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Journal of Thrombosis and Haemostasis :... Aug 2010Vascular smooth muscle cell (VSMC) migration is a critical process in arterial remodeling. Purified plasminogen activator inhibitor-1 (PAI-1) is reported to both promote...
Plasminogen activator inhibitor-1 and vitronectin expression level and stoichiometry regulate vascular smooth muscle cell migration through physiological collagen matrices.
BACKGROUND
Vascular smooth muscle cell (VSMC) migration is a critical process in arterial remodeling. Purified plasminogen activator inhibitor-1 (PAI-1) is reported to both promote and inhibit VSMC migration on two-dimensional (D) surfaces.
OBJECTIVE
To determine the effects of PAI-1 and vitronectin (VN) expressed by VSMC themselves on migration through physiological collagen matrices.
METHODS
We studied migration of wild-type (WT), PAI-1-deficient, VN-deficient, PAI-1/VN doubly-deficient (DKO) and PAI-1-transgenic (Tg) VSMC through three-D collagen gels.
RESULTS
WT VSMC migrated significantly slower than PAI-1- and VN-deficient VSMC, but significantly faster than DKO VSMC. Experiments with recombinant PAI-1 suggested that basal VSMC PAI-1 expression inhibits migration by binding VN, which is secreted by VSMC and binds collagen. However, PAI-1-over-expressing Tg VSMC migrated faster than WT VSMC. Reconstitution experiments with recombinant PAI-1 mutants suggested that the pro-migratory effect of PAI-1 over-expression required its anti-plasminogen activator (PA) and LDL receptor-related protein (LRP) binding functions, but not VN binding. While promoting VSMC migration in the absence of PAI-1, VN inhibited the pro-migratory effect of active PAI-1.
CONCLUSIONS
In isolation, VN and PAI-1 are each pro-migratory. However, via formation of a high-affinity, non-motogenic complex, PAI-1 and VN each buffers the other's pro-migratory effect. The level of PAI-1 expression by VSMC and the concentration of VN in extracellular matrix are critical determinants of whether PAI-1 and VN promote or inhibit migration. These findings help to rectify previously conflicting reports and suggest that PAI-1/VN stoichiometry plays an important role in VSMC migration and vascular remodeling.
Topics: Animals; Aorta; Cell Movement; Collagen; Gels; Gene Expression Regulation; Humans; Low Density Lipoprotein Receptor-Related Protein-1; Mice; Mice, Inbred C57BL; Mice, Transgenic; Muscle, Smooth; Muscle, Smooth, Vascular; Plasminogen Activator Inhibitor 1; Vitronectin
PubMed: 20492459
DOI: 10.1111/j.1538-7836.2010.03907.x -
British Journal of Haematology Jan 1998Urokinase (uPA) and its receptor (uPAR) have been proposed to be involved in monocyte migration by inducing degradation of matrix proteins. In addition, uPAR is also...
Regulation of the uPAR/uPA system expressed on monocytes by the deactivating cytokines, IL-4, IL-10 and IL-13: consequences on cell adhesion to vitronectin and fibrinogen.
Urokinase (uPA) and its receptor (uPAR) have been proposed to be involved in monocyte migration by inducing degradation of matrix proteins. In addition, uPAR is also implicated in cell adhesion to the vascular wall. The adhesive function of uPAR depends on a direct interaction with vitronectin which is increased by uPA and by modification of cell surface integrin (such as CD11b-CD18) when associated to uPAR. In this study we analysed the role of three deactivating cytokines, IL-4, IL-10 and IL-13, on the surface expression of uPA, uPAR and CD11b by monocytes and their consequences on monocyte adhesion to immobilized fibrinogen and vitronectin. IL-10 induced a decrease in uPA and CD11b after 18 h incubation and a delayed decrease in uPAR which was only significant after 48 h incubation. These results may explain the decrease in monocyte adhesion, which was observed after an 18 h incubation with IL-10, on immobilized vitronectin and fibrinogen. In contrast, IL-4 and IL-13 induced a decrease in uPAR after 18 h and a significant increase in uPA both in the cell lysates and at the cell surface, as well as an increase in cell surface associated CD11b. These cytokines did not modify cell adhesiveness to vitronectin or fibrinogen despite the increase in CD11b-CD18. This could be due to the decrease in uPAR because CD11b-CD18/uPAR forms a cell adhesion complex. In addition, the increase in uPA induced by IL-4 could counterbalance the direct interaction of uPAR with vitronectin. The increase in uPA suggests that IL-4 and IL-13 could induce plaque fissuring by monocytes, whereas IL-10 may induce protection against matrix protein degradation by decreasing uPA.
