Academic literature on the topic 'Fibronectin Aggregation in Collagen'
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Journal articles on the topic "Fibronectin Aggregation in Collagen"
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Moon, DG, JE Kaplan, and JE Mazurkewicz. "The inhibitory effect of plasma fibronectin on collagen-induced platelet aggregation." Blood 67, no. 2 (February 1, 1986): 450–57. http://dx.doi.org/10.1182/blood.v67.2.450.450.
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Abstract Plasma fibronectin (Fn) has been proposed to have an antithrombotic effect, protecting against platelet and fibrinogen consumption after injury. The current study was designed to determine the effect of plasma fibronectin on collagen-induced platelet aggregation. In vitro aggregometry using an isolated homologous rat system, demonstrated a significant (P less than .05) inhibitory effect of 120 micrograms/mL Fn on platelet aggregation as induced by 60 micrograms/mL fibrillar collagen (type I). The inhibition was evidenced by a threefold increase in lag time and a significant decrease in the rate and extent of aggregation. The hypothesis was also tested using an in vivo model of collagen-induced platelet aggregation. The model used was intravenous injection of 2 mg/kg of homologous type I collagen into anesthetized Sprague-Dawley rats. Injection of collagen preincubated with 4 mg/kg Fn resulted in significantly less thrombocytopenia and fibrinogen consumption as compared with injection of collagen alone. The results of both the in vitro and in vivo studies are consistent with the proposed antithrombotic effect of plasma fibronectin.
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Moon, DG, JE Kaplan, and JE Mazurkewicz. "The inhibitory effect of plasma fibronectin on collagen-induced platelet aggregation." Blood 67, no. 2 (February 1, 1986): 450–57. http://dx.doi.org/10.1182/blood.v67.2.450.bloodjournal672450.
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Plasma fibronectin (Fn) has been proposed to have an antithrombotic effect, protecting against platelet and fibrinogen consumption after injury. The current study was designed to determine the effect of plasma fibronectin on collagen-induced platelet aggregation. In vitro aggregometry using an isolated homologous rat system, demonstrated a significant (P less than .05) inhibitory effect of 120 micrograms/mL Fn on platelet aggregation as induced by 60 micrograms/mL fibrillar collagen (type I). The inhibition was evidenced by a threefold increase in lag time and a significant decrease in the rate and extent of aggregation. The hypothesis was also tested using an in vivo model of collagen-induced platelet aggregation. The model used was intravenous injection of 2 mg/kg of homologous type I collagen into anesthetized Sprague-Dawley rats. Injection of collagen preincubated with 4 mg/kg Fn resulted in significantly less thrombocytopenia and fibrinogen consumption as compared with injection of collagen alone. The results of both the in vitro and in vivo studies are consistent with the proposed antithrombotic effect of plasma fibronectin.
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Eynard, A. R., M. E. Pasqualini, and R. A. Rovasio. "Exogenous fibronectin modifies the aggregation of collagen-stimulated human platelets." Experientia 46, no. 7 (July 1990): 680–82. http://dx.doi.org/10.1007/bf01939932.
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O'Shea, K. S. "Differential deposition of basement membrane components during formation of the caudal neural tube in the mouse embryo." Development 99, no. 4 (April 1, 1987): 509–19. http://dx.doi.org/10.1242/dev.99.4.509.
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The distribution of basement membrane and extracellular matrix components laminin, fibronectin, type IV collagen and heparan sulphate proteoglycan was examined during posterior neuropore closure and secondary neurulation in the mouse embryo. During posterior neuropore closure, these components were densely deposited in basement membranes of neuroepithelium, blood vessels, gut and notochord; although deposition was sparse in the midline of the regressing primitive streak. During secondary neurulation, mesenchymal cells formed an initial aggregate near the dorsal surface, which canalized and merged with the anterior neuroepithelium. With aggregation, fibronectin and heparan sulphate proteoglycan were first detected at the base of a 3- to 4-layer zone of radially organized cells. With formation of a lumen within the aggregate, laminin and type IV collagen were also deposited in the forming basement membrane. During both posterior neuropore closure and secondary neurulation, fibronectin and heparan sulphate proteoglycan were associated with the most caudal portion of the neuroepithelium, the region where newly formed epithelium merges with the consolidated neuroepithelium. In regions of neural crest migration, the deposition of basement membrane components was altered, lacking laminin and type IV collagen, with increased deposition of fibronectin and heparan sulphate proteoglycan.
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Nitkin, R. M., and T. C. Rothschild. "Agrin-induced reorganization of extracellular matrix components on cultured myotubes: relationship to AChR aggregation." Journal of Cell Biology 111, no. 3 (September 1, 1990): 1161–70. http://dx.doi.org/10.1083/jcb.111.3.1161.
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Agrin, an extracellular matrix-associated protein extracted from synapse-rich tissues, induces the accumulation of acetylcholine receptors (AChRs) and other synaptic components into discrete patches on cultured myotubes. The appearance of agrin-like molecules at neuromuscular junctions suggests that it may direct synaptic organization in vivo. In the present study we examined the role of extracellular matrix components in agrin-induced differentiation. We used immunohistochemical techniques to visualize the spatial and temporal distribution of laminin, a heparan sulfate proteoglycan (HSPG), fibronectin, and type IV collagen on cultured chick myotubes during agrin-induced aggregation of AChRs. Myotubes displayed significant amounts of laminin and HSPG, lesser amounts of type IV collagen, and little, if any, fibronectin. Agrin treatment caused cell surface laminin and HSPG to patch, while collagen and fibronectin distributions were generally unaffected. Many of the agrin-induced laminin and HSPG patches colocalized with AChR patches, raising the possibility of a causal relationship between matrix patching and AChR accumulations. However, patching of AChRs (complete within a few hours) preceded that of laminin or HSPG (not complete until 15-20 h), making it unlikely that matrix accumulations initiate AChR patching at agrin-induced sites. Conversely, when AChR patching was blocked by treatment with anti-AChR antibody mAb 35, agrin was still able to effect patching of laminin and HSPG. Taken together, these findings suggest that agrin-induced accumulations of AChR and laminin/HSPG are not mechanistically linked.
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Huynh, Khon C., Huong T. T. Nguyen, Volker R. Stoldt, Marianna Gyenes, and Rudiger E. Scharf. "Shear-Induced Fibrillar-like Supramolecules of Plasma Fibronectin: A New Form of Fibronectin with Enhanced Activity in Platelet Adhesion and Aggregation." Blood 124, no. 21 (December 6, 2014): 4174. http://dx.doi.org/10.1182/blood.v124.21.4174.4174.
