocytes can become activated, if they meet their cognate antigens transported to Peyer's patches through the gut epithelium. During activation and proliferation, the lymphocytes become imprinted by the local dendritic cells, so that after returning to systemic circulation via the efferent lymphatic vasculature, they preferentially home to lamina propria of the gut to execute their effector functions. In inflammation, the recirculation routes of lymphocytes are altered, and these may explain the pathogenesis of certain extra-intestinal manifestations of gut infections and inflammatory bowel diseases. The increased knowledge on the mechanisms that regulate lymphocyte homing and imprinting has clear applicability in designing more effective vaccination regimens. A detailed understanding of the mucosal homing has recently led to the development of the first successful anti-adhesive therapeutics in human.

Animals↗
PubMed2004

CLEVER-1 mediates lymphocyte transmigration through vascular and lymphatic endothelium.

Common lymphatic endothelial and vascular endothelial receptor-1 (CLEVER-1; also known as stabilin-1 or FEEL-1) is a large multifunctional glycoprotein implicated in scavenging, angiogenesis, and cell adhesion. Here we studied the function of human CLEVER-1 in leukocyte trafficking. Lymphatic vessels expressed CLEVER-1 constitutively in skin in vivo, whereas on vascular endothelium it appeared only upon inflammation. On isolated vascular endothelial cells, CLEVER-1 supported rolling and transmigration of peripheral blood mononuclear cells (PBMCs) under physiologically relevant laminar shear stress. Intriguingly, CLEVER-1 also mediated transmigration of leukocytes through cultured lymphatic endothelium under static conditions. Thus, synthesis of CLEVER-1 is differentially regulated on the 2 anatomically distinct vascular beds, and CLEVER-1 mediates the transmigration step of the leukocyte traffic in both of them. Notably, CLEVER-1 is the first adhesion molecule shown to be involved in the PBMC transmigration through the lymphatic arm of the immune system.

Capillaries↗
PubMed2004

Granulocyte transmigration through the endothelium is regulated by the oxidase activity of vascular adhesion protein-1 (VAP-1).

Polymorphonuclear leukocytes (PMNs) migrate from the blood into areas of inflammation by binding to the endothelial cells of blood vessels via adhesion molecules. Vascular adhesion protein-1 (VAP-1) is one of the molecules mediating leukocyte-endothelial cell interactions. It is also an endothelial cell-surface enzyme (amine oxidase) that produces reactive oxygen species during the catalytic reaction. To study the role of the enzymatic activity of VAP-1 in PMN extravasation, we used an enzymatically inactive VAP-1 mutant, specific amine oxidase inhibitors (including a novel small molecule compound), and anti-VAP-1 antibodies in several flow-dependent models. The enzyme inhibitors diminished PMN rolling on and transmigration through human endothelial cells under conditions of laminar shear stress in vitro. Notably, the enzyme inactivating point mutation abolished the capacity of VAP-1 to mediate transmigration. Moreover, the new VAP-1 inhibitor effectively prevented the extravasation of PMNs in an animal model of inflammation. These data show that the oxidase activity of VAP-1 controls PMN exit from the blood during the relatively poorly understood transmigration step.

Amine Oxidase (Copper-Containing)↗
PubMed2004

A peptide inhibitor of vascular adhesion protein-1 (VAP-1) blocks leukocyte-endothelium interactions under shear stress.

Vascular adhesion protein-1 (VAP-1) is an endothelial adhesion molecule mediating leukocyte interactions with blood vessels during leukocyte extravasation. Molecularly VAP-1 is a cell-surface-expressed ecto-enzyme belonging to the group of semicarbazide-sensitive amine oxidases (SSAO; EC 2.4.6.3), which deaminate primary amines. Here we asked whether peptides displaying a suitable free amine group could be a substrate or inhibitor of SSAO and thus regulate VAP-1-mediated leukocyte adhesion. On the basis of a molecular model of VAP-1, we designed synthetic peptides that fit to the substrate channel of VAP-1. One of these lysine-containing peptides effectively inhibits VAP-1-dependent lymphocyte rolling and firm adhesion to primary endothelial cells under physiologically relevant shear conditions. The same peptide inhibits the SSAO activity of endothelial and recombinant VAP-1 in a selective and long-lasting manner. We also show that all enzymatically active VAP-1 is displayed on the cell surface. Our results suggest that, in addition to soluble amines, specific cell-surface-bound molecules containing free NH(2) groups in a suitable position may modulate the enzymatic activity of SSAO. Moreover, the inhibitory peptide diminishes leukocyte interactions with endothelial cells under conditions of shear, and thus it may be useful to treat inflammatory conditions.

Amine Oxidase (Copper-Containing)↗
PubMed2004

Blockade of vascular adhesion protein-1 inhibits lymphocyte infiltration in rat liver allograft rejection.

Vascular adhesion protein-1 (VAP-1) has been shown to mediate lymphocyte adhesion to endothelia at sites of inflammation, but its functional role in vivo has not been tested in any rodent model. Here we report the effects of VAP-1 blockade on rat liver allograft rejection. BN recipients of PVG liver allografts (known to develop acute rejection by day 7) were treated with 2 mg/kg anti-VAP-1 (a new anti-rat VAP-1 mAb 174-5) or isotype-matched irrelevant antibody (NS1) every other day (n = 6/group) and one group with anti-VAP-1 2 mg/kg daily (n = 7). On day 7, samples were collected for transplant aspiration cytology, histology, and immunohistochemistry. Lymphocyte infiltration to the graft was clearly affected by VAP-blockade. The total inflammation, mainly the number of active lymphoid cells, in transplant aspiration cytology was significantly decreased in animals treated with anti-VAP-1 (4.7 +/- 1.0 and 2.4 +/- 1.0 corrected increment units, respectively) compared to control (6.6 +/- 1.0) (P < 0.05). In histology, the intensity of portal inflammation was significantly decreased (P < 0.05). The amount of T cells expressing activation markers diminished. This is the first demonstration in any prolonged in vivo model that VAP-1 plays an important role in lymphocyte infiltration to sites of inflammation, and, in particular, liver allograft rejection.

Amine Oxidase (Copper-Containing)↗
PubMed2003

The same endothelial receptor controls lymphocyte traffic both in vascular and lymphatic vessels.

The mechanisms controlling the exit of lymphocytes from tissues via lymphatics are practically unknown. We have now identified a 270-300-kDa molecule designated common lymphatic endothelial and vascular endothelial receptor-1 (CLEVER-1) on human lymphatic endothelium and high endothelial venules. We show that it mediates binding of lymphocytes both to high endothelial venules and to lymphatic vessels. Moreover, blocking of the function of CLEVER-1 results in significant reduction of lymphocyte traffic in vivo. Notably, CLEVER-1 is also an inducible vascular adhesion molecule for other classes of leukocytes at sites of inflammation in peripheral tissues. These findings suggest that CLEVER-1 is involved in regulation of lymphocyte recirculation and migration of leukocytes to sites of inflammation and is a potential new target to control inflammation.

Animals↗