Topics: Cell Adhesion; Cell Movement; Fibrinogen; Humans; Interleukin-10; Interleukin-13; Interleukin-4; Monocytes; Receptors, Cell Surface; Receptors, Urokinase Plasminogen Activator; Urokinase-Type Plasminogen Activator; Vitronectin
PubMed: 9450789
DOI: 10.1046/j.1365-2141.1998.00528.x -
The Journal of Biological Chemistry May 2007The serine proteinase inhibitor, plasminogen activator inhibitor type-1 (PAI-1), binds to the adhesion protein vitronectin with high affinity at a site that is located...
The serine proteinase inhibitor, plasminogen activator inhibitor type-1 (PAI-1), binds to the adhesion protein vitronectin with high affinity at a site that is located directly adjacent to the vitronectin RGD integrin binding sequence. The binding of PAI-1 to vitronectin sterically blocks integrin access to this site and completely inhibits the binding of purified integrins to vitronectin; however, its inhibition of endothelial and smooth muscle cell adhesion to vitronectin is at most 50-75%. Because PAI-1 binds vitronectin with approximately 10-100-fold higher affinity than purified integrins, we have analyzed the mechanism whereby these cells are able to overcome this obstacle. Our studies exclude proteolytic removal of PAI-1 from vitronectin as the mechanism, and show instead that cell adhesion in the presence of PAI-1 is dependent on integrin-cytoskeleton engagement. Disrupting endothelial or smooth muscle cell actin polymerization and/or focal adhesion assembly reduces cell adhesion to vitronectin in the presence of PAI-1 to levels similar to that observed for the binding of purified integrins to vitronectin. Furthermore, endothelial cell, but not smooth muscle cell adhesion to vitronectin in the presence of PAI-1 requires both polymerized microtubules and actin, further demonstrating the importance of the cytoskeleton for integrin-mediated adhesion. Finally, we show that cell adhesion in the presence of PAI-1 leads to colocalization of PAI-1 with the integrins alphavbeta3 and alphavbeta5 at the cell-matrix interface.
Topics: Actins; Animals; Cell Adhesion; Cytoskeleton; Endothelial Cells; Humans; Integrin alphaVbeta3; Integrins; Myocytes, Smooth Muscle; Plasminogen Activator Inhibitor 1; Protein Binding; Receptors, Vitronectin; Vitronectin
PubMed: 17403662
DOI: 10.1074/jbc.M702125200 -
Frontiers in Immunology 2023Microvascular immunothrombotic dysregulation is a critical process in the pathogenesis of severe systemic inflammatory diseases. The mechanisms controlling...
Microvascular immunothrombotic dysregulation is a critical process in the pathogenesis of severe systemic inflammatory diseases. The mechanisms controlling immunothrombosis in inflamed microvessels, however, remain poorly understood. Here, we report that under systemic inflammatory conditions the matricellular glycoproteinvitronectin (VN) establishes an intravascular scaffold, supporting interactions of aggregating platelets with immune cells and the venular endothelium. Blockade of the VN receptor glycoprotein (GP)IIb/IIIa interfered with this multicellular interplay and effectively prevented microvascular clot formation. In line with these experimental data, particularly VN was found to be enriched in the pulmonary microvasculature of patients with non-infectious (pancreatitis-associated) or infectious (coronavirus disease 2019 (COVID-19)-associated) severe systemic inflammatory responses. Targeting the VN-GPIIb/IIIa axis hence appears as a promising, already feasible strategy to counteract microvascular immunothrombotic dysregulation in systemic inflammatory pathologies.