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Abstract Introduction: Plasma fibronectin (FN) is synthesized by hepatocytes and secreted into the circulation in a soluble, compact and non-fibrillar form. Plasma FN is assembled by cells or adherent platelets into functional fibrils. Reports have indicated that the process to incorporate FN into multimer fibrils can also occur in cell-free models in vitro by incubation with denaturants, reducing agents, or anastellin (FN peptidic fragment). Here, we report on (1) the formation of insoluble fibrillar-like supramolecules of plasma FN (FN fibrils) by exposing the molecules to increasing shear rates and (2) the functional characterization in platelet adhesion and aggregation. Methods: To induce the formation of FN fibrils, 1 ml of plasma FN solution (100 μg/ml) was added to plates pre-coated with FN (100 μg/ml). Subsequently, the FN solutions were exposed to shear (50 to 5000 s-1 within 5 min and subsequently 5000 to 50 s-1 within 5 min) generated by a cone-plate rheometer (Haaka Rheostress 1; Thermo Scientific). Viscosities of FN solutions were recorded. To quantify the formation of FN fibrils, FN solutions after exposure to shear were collected and incubated with 2% deoxycholate (DOC). The DOC-insoluble pellets containing FN fibrils were isolated by centrifugation at 19,000 g for 20 min at 4°C and resuspended in 1% SDS buffer for Western blot analyses. For adhesion experiments, washed platelets (107/ml) in HEPES Tyrode’s buffer were labeled with 10 μM 5-chloromethylfluorescein diacetate and placed on 96-well plates pre-coated with FN or FN fibrils (25 µg/ml) for 30 min at 37°C. In parallel experiments, platelets resuspended in FN-depleted plasma (107/ml) were placed onto immobilized collagen, fibrinogen, FN (10 µg/ml) in the presence of FN (300 µg/ml) or FN fibrils (10 µg/ml). For aggregation experiments, FN (5, 10, 300 µg/ml) or FN fibrils (5, 10 µg/ml) was added to platelet-rich plasma (PRP) or platelets resuspended in FN-depleted plasma (2.5 × 108/ml). Aggregation was induced by 400 nM PMA (Phorbol 12-myristate 13-acetate), or 10 µg/ml collagen. Results: The initial viscosities (mPa's) of FN solutions were 7.62 ± 0.98. Upon exposure to dynamic shear for 10 min, viscosities increased to 10.98 ± 1.81 (p = 0.02, n = 4), suggesting conformational changes of FN. Western blot analyses of DOC-insoluble fractions revealed bands of FN in the range of 220 – 250 kDa (reducing condition), indicative of the formation of insoluble fibrils in FN solutions after exposure to shear. Platelet adhesion and aggregation experiments were performed to compare the activity of FN fibrils with normal plasma FN. For adhesion experiments, washed platelets in HEPES Tyrode’s buffer were placed onto immobilized FN or FN fibrils (25 µg/ml). The adhesion rates (mean fluorescence signal ± SD) of washed platelets were higher onto surfaces coated with FN fibrils (0.5 ± 0.06) than onto surfaces coated with plasma FN (0.4 ± 0.01) (p = 0.04, n = 3). In parallel adhesion experiments using platelets resuspended in FN-depleted plasma, addition of plasma FN (300 µg/ml) increased platelet adhesion rates onto immobilized collagen (from 0.14 ± 0.005 to 0.2 ± 0.01, p = 0.0007), fibrinogen (from 0.16 ± 0.03 to 0.22 ± 0.01, p = 0.03), and FN (from 0.14 ± 0.01 to 0.18 ± 0.02, p = 0.04) (n = 3). Addition of FN fibrils at low concentration of 10 µg/ml had a similar supportive effect. FN showed an inhibitory effect in platelet aggregation. Activation by 400 nM PMA induced aggregation of PRP by 81% (amplitude). In the presence of plasma FN at 5, 10, 300 µg/ml, platelet aggregation was reduced to 50 %, 41 %, and 29.5 %, respectively. A stronger inhibition on platelet aggregation was seen when FN fibrils were added. PRP aggregated by 35.4 % and 17 % in the presence of 5 and 10 µg/ml FN fibrils, respectively. The same phenomenon was observed in aggregation assays using platelets resuspended in FN-depleted plasma and collagen (10 µg/ml) as activating agonist. Conclusion: Our study shows that dynamic shear rates induce the formation of insoluble fibrillar-like form of plasma FN in cell-free model in vitro. Fibril formation of FN can be monitored by measuring viscosities of FN solutions during exposure to shear and quantified by Western blot. Shear-induced formed FN fibrils have an explicitly stronger activity in supporting platelet adhesion and inhibiting platelet aggregation than normal plasma FN. This finding emphasizes the importance of FN assembly on its activity in platelet functions. Disclosures No relevant conflicts of interest to declare.
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Diehl-Seifert, B., B. Kurelec, R. K. Zahn, A. Dorn, B. Jericevic, G. Uhlenbruck, and W. E. Muller. "Attachment of sponge cells to collagen substrata: effect of a collagen assembly factor." Journal of Cell Science 79, no. 1 (November 1, 1985): 271–85. http://dx.doi.org/10.1242/jcs.79.1.271.
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Collagen, isolated from the sponge Geodia cydonium in the absence of denaturing agents, had the typical amino acid composition and was associated with the carbohydrates galactose and glucose. The resulting individual fibrils with a diameter of 23 nm, displayed a 19.5 nm periodicity with one intraperiod band. A collagen assembly factor (CAF) was identified in and partially purified from the extracellular space. The CAF reacted with antibodies against intact Geodia cells but not with antibodies against Geodia lectin and Geodia aggregation factor. In the presence of the CAF, the collagen fibrils reconstituted collagen bundles in an ordered sequence of events, which were followed by electron-microscopical and biochemical methods. Bundle formation was not dependent on the presence of the homologous lectin, glycoconjugates or aggregation factor. Homologous cells (Geodia archaeocytes) were determined to attach only to those Geodia collagen substrates that contained CAF. The attachment of these cells did not require fibronectin or Geodia lectin. Homologous glycoconjugates or NaOH-treated collagen inhibited cell attachment. Collagen from the sponge Chondrosia reniformis, even in the presence of Geodia CAF, was no appropriate substrate for Geodia cell attachment. Whether collagen is a component of cell-matrix interactions in sponge systems also in vivo is discussed.
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Parmentier, S., B. Catimel, L. McGregor, LL Leung, and JL McGregor. "Role of glycoprotein IIa (beta 1 subunit of very late activation antigens) in platelet functions." Blood 78, no. 8 (October 15, 1991): 2021–26. http://dx.doi.org/10.1182/blood.v78.8.2021.2021.
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Abstract Very late activation antigens (VLAs) are glycoproteins (GPs) that play a major role in platelet adhesion to extracellular matrix. These GPs, members of the integrin family, are heterodimer complexes with different alpha subunits noncovalently associated with a common beta 1 subunit known as GPIIa. GPIa-IIa (also known as VLA2), GPIc-IIa (VLA5), and GPIc*-IIa (VLA6) are involved, respectively, in platelet adhesion to collagen, fibronectin, and laminin. At this stage, very little is known about the role of GPIIa in platelet adhesive functions. In this study, we have generated a monoclonal antibody (MoAb) (LYP22) directed against GPIIa. Immunoaffinity chromatography using LYP22 combined with two-dimensional nonreduced-reduced sodium dodecyl sulfate- polyacrylamide gel electrophoresis shows that the antibody brings down all VLA subunits. Western blots indicate that the binding site of LYP22 on GPIIa is disulfide bridge-dependent. The number of LYP22 binding sites is not increased on stimulation with thrombin and is in the range of what is observed with another anti-GPIIa MoAb (A-1A5). LYP22 is the first anti-GPIIa MoAb to inhibit aggregation and secretion of washed platelets stimulated with collagen, thrombin, or arachidonic acid. Moreover, the lag-phase usually observed on collagen stimulation is significantly prolonged (by 60 seconds) in the presence of LYP22. This lag-phase, mediated by LYP22, is also observed in the presence of plasma proteins and is coupled with a reduced effect on collagen- induced platelet aggregation. In addition, LYP22 affects the adhesion of resting platelets to type III collagen, but not to fibronectin, laminin, or type I collagen. These results strongly indicate that the site on GPIIa, bearing the LYP22 epitope, is an active participant in signal transduction controlling platelet functions.
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Parmentier, S., B. Catimel, L. McGregor, LL Leung, and JL McGregor. "Role of glycoprotein IIa (beta 1 subunit of very late activation antigens) in platelet functions." Blood 78, no. 8 (October 15, 1991): 2021–26. http://dx.doi.org/10.1182/blood.v78.8.2021.bloodjournal7882021.
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Very late activation antigens (VLAs) are glycoproteins (GPs) that play a major role in platelet adhesion to extracellular matrix. These GPs, members of the integrin family, are heterodimer complexes with different alpha subunits noncovalently associated with a common beta 1 subunit known as GPIIa. GPIa-IIa (also known as VLA2), GPIc-IIa (VLA5), and GPIc*-IIa (VLA6) are involved, respectively, in platelet adhesion to collagen, fibronectin, and laminin. At this stage, very little is known about the role of GPIIa in platelet adhesive functions. In this study, we have generated a monoclonal antibody (MoAb) (LYP22) directed against GPIIa. Immunoaffinity chromatography using LYP22 combined with two-dimensional nonreduced-reduced sodium dodecyl sulfate- polyacrylamide gel electrophoresis shows that the antibody brings down all VLA subunits. Western blots indicate that the binding site of LYP22 on GPIIa is disulfide bridge-dependent. The number of LYP22 binding sites is not increased on stimulation with thrombin and is in the range of what is observed with another anti-GPIIa MoAb (A-1A5). LYP22 is the first anti-GPIIa MoAb to inhibit aggregation and secretion of washed platelets stimulated with collagen, thrombin, or arachidonic acid. Moreover, the lag-phase usually observed on collagen stimulation is significantly prolonged (by 60 seconds) in the presence of LYP22. This lag-phase, mediated by LYP22, is also observed in the presence of plasma proteins and is coupled with a reduced effect on collagen- induced platelet aggregation. In addition, LYP22 affects the adhesion of resting platelets to type III collagen, but not to fibronectin, laminin, or type I collagen. These results strongly indicate that the site on GPIIa, bearing the LYP22 epitope, is an active participant in signal transduction controlling platelet functions.
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Baldwin, Samuel P., Christine E. Krewson, and W. Mark Saltzman. "PC12 CELL AGGREGATION AND NEURITE GROWTH IN GELS OF COLLAGEN, LAMININ AND FIBRONECTIN." International Journal of Developmental Neuroscience 14, no. 3 (June 1996): 351–64. http://dx.doi.org/10.1016/0736-5748(96)00018-4.
Full textDissertations / Theses on the topic "Fibronectin Aggregation in Collagen"
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Reyes, Catherine Diane. "Collagen- and Fibronectin-Mimetic Integrin-Specific Surfaces That Promote Osseointegration." Diss., Georgia Institute of Technology, 2006. http://hdl.handle.net/1853/11599.
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Cell adhesion to the extracellular matrix through cell-surface integrin receptors is essential to development, wound healing, and tissue remodeling and therefore represents a central theme in the design of bioactive surfaces that successfully interface with the body. This is especially significant in the areas of integrative implant coatings since adhesion triggers signals that regulate cell cycle progression and differentiation in multiple cellular systems. The interactions of osteoblasts with their surrounding extracellular matrix are essential for skeletal development and homeostasis and the maintenance of the mature osteoblastic phenotype. Our objective was to engineer integrin-specific bioactive surfaces that support osteoblastic differentiation and promote osseointegration by mimicking these interactions. We target two specific integrins essential to osteoblast differentiation the type I collagen receptor alpha2beta1 and the fibronectin receptor alpha5beta1. The central hypothesis of this project was that the controlled presentation of type I collagen and fibronectin binding domains onto well-defined substrates would result in integrin-specific bioadhesive surfaces that support osteoblastic differentiation, matrix mineralization, and osseointegration. We have demonstrated that these biomimetic peptides enhance bone formation and mechanical osseointegration on titanium implants in a rat tibia cortical bone model. We have also shown that the presentation of multiple integrin-binding ligands synergize to enhance intracellular signaling and proliferation. Finally, we demonstrate the advantage of the short biomimetic peptides over the native ECM proteins. This research is significant because it addresses current orthopaedic implant limitations by specifically targeting cellular responses that are critical to osteoblastic differentiation and bone formation. This biomolecular approach provides a versatile and robust strategy for developing bioactive surfaces that enhance bone repair and osseointegration of orthopaedic implants.
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Millard, Christopher John. "Structural and functional characterisation of the collagen binding domain of fibronectin." Thesis, University of Oxford, 2007. http://ora.ox.ac.uk/objects/uuid:af0ec9b5-8789-498e-a1b7-5887ca1ad03f.
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Fibronectin is an extracellular multidomain glycoprotein that directs and regulates a variety of cell processes such as proliferation, development, haemostasis, embryogenesis, and wound healing. As a major component of blood, fibronectin exists as a soluble disulphide linked dimer, but it can also be incorporated into an insoluble cross-linked fibrillar network to form a major component of the extracellular matrix. Fibronectin is composed of an extended chain of module repeats termed Fn1, Fn2, and Fn3 that bind to a wide range of transmembrane receptors and extracellular matrix components, including collagen. The gelatin binding domain of fibronectin was first isolated as a 45kDa proteolytic fragment and has since been found to be composed of six modules: 6Fn1-1Fn2-2Fn2-7Fn1-8Fn1-9Fn1 (in this notation nFX represents the nth type X module in the native protein). This domain has been reported to bind to both collagen and denatured collagen (gelatin), but with 10-100 times higher affinity to the latter; it can be purified to homogeneity on a gelatin affinity column. In the work presented here, fragments of the gelatin binding domain are expressed in P. pastoris, purified to homogeneity, and investigated at the molecular level. Through a dissection approach, surface plasmon resonance (SPR) is used to characterise the recombinantly produced protein, to accumulate more information about the function of the full domain. NMR is used to assess the folding of the protein fragments at atomic resolution. In particular, the secondary structure of 8Fn1-9Fn1 is mapped using inter-strand NOEs, which suggests that the construct takes the fold of a pair of typical Fn1 modules. Gelatin affinity chromatography is used to confirm that both Fn1 and Fn2 modules contribute to gelatin binding, possibly in two clusters (1Fn2-2Fn2 and 8Fn1-9Fn1). The 7Fn1 module may perform a structural role in linking together these two interaction sites, in the same way as suggested for 6Fn1, which is thought to act in a structural manner to enhance the binding of 1Fn2-2Fn2 to gelatin. Three carbohydrate moieties are found on this domain, one on 2Fn2 and two on 8Fn1. Here, by means of expressing different protein length fragments, and by site directed mutagenesis, the role of each sugar chain is investigated independently. The sugar chain on 2Fn2 does not appear to promote binding to collagen, nor does the first sugar chain on 8Fn1 (N-linked to N497), implying another role for these sugars such as protection from proteolysis. However, the presence of at least a single GlcNAc sugar residue on the second sugar chain site on 8Fn1 (N- linked to N511) is essential for full affinity binding to collagen. Direct binding of the 8Fn1-9Fn1 module pair to collagen is assessed with a short collagen peptide and the binding is monitored by NMR. The peptide appears to bind, predominantly to the final strand of 8Fn1, the first β- strand of 9Fn1, and the linker between the two modules, with μM affinity. A model for bound peptide is proposed. The highly conserved amino acid motif Ile-Gly-Asp (IGD) is found on four of the nine N-terminal Fn1 modules of fibronectin. Tetrapeptides containing the IGD were demonstrated to promote the migration of fibroblast cells into a native collagen matrix. Two of these “bioactive” IGD motifs are found within the gelatin binding domain, one on 7Fn1 and one on 9Fn1. In this study, the motif in the 8Fn1-9Fn1 module pair is shown to be located in a tightly constrained loop within 9Fn1. By site directed mutagenesis, the IGD motifs of 7Fn1 and 9Fn1 are subjected to single amino acid substitutions, and their ability to stimulate cell migration assessed in our assay. By NMR, the fold of the IGD mutant proteins is found to be unaffected by the mutation with respect to the wild type, with the exception of small perturbations around the substitution site. While the wild type module is able to stimulate fibroblast migration, the mutant proteins show reduced or negligible bioactivity. The larger fragments show far more potency in stimulating fibroblast migration, with 8Fn1-9Fn1 (one IGD motif) 104 times more potent than the IGD peptide, and the full gelatin binding domain (two IGD motifs) 106 times more potent than the 8Fn1-9Fn1. Potential mechanisms for this enormous enhancement of the IGD potency in different contexts are discussed.
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Asadishekari, Maryam. "Design and Engineering of 3D Collagen-Fibronectin Scaffolds for Wound Healing and Cancer Research." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/38378.
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Despite our understanding of the importance of the 3D environment on the behaviour of virtually every cell, most studies are still performed within 2D engineered cell culture devices. In this project, the main goal was to design and engineer tunable three-dimensional (3D) extracellular matrix (ECM)-mimicking scaffolds made of collagen and fibronectin (namely the two major building blocks of the ECM) that recapitulate the ECM structural and mechanical properties essential for wound healing and cancer research. Two different methods were implemented to fabricate 3D scaffolds.
First, 3D collagen scaffolds with a ‘porous’ structure (fabricated by a previous student via an ice-templating technique) were used. It was shown that, by increasing collagen concentration to 1.25 wt.%, homogenous scaffolds with interconnected pores (needed for cell invasion through the entire scaffold) were obtained. Fibronectin (Fn) was then incorporated using thermal and mechanical gradients to modify protein content and tune scaffolds microarchitecture. The effect of Fn coating of the collagen underlying structure on cell behaviour such as cell adhesion, invasion and matrix deposition was studied. Results showed that overall more cells adhered to Fn-coated scaffolds with respect to pure collagen scaffolds. Furthermore, our findings indicated that cells were also able to sense the conformation of the Fn coating (as assessed by Fluorescence Resonance Energy Transfer, FRET) since they deposited a more compact ECM on compact Fn coating while a more unfolded and stretched ECM was deposited on unfolded Fn coating.
Second, 3D more complex physiologically relevant scaffolds with a ‘fibrillar’ structure were fabricated via a cold/warm casting technique. Pure collagen scaffolds were first generated: in cold-cast scaffolds, clear thin and long collagen fibers were observed while warm-cast scaffolds were denser and comprised shorter collagen fibers. The effect of both collagen concentration and casting temperature on scaffolds’ microstructure was studied. Our results indicate a preponderant effect of temperature. We further engineered dual-protein fibronectin-collagen fibrillar scaffolds by incorporating Fn fibers using thermal gradient. Clear Fn fibers were observed in some conditions. FRET assessment of Fn fibers also showed significant difference of Fn conformation. In this more advanced casting technique, cells were initially embedded into the scaffolds, which provided a more homogeneous cell distribution and a better tissue-mimicking setting. In each case, the effect of resulting ECM properties was tested via cell viability assays. Our data indicate that cells were viable after 72 hours, they could proliferate inside the scaffolds and were able to spread in some conditions.