Topics: Humans; Vitronectin; COVID-19; Blood Platelets; Platelet Glycoprotein GPIIb-IIIa Complex; Microvessels
PubMed: 36845099
DOI: 10.3389/fimmu.2023.1078005 -
American Journal of Respiratory Cell... Jun 2012Vitronectin is present in large concentrations in serum and the extracellular matrix. Although vitronectin is known to modulate neutrophil adhesion and chemotaxis, and...
Vitronectin is present in large concentrations in serum and the extracellular matrix. Although vitronectin is known to modulate neutrophil adhesion and chemotaxis, and to contribute to neutrophil-associated proinflammatory processes, a role in apoptosis has not been demonstrated. In the present studies, we found that neutrophils demonstrated more rapid progression to spontaneous or TNF-related apoptosis-inducing ligand-induced apoptosis when incubated under vitronectin-free conditions than when vitronectin was present. The ability of native vitronectin to delay neutrophil apoptosis was not recapitulated by the vitronectin somatomedin B domain. In contrast, inclusion of the cyclo[Arg-Gly-Asp-D-Phe-Val] peptide in cultures containing vitronectin resulted in enhanced neutrophil apoptosis, showing that the vitronectin RGD motif (Arg-Gly-Asp motif) was responsible for the antiapoptotic effects of vitronectin. Addition of antibodies to β(1), β(3), or β(5), but not to β(2) or β(4) integrins, reversed the ability of vitronectin to diminish neutrophil apoptosis. The ability of vitronectin to enhance neutrophil viability was dependent on activation of phosphatidylinositol 3-kinase and extracellular signal-regulated kinase 1/2 kinases, but not on the p38 kinase. Increased numbers of apoptotic neutrophils were present in the lungs of LPS-treated transgenic vitronectin-deficient mice, as compared with control mice. These results demonstrate a novel antiapoptotic function for vitronectin.
Topics: Animals; Apoptosis; Integrins; Mice; Mice, Inbred C57BL; Neutrophils; Phosphatidylinositol 3-Kinases; Signal Transduction; Vitronectin
PubMed: 22281987
DOI: 10.1165/rcmb.2011-0187OC -
PloS One 2012Oligodendrocyte differentiation is temporally regulated during development by multiple factors. Here, we investigated whether the timing of oligodendrocyte...
Oligodendrocyte differentiation is temporally regulated during development by multiple factors. Here, we investigated whether the timing of oligodendrocyte differentiation might be controlled by neuronal differentiation in cerebellar organotypic cultures. In these cultures, the slices taken from newborn mice show very few oligodendrocytes during the first week of culture (immature slices) whereas their number increases importantly during the second week (mature slices). First, we showed that mature cerebellar slices or their conditioned media stimulated oligodendrocyte differentiation in immature slices thus demonstrating the existence of diffusible factors controlling oligodendrocyte differentiation. Using conditioned media from different models of slice culture in which the number of Purkinje cells varies drastically, we showed that the effects of these differentiating factors were proportional to the number of Purkinje cells. To identify these diffusible factors, we first performed a transcriptome analysis with an Affymetrix array for cerebellar cortex and then real-time quantitative PCR on mRNAs extracted from fluorescent flow cytometry sorted (FACS) Purkinje cells of L7-GFP transgenic mice at different ages. These analyses revealed that during postnatal maturation, Purkinje cells down-regulate Sonic Hedgehog and up-regulate vitronectin. Then, we showed that Sonic Hedgehog stimulates the proliferation of oligodendrocyte precursor cells and inhibits their differentiation. In contrast, vitronectin stimulates oligodendrocyte differentiation, whereas its inhibition with blocking antibodies abolishes the conditioned media effects. Altogether, these results suggest that Purkinje cells participate in controlling the timing of oligodendrocyte differentiation in the cerebellum through the developmentally regulated expression of diffusible molecules such as Sonic Hedgehog and vitronectin.