Collectively, our 3D ECM-mimicking scaffolds represent a new tunable platform for biological and biomaterial research with many potential applications in tissue engineering and regenerative medicine. Investigating cell behaviour in 3D ECM-mimicking environment will provide valuable insights to understand cancer progression and approaches to limit the progression and ultimately prevent metastasis.
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Stanton, Heather. "The role of matrix metalloproteinases in cell-matrix interactions." Thesis, Open University, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.284376.
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Stupack, Dwayne G. P. "Utilization and regulation of integrins by lymphoid cells to adhere to fibronectin and collagen." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1996. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq23671.pdf.
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Väisänen, M. R. (Marja-Riitta). "Type XIII collagen:characterization of ectodomain shedding and its biological implications in mammalian cells, characterization of type XIII collagen expression in human cancers." Doctoral thesis, University of Oulu, 2005. http://urn.fi/urn:isbn:9514279085.
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Abstract
Type XIII collagen is an integral membrane protein in type II orientation. In cells and tissues type XIII collagen has been located in various adhesive structures, like focal adhesions. Due to this, its biological role has been implicated in cell adhesion. This collagen also exists as a soluble protein due to the release of the ectodomain from the plasma membrane.
In this thesis, ectodomain shedding, i.e. enzymatic release of the extracellular domain, was studied in detail, focusing on the phenomenon as it occurs in mammalian cells. It was found that the ectodomain is released by members of the mammalian proprotein convertase family, e.g. furin. Shedding was shown to take place at the cell surface, but based on additional observations, this cleavage may also take place intracellularly in the Golgi apparatus. Various intracellular mechanisms, depending on cell type, were found to be involved in the regulation of ectodomain shedding. Apparently, due to the liberation of the ectodomain, the level of type XIII collagen on the plasma membrane is maintained at a relatively even amount.
The released ectodomain was shown to retain biological activity. It showed distinct matrix-specificity so that on vitronectin its influence on cell functions was anti-adhesive, anti-migratory, anti-proliferative and non-supportive of cell spreading. It was also demonstrated to affect the fibronectin matrix assembly in a manner that resulted in reduced amounts of the fibrillar fibronectin matrix.
A large collection of human epithelial and mesenchymal cancer samples were screened for type XIII collagen mRNA expression and compared to the expression levels of pre-malignant and normal samples. It was discovered that malignant transformation upregulates the expression of type XIII collagen in mesenchymal cancers and particularly in the stroma of epithelial cancers, more so than in cancer epithelia. TGF-β1 was demonstrated as one factor contributing to the stimulation of expression. Based on cell culture experiments in this study, it was also deduced that the upregulated expression of type XIII collagen and the concomitant shedding of the ectodomain can remodel the tumour stroma, making it inauspicious for adhesion-dependent cell functions, particularly in vitronectin-rich milieu.
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Li, Chuen-wai, and 李鑽偉. "Dynamic compression and exogenous fibronectin regulates cell-matrix adhesions and intracellular signaling proteins of human mesenchymal stem cells in 3D collagen environment." Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2013. http://hdl.handle.net/10722/197553.
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The fundamental principle of tissue engineering is to use appropriate cell source, combined with scaffolds and bioactive factors to develop tissue constructs which restore, maintain or improve tissue function. There is increasing data emphasizing the importance of mechanical signals and extracellular matrix (ECM) proteins presented by the scaffold in determining stem cell fate/functions which are critical to tissue construct maturation and success of stem cell-based therapies. Cell-matrix adhesions are one of the major mechanosensing machineries cells use to convert information provided by ECM ligands and mechanical signals presented by scaffolds into intracellular biochemical signaling cascades which lead to particular functional responses. Therefore, understanding how ECM ligands and mechanical signals regulate cell-matrix adhesion formation and activation of associated intracellular signaling proteins is fundamental to rational design of biomaterial and loading protocol for optimal cell functional responses in tissue constructs.
In this study, we attempted to understand the regulatory effects of external mechanical signal and exogenous ECM protein on cell-matrix adhesion formation and associated intracellular signaling proteins of human mesenhymal stem cells, and in particular, to test the hypothesis that mechanical stimulation or exogenous ECM protein can lead to adhesion maturation into 3D-matrix adhesions in 3D collagen environment. We used microencapsulation technique to embed cells in 3D collagen environment, forming disc-shaped hMSC-collagen constructs. By immunofluorescent staining and confocal microscopy, we visualized changes in size, morphologies and molecular composition of the adhesions.
First of all, 2D adhesions of hMSCs were characterized. We showed that hMSCs form well-organized αv integrin-based focal adhesions and fibrillar adhesions in 2D culture. To investigate the regulatory effects of mechanical signals on adhesion signaling and maturation, we used micromanipulator-based loading device to impose dynamic compression to hMSC-collagen constructs. We found that dynamic compression lead to enlargement of integrin αv adhesions which recruit focal adhesion kinase (FAK), vinculin and extracellular signal-regulated kinase (ERK). In addition, FAK was activated at enlarged integrin αv adhesions and translocated to peri-nuclear region after compression, suggesting that loading induces activation of FAK signaling pathways through increased integrin αv clustering. Moreover, we demonstrated that dynamic compression can induce 3D-matrix adhesion formation, indicating the role of external force in integrin α5-based adhesion maturation in 3D collagen environment.
We explored the effect of exogenous ECM proteins on adhesion maturation of hMSCs by adding fibronectin into cell-collagen mixture during fabrication of collagen constructs. Our results demonstrated that the exogenous fibronectin can induce α5 integrin-based adhesion maturation into 3D-matrix adhesions in our collagen constructs in a dose-dependent manner.
This study demonstrated that the effect of external mechanical signals and exogenous ECM ligands on adhesion signaling and maturation of hMSCs in 3D collagen environment. Our findings contribute towards mechanobiology of hMSCs in 3D context. In particular, our results showed that exogenous proteins or external loading can lead to 3D-matrix adhesion formation, which may serve as a potential way to enhance biological functions of hMSCs in collagen constructs, facilitating stem cell-based therapies.published_or_final_version
Mechanical Engineering
Doctoral
Doctor of Philosophy
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Iskender, Banu. "Investigating the effects of extracellular matrix molecules on human embryonic stem cells." Thesis, University of Manchester, 2012. https://www.research.manchester.ac.uk/portal/en/theses/investigating-the-effects-of-extracellular-matrix-molecules-on-human-embryonic-stem-cells(e6b6df88-645a-4516-92da-63b3efee3cdb).html.
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Human embryonic stem cells are pluripotent cells that have indefinite replicative potential and ability to differentiate into derivatives of three germ layers. HESCs are conventionally derived and grown on mitotically inactivated mouse embryonic fibroblasts and there are some alternative feeder types of human origin that have been used to replenish hESCs while trying to prevent cross-species contamination. The trophic factors that are secreted by the feeders are found to be important for long-term pluripotency but there are also supportive culture systems for hESCs lacking feeder cells which might suggest that not only the interactions with the feeders affect the behaviour of hESCs but also the components of the niche may take part in the decision of self-renewal or differentiation. Extracellular matrix components are known to exert their stimulatory or inhibitory effects by localising cells into a specific microenvironment in natural niches but have been relatively little investigated for hESCs. The aim of this study was to investigate ECM components which might have a role in the maintenance of hESCs. I have first investigated human placental stromal fibroblasts and immortalised human placental stromal fibroblasts for the support hESC pluripotency as an anlternative feeder type to conventional mouse embryonic fibroblasts. Secondly, the matrices derived from hPSFs and ihPSFs were assessed for their ability to support hESC pluripotency. Tandem mass spectrometry was used to identify ECM components released by human feeders in order to characterise the range of extracellular matrix proteins that support the growth of self-renewing hESCs. The majority of the molecules was shared between the cell types irrespective of hPSF cell derived matrix was not being supportive for hESC pluripotency, with some ECM components being unique ihPSFs. Collagen VI, tenascin C and versican were tested for hESC attachment and as substrates for feeder-free culture system in order to develop an optimised feeder-free system. Furthermore, integrin receptor profile of different hESC lines was also determined in order to identify the mechanisms of substrate attachment. Integrin attachment was shown to be vital for hESC engagement to fibronectin and vitronectin in feeder-free systems. The components of the integrin signalling machinery were identified in hESCs and the significance of integrin-mediated signalling in hESC self-renewal was demonstrated by blocking integrin β1 on fibronectin and integrin aVβ5 on vitronectin. Moreover, intracellular signalling mediator c-Src was shown to involve in ECMregulated signalling by affecting the phosphorylation of Focal Adhesion Kinase. Inhibition of Src led to a decrease in the expression of pluripotency-associated markers. Finally, the effects of growth factor supplementation on the maintenance of pluripotency in defined feeder-free conditions were studied by withdrawal of growth factors and blocking FGF Receptors. FGF-2 was shown to be essential for long-term self-renewal while the effects on pluripotency deteriorated in the absence of both FGF-2 and Activin A. Taken together this project highlighted the importance of substrate attachment and growth factors on the regulation of hESC self-renewal.
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Linnola, R. (Reijo). "The sandwich theory:a bioactivity based explanation for posterior capsule opacification after cataract surgery with intraocular lens implantation." Doctoral thesis, University of Oulu, 2001. http://urn.fi/urn:isbn:9514259793.
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Abstract
This study was undertaken to identify mechanisms of adhesion of intraocular lenses (IOLs) to the
capsular bag after cataract surgery and IOL implantation. It was also done to challenge the sandwich
theory presented for posterior capsular opacification (PCO): If the IOL is made of a bioactive material
it would allow a single lens epithelial cell layer to bond both to the IOL and the posterior capsule at
the same time. This would produce a sandwich pattern including the IOL, the cell monolayer and the
posterior capsule. The sealed sandwich structure would prevent further epithelial ingrowth. The degree
of bioactivity of the IOL could explain the basic difference in the incidence of PCO and capsulotomy
rates with different IOL materials.
The sandwich theory was put forward on the basis of a search for a keratoprosthesis material,
which would allow maximal adhesion of the prosthesis to corneal tissue. Titanium and glass-ceramic
coated titanium were found to develop better adhesion than poly (methyl methacrylate) (PMMA). The
adhesion of PMMA to the corneal stromal tissue was loose, and down growth of corneal epithelial cells
was seen around the prosthesis.
The differences between various IOL materials were first tested with rabbit corneal tissue
cultures. There was better adhesion of corneal tissue to soft, hydrophobic acrylate than to PMMA,
heparin surface modified (HSM)-PMMA, silicone or hydrogel IOLs.
To assess differences in protein adhesion to IOL surfaces, different IOLs were incubated for 24
hours with radioactive iodine labeled fibronectin. Soft hydrophobic acrylate (AcrySof®) showed the
highest binding of fibronectin, and the differences relative to all the other materials were significant
(p < 0.01-0.001), except to PMMA (p = 0.31).
The sandwich theory and the results with rabbit corneal tissue cultures and the protein adhesion
study in vitro were evaluated against the results found in pseudophakic autopsy eyes. Altogether, 70
autopsy eyes were analyzed. From 38 autopsy eyes containing PMMA, silicone, soft hydrophobic acrylate or
hydrogel IOLs histological sections were prepared from the capsular bag and immunohistochemical analyses
were performed for fibronectin, vitronectin, laminin and collagen type IV. A total of 152 specimens were
analyzed. From 32 autopsy eyes containing IOLs made of PMMA, silicone, acrylate or hydrogel, IOLs were
explanted from the capsular bag and immunohistochemical analysis was done on both sides of the IOLs for
fibronectin, vitronectin, laminin or collagen type IV. Soft hydrophobic acrylate IOLs had significantly
more adhesion of fibronectin to their surfaces than PMMA or silicone IOLs. Also, more vitronectin was
attached to acrylate IOLs than to the other IOL materials. Silicone IOLs had more collagen type IV
adhesion in comparison to the other IOL materials studied. In histologic sections a sandwich-like
structure (anterior or posterior capsule-fibronectin-one cell layer-fibronectin-IOL surface) was seen
significantly more often in eyes with acrylate IOLs than in PMMA, silicone or hydrogel IOL eyes.
These studies support the sandwich theory for posterior capsule opacification after
cataract surgery with IOLs. The results suggest that fibronectin may be the major extracellular protein
responsible for the attachment of acrylate IOLs to the capsular bag. This may represent a true bioactive
bond between the IOL and the lens epithelial cells, and between the IOL and the capsular bag. This may
explain the reason for clinical observations of less posterior capsular opacification and lower
capsulotomy rates with the soft hydrophobic acrylate material of AcrySof® IOLs compared to the
other IOL materials studied.
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Huynh, Khon Verfasser], Rüdiger E. [Akademischer Betreuer] Scharf, and Dieter [Akademischer Betreuer] [Willbold. "Role of fibronectin in platelet adhesion and aggregation: impact of biomechanics and β3 integrin on fibrillogenesis / Khon Huynh. Gutachter: Rüdiger E. Scharf ; Dieter Willbold." Düsseldorf : Universitäts- und Landesbibliothek der Heinrich-Heine-Universität Düsseldorf, 2012. http://d-nb.info/1029350108/34.
Full textBooks on the topic "Fibronectin Aggregation in Collagen"
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Kirkpatrick, Peter. Immunohistochemical demonstration of fibronectin, laminin and collagen type III in human tissues. Uxbridge: Brunel University, 1988.
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Parkinson, John. Mathematical modelling of collagen fibril formation using diffusion limited aggregation. Manchester: University of Manchester, 1995.
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Badimon, Lina, Felix C. Tanner, Giovanni G. Camici, and Gemma Vilahur. Pathophysiology of thrombosis. Oxford University Press, 2017. http://dx.doi.org/10.1093/med/9780198755777.003.0018.
Full textAbstract:
Ischaemic heart disease and stroke are major causes of death and morbidity worldwide. Coronary and cerebrovascular events are mainly a consequence of a sudden thrombotic occlusion of the vessel lumen. Arterial thrombosis usually develops on top of a disrupted atherosclerotic plaque because of the exposure of thrombogenic material, such as collagen fibrils and tissue factor (TF), to the flowing blood. TF, either expressed by subendothelial cells, macrophage- and/or vascular smooth muscle-derived foam-cells in atherosclerotic plaques, is a key element in the initiation of thrombosis due to its ability to induce thrombin formation (a potent platelet agonist) and subsequent fibrin deposition at sites of vascular injury. Adhered platelets at the site of injury also play a crucial role in the pathophysiology of atherothrombosis. Platelet surface receptors (mainly glycoproteins) interact with vascular structures and/or Von Willebrand factor triggering platelet activation signalling events, including an increase in intracellular free Ca2+, exposure of a pro-coagulant surface, and secretion of platelet granule content. On top of this, interaction between soluble agonists and platelet G-coupled protein receptors further amplifies the platelet activation response favouring integrin alpha(IIb)beta(3) activation, an essential step for platelet aggregation. Blood-borne TF and microparticles have also been shown to contribute to thrombus formation and propagation. As thrombus evolves different circulating cells (red-blood cells and leukocytes, along with occasional undifferentiated cells) get recruited in a timely dependent manner to the growing thrombus and further entrapped by the formation of a fibrin mesh.
Book chapters on the topic "Fibronectin Aggregation in Collagen"
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Wadström, Torkel, Janos Erdei, Marianne Paulsson, and Åsa Ljungh. "Fibronectin, Collagen and Vitronectin Binding of Coagulase-Negative Staphylococci." In Pathogenesis of Wound and Biomaterial-Associated Infections, 339–47. London: Springer London, 1990. http://dx.doi.org/10.1007/978-1-4471-3454-1_41.
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Franchi, Marco, Valentina Masola, Konstantinos-Athanasios Karamanos, Leonardo Franchi, Konstantina Kyriakopoulou, Maurizio Onisto, and Concettina Cappadone. "Basement Membrane, Collagen, and Fibronectin: Physical Interactions with Cancer Cells." In The Extracellular Matrix and the Tumor Microenvironment, 247–77. Cham: Springer International Publishing, 2022. http://dx.doi.org/10.1007/978-3-030-99708-3_10.
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Khon, Huynh, Huong T. T. Nguyen, Phong Le, Thao Nguyen, Thi-Hiep Nguyen, Toi Vo Van, and Volker R. Stoldt. "Shear-Induced Fibrillar-Like Supramolecule of Plasma Fibronectin: A New Form of Fibronectin with Enhanced Activity in Platelet Adhesion and Aggregation." In 6th International Conference on the Development of Biomedical Engineering in Vietnam (BME6), 805–8. Singapore: Springer Singapore, 2017. http://dx.doi.org/10.1007/978-981-10-4361-1_137.