Topics: Animals; Animals, Newborn; Cell Differentiation; Cell Proliferation; Cerebellum; Down-Regulation; Hedgehog Proteins; Mice; Mice, Transgenic; Oligodendroglia; Purkinje Cells; Up-Regulation; Vitronectin
PubMed: 23155445
DOI: 10.1371/journal.pone.0049015 -
FEBS Letters Aug 2010Vitronectin is a multi-functional protein found predominantly as a monomer in blood and as an oligomer in the extracellular matrix. We have dissected the minimal regions...
Vitronectin is a multi-functional protein found predominantly as a monomer in blood and as an oligomer in the extracellular matrix. We have dissected the minimal regions of vitronectin protein needed for effective integrin dependent cell adhesion and spreading. A fragment of vitronectin containing the RGD integrin binding site showed similar binding affinity as that of full vitronectin protein to purified integrin alphavbeta3 but had diminished cell adhesion and spreading function in vivo. We demonstrate that the oligomeric state of the protein is responsible for this effect. We provide compelling evidence for the involvement of the heparin binding domain of vitronectin in the oligomerization process and show that such oligomerization reinforces the activity of vitronectin in cell adhesion and spreading.
Topics: Cell Adhesion; Cell Movement; Cell Shape; Chromatography, Gel; Electrophoresis, Polyacrylamide Gel; Fibroblasts; Heparin; Humans; Integrin alphaVbeta3; Peptide Fragments; Protein Binding; Protein Structure, Quaternary; Protein Structure, Tertiary; Recombinant Proteins; Structure-Activity Relationship; Vitronectin
PubMed: 20600001
DOI: 10.1016/j.febslet.2010.06.023 -
Current Protocols in Stem Cell Biology Sep 2020The discovery of induced pluripotent stem cells (iPSCs) revolutionized the approach to cell therapy in regenerative medicine. Reprogramming of somatic cells into an...
The discovery of induced pluripotent stem cells (iPSCs) revolutionized the approach to cell therapy in regenerative medicine. Reprogramming of somatic cells into an embryonic-like pluripotent state provides an invaluable resource of patient-specific cells of any lineage. Implementation of procedures and protocols adapted to current good manufacturing practice (cGMP) requirements is critical to ensure robust and consistent high-quality iPSC manufacturing. The technology developed at Allele Biotechnology for iPSC generation under cGMP conditions is a powerful platform for derivation of pluripotent stem cells through a footprint-free, feeder-free, and xeno-free reprogramming method. The cGMP process established by Allele Biotechnology entails fully cGMP compliant iPSC lines where the entire manufacturing process, from tissue collection, cell reprogramming, cell expansion, cell banking and quality control testing are adopted. Previously, we described in this series of publications how to create iPSCs using mRNA only, and how to do so under cGMP conditions. In this article, we describe in detail how to culture, examine and storage cGMP-iPSCs using reagents, materials and equipment compliant with cGMP standards. © 2020 The Authors. Basic Protocol 1: iPSC Dissociation Support Protocol 1: Stem cell media Support Protocol 2: ROCK inhibitor preparation Support Protocol 3: Vitronectin coating Basic Protocol 2: iPSC Cryopreservation Basic Protocol 3: iPSC Thawing.