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Peters, Theodorus H. F., Peter L. de Jong, Lennart Klompe, Rolf M. F. Berger, Pramod R. Saxena, Hari S. Sharma, and Ad J. J. C. Bogers. "Right ventricular collagen and fibronectin levels in patients with pulmonary atresia and ventricular septal defect." In Biochemistry of Hypertrophy and Heart Failure, 27–32. Boston, MA: Springer US, 2003. http://dx.doi.org/10.1007/978-1-4419-9238-3_4.
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Felgueiras, H., V. Migonney, S. Sommerfeld, N. S. Murthy, and J. Kohn. "Competitive Adsorption of Albumin, Fibronectin and Collagen Type I on Different Biomaterial Surfaces: A QCM-D Study." In IFMBE Proceedings, 1597–600. Cham: Springer International Publishing, 2014. http://dx.doi.org/10.1007/978-3-319-00846-2_394.
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Clément, B., M. Rissel, S. Peyrol, J. A. Grimaud, and A. Guillouzo. "LIGHT AND ELECTRON MICROSCOPIC IMMUNOLOCALIZATION OF FIBRONECTIN AND TIPE I COLLAGEN IN ADULT RAT HEPATOCYTES DURING PRIMARY CULTURE." In Proceedings of the Third Symposium, Lyon, France, June 26–28, 1985, edited by Jacques Bienvenu, J. A. Grimaud, and Philippe Laurent, 411–16. Berlin, Boston: De Gruyter, 1986. http://dx.doi.org/10.1515/9783110860757-054.
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Teng, Che-Ming, Feng-Nien Ko, Chien-Huan Lin, and Chung-Nan Lin. "Collagen-Induced Platelet Aggregation was Mimicked by Snake Venom Protein and Inhibited by Plant Component." In Current Aspects of Blood Coagulation, Fibrinolysis, and Platelets, 147–51. Tokyo: Springer Japan, 1993. http://dx.doi.org/10.1007/978-4-431-68323-0_25.
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Shibasaki, Y., S. Hirohara, K. Terada, T. Ando, and M. Tanihara. "Collagen-Like Polypeptide Poly(Pro-Hyp-Gly) Conjugated with Fibronectin- Derived Peptides Enhances the Cell Adhesion, Migration and Stratification." In IFMBE Proceedings, 1086–89. Berlin, Heidelberg: Springer Berlin Heidelberg, 2011. http://dx.doi.org/10.1007/978-3-642-23508-5_282.
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Drakes, Maureen L., Lina Lu, Hilary J. Mckenna, and Angus W. Thomson. "The Influence of Collagen, Fibronectin, and Laminin on the Maturation of Dendritic Cell Progenitors Propagated from Normal or Flt3-Ligand-Treated Mouse Liver." In Advances in Experimental Medicine and Biology, 115–20. Boston, MA: Springer US, 1997. http://dx.doi.org/10.1007/978-1-4757-9966-8_19.
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Moortgat, Peter, Mieke Anthonissen, Ulrike Van Daele, Jill Meirte, Tine Vanhullebusch, and Koen Maertens. "Shock Wave Therapy for Wound Healing and Scar Treatment." In Textbook on Scar Management, 485–90. Cham: Springer International Publishing, 2020. http://dx.doi.org/10.1007/978-3-030-44766-3_55.
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AbstractShock Wave Therapy (SWT) meets all the requirements for the ideal non-invasive scar treatment. It is safe, well tolerated by patients, cost-effective, easy to apply, has low complication rates, and can be used in an outpatient setting. The overall effect of SWT is an improvement of tissue homeostasis, accompanied by an improvement of the tissue self-healing abilities, and it seems to focus on inducing tissue regeneration and matrix remodeling in vivo by means of mechanotransduction.SWT has a beneficial effect on wound healing and is characterized by an upregulation of the angio-active factors as nitric oxide (NO) and vascular endothelial growth factor (VEGF) leading to induced angiogenesis. A downregulation of alpha-SMA expression, myofibroblast phenotype, TGF-β1 expression, fibronectin, and collagen type I are measured after SWT on scars, leading to improvement of several relevant scar parameters like height, pliability, vascularity, and pigmentation, and thus ameliorating function.For a full treatment outline, the energy flux density (EFD), the number of pulses, the pulse frequency, and the number and interval of treatments are the most relevant parameters. The EFD for soft tissue indications is typically in the range of 0.08–0.25 mJ/mm2, while scars and fibrosis are treated with an EFD ranging between 0.15 and 0.33 mJ/mm2. These settings seem to be ideal to induce the optimal cell responses for each indication.All the presented findings are fundamental knowledge for further investigation of SWT to reduce the fibrous component in regenerating and remodeling tissues. However, the full potential of SWT in wound healing and scar treatment needs further unraveling.
Conference papers on the topic "Fibronectin Aggregation in Collagen"
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Kenneally, D. A., P. J. Thurlow, and J. M. Cornellan. "MONOCLONAL ANTIBODY (ANTI-2B6D4) TO PLASMA FIBRONECTIN INHIBITS COLLAGEN AND THROMBIN INDUCED AGGREGATION OF WASHED PLATELETS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643861.
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Fibronectins(Fns) constitute a family of large glycoproteins which are known to bind to a wide range of biological molecules eg.collagen, gelatin, fibrin, heparin and DNA,and to many cells including platelets via discrete structural domains. A murine monoclonal antibody (anti-2B6D4) produced by imnunizing BALB/c mice with plasma fn, was used to study the structure and function of fn and its platelet interaction Anti-2B6D4 reacted specifically with plasma fn was unreactive with FVIII/vWF,BTG,PF4 collagen and fibrinogen nor was it reactive with platelets (unstiraulated), human PBLs or a range of tumour cell lines. Iirmuno-blotting studies( 8% SDS-PAGE) using thermolysin-digested plasma fn with anti-2B6D4 indicated that the 2B6D4 epitope was present on only 2 of the 7 fragments detected by Indian ink. The 2 fragments had an Mr of 145 and 155 K dal tons and have been reported to each contain domains which bind cells, DNA and heparin. These fragments were studied further by examining the effect of anti-2B6D4 (Fabs) on the binding of 125I-fn to thrombin-stimulated platelets and demonstrated that anti-2B6D4 binding was inhibited by 50% thus implicating the 2B6D4 epitope as a platelet binding site within the two cell binding domains. Competitive binding analysis of 125I-fn to solid-phase macromolecules i.e.collagen, gelatin, fibrin, heparin, DNA and Con A demonstrated that anti-2B6D4 (Fabs) inhibited the binding of fn to DNA by 50%,but not to the other macromolecules. Therefore, either the DNA and platelet binding sites are shared or the inhibition is due to steric hindrance. However, as Fab fragments of anti-2B6D4 were used, it is more likely that the binding sites are shared. Functional studies were performed to investigate the role of 2B6D4 in platelet-platelet interaction. Anti-2B6D4 totally blocked the aggregation of washed platelets stimulated by low dose collagen (1.6ug/ml) and thrombin (0.05U/ml), partially inhibited arachidonic acid (250ugs/ml) induced platelet aggregation and had no effect on aggregation induced by A23187 (30uM). One other report had demonstrated that fn is a requirement for A23187 and low dose thrombin induced platelet aggregation. We conclude that fn plays an essential role in platelet aggregation induced by low dose collagen and thrombin.
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FAUVEL-LAFEVE, F., and Y. J. LEGRAND. "IMMUNOCHEMICAL IDENTIFICATION OF A THROMBOSPONDIN -LIKE ANTIGEN IN ARTERIAL THROMBOGENIC MICROFIBRILS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643823.
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The biochemical structure of arterial microfibrils (MFS) is unknown. Presently, the most probable hypothesis is that elastin associated MFS contain several antigenic determinants with MW varying between 31 and 200 KD.From our previous studies we know that MFS extracted by 6 M GuCl contain a major glycoprotein with a 128 KD MV (GP128). GP 128 is essential for the reactivity of MFS towards blood platelets but due tc the high insolubility of the extracted material it was not possible to isolate and study this GP 128. We have used immunoblotting to determine if MFS contain determinants recognized by antibodies against connective tissue glycoproteins such as fibronectin, type VI collagen or anti-platelet thrombospondin (TSP). 'The results showed that MFS do not contain fibronectin or type VI collagen but that anti-TSP IgG reacted with GP 128. Furthermore, the Fab fragments from anti-TSP IgG inhibited platelet aggregation induced by MFS but not by collagen or ADP . In a second step,to raise antibodies against GP 128, we prepared blots from entire MFS, the nitrocellulose band corresponding to GP 128 was cut, dissolved in DMSO, and wrs injected to rabbits. Such obtained antibodies recognized only GP 128 in arterial MFS and also TSP in a platelet lysate confirming that GP 128 and TSP have a common antigenic structure. IgG from anti-GP 128 inhibit platelet aggregation induced by MFS but not by collagen or ADP. Previously reported observations showed that tissue TSP and endothelial cells derived GP 128 have a similar affinity for chromatography supports and have the same effect on platelet-MFS interactions. All these results led us to propose that TSP, GP 128, and MFS recognize a common determinant on platelet membrane. This assumption would be strenghened if GP 128 indeed is derived from tissue TSP.
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Nievelstein, P. F. E. M., M. Ottenhof-Rovers, M. D. Pierschbacher, and J. J. Sixma. "THE ARG-GLY-ASP(SER) SEQUENCE OF FIBRONECTIN, AND THE GLYCOPROTEIN IIB-IIIA COMPLEX ARE NOT INVOLVED IN FIBRONECTIN DEPENDENT PLATELET ADHESION IN FLOW." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643590.
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Activated blood platelets interact with fibronectin through it to the glycoprotein IIb-IIIa(GPIIb-IIIa)-complex. The cell attachment site of fibronectin with its crucial arg-gly-asp-(-ser) (RGD(S))sequence is involved in this binding. We have studied the importance of this interaction for the fibronectin dependence of platelet adhesion under flow conditions. An RGDS-containing hexapeptide (GRGDSP) was compared with a non-reactive control peptide (GRGESP). The GRGDSP-peptide inhibited thrombin induced aggregation and adhesion under static conditions at 0.1 mM. This concentration had no effect on platelet adhesion to nonfibrillar collagen type I in flow. GRGDSP at 1 mM had a significant inhibitory effect at 1500 s™1 (8.8 ± 1.4 111In platelets* 105 /cm2, versus 19.8 ± 0.5 for the control). At lower shear rates of 800 and 300 s™1 , where platelet adhesion is also fibronectin dependent, no significant differences were obtained (respectively 11.7 ± 1.1 versus 15.2 ± 2.1, and 11.4 ± 1.0 versus 13.1 ± 0.7).The relation between GPIIb-IIIa and fibronectin dependence was investigated with platelets of a patient with Glanzmann’s thrombasthenia and monoclonal antibodies to GPIIb-IIIa, using endothelial cell matrix (ECM) as a surface. Platelets of normal controls or a patient with Glanzmann’s thrombasthenia showed a inhibition of adhesion in fibronectin free plasma, after the ECM had been preincubated with anti-fibronectin F(ab’)2, of respectively _J5 and 30 percent at 300 s™1 , and 43 and 65 percent at 1300 s™1 . Incubation of platelets with anti GPIIb-IIIa showed inhibition of platelet adhesion at high shear rates. Dependence on fibronectin for platelet adhesion was still observed, even though separate experiments had shown that these anti GPIIb-IIIa antibodies could block binding of radiolabeled fibronectin to thrombin activated platelets. These data suggest the existence of a second binding system from the RGD/GPIIb-IIIa system separate for the interaction of platelets with fibronectin, which may only function when fibronectin is present on a surface.
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Fresslnaud, E., J. E. Sadler, J. P. Girma, H. R. Baumgartner, and D. Meyer. "SYNTHETIC RGD-CONTAINING PEPTIDES OF VON WILLEBRAND FACTOR INHIBIT PLATELET ADHESION TO COLLAGEN." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643591.
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The Arg-Gly-Asp (RGD) sequence is common to fibrinogen (Fg), fibronectin (Fn) and von Willebrand Factor (vWF). RGD-containing peptides compete for binding of these adhesive proteins to platelet membrane GPIIb/IIIa and inhibit thrombin-induced platelet aggregation as does an unrelated dodecapeptide from the γ Fg COOH terminus (γFg 400-411). We compared in flowing blood the effect of γ Fg 400-411 and of 3 synthetic peptides derived from the sequence of human vWF upon platelet adhesion to collagen. The 3 vWF peptides (13 or 18 aminoacids) contained an RGD sequence in the NH2 (peptide 03), central (peptide 07) or COOH (peptide 02) portions. Collagen was coated onto plastic coverslips and exposed in a parallel-plate perfusion chamber to reconstituted human blood at a shear rate of 2,600 s™1 for 3 min at 37°C. Perfusates contained physiological concentrations of 51 Cr-platelets and red cells in either citrated autologous plasma or modified Tyrode buffer containing 4% human albumin ; in the latter case, the collagen-coated coverslips were preincubated with normal plasma or purified human vWF prior to perfusion. Platelet-collagen interactions were estimated by radioactivity counting and quantitative morphometry. RGD peptides 02, 03 and 07 inhibited platelet-collagen interactions in a dose-dependent manner. With peptide 07, deposition of 51 Cr-platelets decreased from 283.8 ± 32.5 × 105/cm2 (mean ± SEM, n = 3) with buffer to 169.6 ± 33.0 in the presence of 50 μM peptide (p < 0.05), 133.7 ± 26.4 with 150 uM (p <0.012) and 101.8 ± 27.1 with 300 uM (p <0.005). The inhibitory effect of γ Fg 400-411 upon platelet deposition was less significant than that of the RGD peptides at 50 and 150 uM concentrations (224.4 ± 39.8, N.S. and 139.5 ± 55.3, p < 0.05, respectively). RGD peptide 07 also inhibited in a dose-dependent way both platelet adhesion to collagen and thrombus growth. Similar results were observed with peptides 02 and 03, indicating that the position of the RGD sequence is not critical. No synergetic effect between RGD and γFg 400-411 peptides was observed. These results with vWF peptides confirm that GPIIb/IIIa is involved not only In platelet aggregation (thrombus growth) but also in vWF-mediated platelet adhesion to collagen.
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Kantak, Ameya, Yuri Lvov, David K. Mills, James G. Spaulding, and Steven A. Jones. "Platelet Function Assessment in a Microfabricated Device." In ASME 2002 International Mechanical Engineering Congress and Exposition. ASMEDC, 2002. http://dx.doi.org/10.1115/imece2002-33135.
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Although platelets are small and simple in shape, they are complicated in their physiologhy. Their alpha granules and dense bodies secrete a large number of agents that are involved in haemostasis, and the glycoproteins on their surfaces form the linkages with proteins like fibrinogen, fibronectin, collagen and von Willebrand factor that are necessary for adhesion and aggregation (Frojmovic, 1998). Diseases such as heart attack (Meade, 1992), stroke (Harker, 1998) and eclampsia (Schindler et al., 1990), can be the result of pathologies in platelets. Although devices have been recently developed to diagnose platelet function (Nicholson et al., 1998), there is still a need for devices that can examine the multiple factors of platelet physiology that affect thrombus formation. Because fluid shear is a key factor in platelet adhesion and aggregation (Colantuoni et al., 1977), it is necessary to test platelet function under conditions of flow. Also, because of the large number of adhesion receptors and secreted agents that make up a platelet’s physiology, a complete diagnosis of platelet function should be performed on a variety of substrates.
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Hawiger, J. "PLATELET RECEPTOR RECOGNITION DOMAINS AND THEIR SYNTHETIC PEPTIDE ANALOGS." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643726.