Topics: Cell Culture Techniques; Cell Shape; Cryopreservation; Culture Media; Cyclic GMP; Humans; Induced Pluripotent Stem Cells; Protein Kinase Inhibitors; Vitronectin
PubMed: 32649060
DOI: 10.1002/cpsc.117 -
Journal of Zhejiang University.... Nov 2012This study was aimed at assessing the dynamics of vitronectin (VN), laminin (LN), and heparan sulfate/heparin (HS/HP) content changes during experimental burn healing.
OBJECTIVE
This study was aimed at assessing the dynamics of vitronectin (VN), laminin (LN), and heparan sulfate/heparin (HS/HP) content changes during experimental burn healing.
METHODS
VN, LN, and HS/HP were isolated and purified from normal and injured skin of domestic pigs, on the 3rd, 5th, 10th, 15th, and 21st days following thermal damage. The wounds were treated with apitherapeutic agent (propolis), silver sulfadiazine (SSD), physiological salt solution, and propolis vehicle. VN and LN were quantified using an immunoenzymatic assay and HS/HP was estimated by densitometric analysis.
RESULTS
Propolis treatment stimulated significant increases in VN, LN, and HS/HP contents during the initial phase of study, followed by a reduction in the estimated extracellular matrix molecules. Similar patterns, although less extreme, were observed after treatment with SSD.
CONCLUSIONS
The beneficial effects of propolis on experimental wounds make it a potential apitherapeutic agent in topical burn management.
Topics: Animals; Anti-Infective Agents, Local; Burns; Heparin; Laminin; Propolis; Silver Sulfadiazine; Swine; Vitronectin; Wound Healing
PubMed: 23125086
DOI: 10.1631/jzus.B1100310 -
Urology Journal Mar 2016To detect the expression of vitronectin (VTN) in the tissues and blood serum of prostate cancer (PCa) patients, and evaluate its clinical significance and to evaluate...
Evaluation of Vitronectin Expression in Prostate Cancer and the Clinical Significance of the Association of Vitronectin Expression with Prostate Specific Antigen in Detecting Prostate Cancer.
PURPOSE
To detect the expression of vitronectin (VTN) in the tissues and blood serum of prostate cancer (PCa) patients, and evaluate its clinical significance and to evaluate the significance of the combined assay of VTN and prostate specific antigens (PSA) in PCa diagnosis.
MATERIALS AND METHODS
To detect the expression of VTN as a potential marker for PCa diagnosis and prognosis, immunohistochemistry was performed on the tissues of 32 patients with metastatic PCa (PCaM), 34 patients with PCa without metastasis (PCa), and 41 patients with benign prostatic hyperplasia (BPH). The sera were then subjected to Western blot analysis. All cases were subsequently examined to determine the concentrations of PSA and VTN in the sera. The collected data were collated and analyzed.
RESULTS
The positive expression rates of VTN in the tissues of the BPH and PCa groups (including PCa and PCaM groups) were 75.61% and 45.45%, respectively (P = .005). VTN was more highly expressed in the sera of the BPH patients (0.83 ± 0.07) than in the sera of the PCa patients (0.65 ± 0.06) (P < .05). It was also more highly expressed in the sera of the PCa patients than in the sera of the PCaM patients (0.35 ± 0.08) (P < .05). In the diagnosis of BPH and PCa, the Youden indexes of PSA detection, VTN detection, and combined detection were 0.2620, 0.3468, and 0.5635; the kappa values were 0.338, 0.304, and 0.448, respectively, and the areas under the receiver operating characteristic curve were 0.625, 0.673, and 0.703 (P < .05), respectively.
CONCLUSION
VTN levels in sera may be used as a potential marker of PCa for the diagnosis and assessment of disease progression and metastasis. The combined detection of VTN and PSA in sera can be clinically applied in PCa diagnosis. .
Topics: Aged; Biomarkers, Tumor; Biopsy; Blotting, Western; Humans; Immunohistochemistry; Male; Prognosis; Prostate; Prostate-Specific Antigen; Prostatic Neoplasms; ROC Curve; Vitronectin
PubMed: 26945657
DOI: No ID Found