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Adhesive molecules and their receptorsplay an essential role in hemostasis and thrombosis. Platelet thrombi are formed through the interaction of cell adhesion molecules (CAMs) with intercellular adhesion molecules (IAMs)and substrate adhesion molecules (SAMs). Platelet CAMs encompass membrane glycoproteins lb, lib, Ilia,and possibly la and IV, which constitutemembrane receptors for IAMs(e.g., fibrinogen) and for SAMs encompassingvon Willebrand Factor (vWF), fibronectin, vitronectin, collagen, and thrcmbospondin. Receptorfunction of platelet CAMs can be specific,i.e., only one adhesive protein among IAMs and SAMs is selected forbinding as exemplified by GPIb and vWF. Alternatively,more than one adhesive protein can interact with platelet CAMs comprising the GPIIb/IIIa complex.This common adhesive receptor mechanism switched on by thrombin, ADP, phorbol ester or ionophore A23187 is turned off by a rise in intraplatelet cyclic AMP which provides a negative control.Fibrinogen, the most abundant adhesiveprotein in plasma, interacts with platelet CAMs via receptor recognition domains on gamma and alpha chains. Pinpointing platelet receptor recognition domain to a carboxy-terminal segment of the gamma chain encompassing residues 400-411gave rise to a series of synthetic peptide analogs which do not interfere with themetabolic pathways of platelets but blockbinding of I fibrinogen to its receptors on stimulated platelets, inhibit their aggregation in vitro, and formation of a platelet thrombus in vivo. The alpha chain of human fibrinogen contains the sequenceRGD (residues 95-97 and 572-574). Synthetpeptide analogs of the RGD sequence, which constitute the "cell adhesion site" of fibronectin, also inhibit binding of 125I-fibrinogen to stimulated platelets. However, these synthetic peptides are not "specific" for fibrinogen chains because thealpha chain of human fibrinogen which hasnosequence homology with gamma 400-411 is prevented by a peptide gamma 400-411 from interaction with platelet receptors. Viceversa, the human gamma chain is blocked by tetrapeptide RGDS not expressed in the human gamma chain. Interaction of human vWF with human platelets is blocked by synthetic peptide analogs of gamma 400-411 (not present in vWF)and of RGD sequence (present in vWF).These synthetic peptides inhibite "common" receptor pathwaystimulated with ADP, thrombin, or phorbolester, but they do not interfere with binding of 125I-vWF via a "specific" pathvoy induced with ristocetin and involving GPIb.The design of synthetic peptide analogs which inhibit platelet receptors for adhesive molecules includes the following considerations: ligand specificity (is thepeptide inhibitory toward binding of one or more adhesive molecules?),cell speciicity (is the peptide specific for platelets or does it perturb the adhesive properties of other cells, e.g.,endothelium?);the hydrophilic character; protection against degradation by peptidases; and a sufficiently long half-life to achieve platelet inhibitory potency in vivo without overloading the blood with excessive amounts of peptide.This is accomplished by constructing a peptide-albumin conjugate with ahalf-life extended at least 30 times.Whenpeptides are modeled with predominantly hydrophilic or hydrophobic residues, only the hydrophilic peptide remained active to block the platelet receptor. This agreed with the general observation that sequences on adhesive molecules that are knownto interact with cellular receptors have a hydrophilic rather than a hydrophobic character. Furthermore, changing the charge of synthetic peptides toward the negative reduced the reactivity, whereas introducing additional arginine residues enhanced the reactivity toward platelet receptors. Localization of the functionally important binding domain in the flexible segment of an adhesive protein increases the likelihood that the synthetic peptide will assume the conformation mimicking such a domain in the native adhesive protein. Structure-function studies of the receptor recognition domains on adhesive molecules led to development of a new class of platelet inhibitors acting at the membranereceptors responsible for anchoring of platelets to the vessel wall and linking them to each other.
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Twomey, John R., Vivek Sundaram, Krishna Madhavan, and Wei Tan. "Characterization of Carbon Nanotube-Conjugated Collagen Composite Matrix Mechanics." In ASME 2007 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2007. http://dx.doi.org/10.1115/sbc2007-176643.
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In the case of vascular grafts, enhanced mechanical properties of engineered tissue constructs are required in order to function properly in mechanically-active physiologic conditions. It is proposed that a composite matrix constructed of type I collagen, fibronectin, and covalently-functionalized single-walled carbon nanotubes (SWNTs) will provide the desired mechanical properties required for the development of implantable tissues capable of withstanding high-stress environments.
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Guterl, Clare Canal, Tyler Kim, Steven B. Nicoll, and James C. Iatridis. "Genipin-Crosslinked Fibrin Hydrogels Modified With Collagen or Fibronectin as an Annulus Fibrosus Sealant." In ASME 2012 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2012. http://dx.doi.org/10.1115/sbc2012-80913.
Full textAbstract:
Low back pain (LBP) affects 80% of all adults at some point in their lives [1], and is often associated with intervertebral disc (IVD) degeneration, therefore, restorative solutions are of high importance. The IVD is composed of an outer ring of annulus fibrosus (AF) containing the pressurized nucleus pulposus (NP) center. NP replacement approaches and IVD procedures such as discectomy or discography involve rupture of the AF tissue for diagnosis or repair purposes. Even small defects to the outer AF can accelerate IVD degeneration [2]. Current closure techniques are limited to sutures and offer little in the way of tissue replacement or mechanical restoration of the AF [3]. A genipin crosslinked fibrin hydrogel offers some promise as an adhesive AF sealant [4]. Fibrin is FDA approved and genipin is a plant based chemical crosslinker with low cytotoxicity used with a variety of materials including fibrin for articular cartilage engineering [5]. Genipin crosslinked fibrin is a tunable material with comparable mechanical properties to native AF tissue and although cells remained viable on the gel, the relatively slow proliferation and presence of abnormal cells with rounded morphology motivated further optimization. To improve cytomorphology, key extracellular matrix proteins, collagen and fibronectin, were added to the formulation. It was hypothesized that the addition of these proteins would maintain the mechanical properties of this gel while improving cell morphology and viability. Several additional analyses were included to further characterize this gel including push-out strength, degradation and contraction tests.
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Bastida, E., G. Escolar, R. Castillo, A. Ordinas, and J. J. Sixma. "FIBRONECTIN IS REQUIRED FOR PLATELET ADHESION AND FOR THROMBUS FORMATION ON SUBENDOTHELIUM AND COLLAGEN SURFACES." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643558.
Full textAbstract:
Fibronectin (FN) plays a role in several adhesion mediated functions including the interaction of platelets with subendothelium.We investigated the role of plasma FN in platelet adhesion and platelet thrombus formation under flow conditions.To do this we used two different perfusion models:1)the annular chamber with α -chymotrypsin-treated rabbit vessel segments and 2)the flat chamber with coverslips coated with fibrillar purified human collagen type III.Perfusates consisted of washed platelets, and washed red blood celIs,suspended in normal or FN-depleted plasma.Perfusions were carried out for 10 min at shear rates of 300 or 1300 sec™1 Platelet deposition and thrombus dimensions were morphometrically evaluated by a computerized system. We found that depletion of plasma FN significantly reduced the percentage of total coverage surface and percentage of platelet thrombus, at both shear rates studied, and in both perfusion systems (p < 0.01)(p < 0.01).The dimensions of the platelet thrombi formed in perfusions at high shear rate were also significantly reduced in perfusions carried out with FN-depleted plasma.(p < 0.01). Addition of purified FN to FN-depleted perfusates restored all the values to those measured in the control perfusions.These results indicate that, in addition to supporting platelet adhesion to the subendothelium and to fibrillar collagen, FN contributes to platelet thrombus formation under flow conditions.
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Kioczko, J., M. Wojtukiewicz, M. Bielawiec, and E. Pilecka. "FACTOR XIII AND ITS NATURAL PLASMATIC TARGETS:FIBRINO-GEN,FIBRONECTIN AND ALPHA2-ANTIPLASMIN IN MALIGNANT DISEASE." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1643208.
Full textAbstract:
The relevance of fibrin formation and its stabilization in tumour growth and metastasis formation seems to be important but it still remains unclear.The role of F.XIII in this process may emerge from a variety of mechanisms it is involved in,including crosslink of fibrin molecules,α2AP incorporation into fibrin and reciprocal crosslink of fibronectin,fibrin and collagen. We studied F.XIII and its natural targets such as fibrinogen , fibronectin and α2AP in 61 patients with various inoperable neoplasms (21 with Ca mammae,18 with Ca ventriculi and 22 with Melanoma malignum).F.XIII activity was measured by dansylcadaverine incorporation method acc.to Lorand et α2F.XIII subunit “a” and “b”, f ibronect in , α2AP , AT III, α22M , α2AT and C-I plasma concentrations were estimated by means of rocket immunoelectrophoresis acc.to Laurell using monospecific antisera (Behringwerke AG,Marburg).The following tests were performed in all cases:fibrinogen concentration, euglobulin lysis time and serum FDP.In comparison with healthy subjects the patients revealed statistically significant decrease in F.XIII activity and its subunit “a“ and “b“ concentrations concomitant with lowered fibronectin and α2AP levels.No statistically significant correlations were found between F.XIII and its natural plasmatic substrates.Furthermore,the malignant patients showed decreased AT III concentrations,whereas α2CiI and α2AT levels were elevated.Prolongation of ELT concomitant with an increase of FDP concentration were also found. Differences in F.XIII level and in other haemostasis parameters were stated between different tumour types. Our data indicate that the role of F.XIII in malignancy is not limited to fibrin stabilization but its interactions with fibronectin and α2AP should be taken into account.