| 1. |
- Andersson, Jonas, et al.
(författare)
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Optimal heparin surface concentration and antithrombin binding capacity as evaluated with human non-anticoagulated blood in vitro
- 2003
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Ingår i: Journal of Biomedical Materials Research. - : Wiley. - 0021-9304 .- 1097-4636. ; 67:2, s. 458-466
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Tidskriftsartikel (refereegranskat)abstract
- Contact between blood and a biomaterial surface takes place in many applications and is known to activate the coagulation and complement systems. Heparin surface coatings have been shown to reduce blood activation upon contact with artificial surfaces. To establish the optimal heparin surface concentration, blood was incubated in a tubing loop model at 37 degrees C. The tubing was coated with different surface concentrations of heparin and rotated at three different velocities. We demonstrate that the blood compatibility of a surface with regard to coagulation, complement, and platelet activation can be improved by increasing the heparin surface concentration in the 6-12 pmol antithrombin/cm2 concentration interval. The binding of factor H is not influenced by the increased heparin surface concentration, suggesting that this factor is not the primary regulator of complement on heparin surfaces. In addition, the heparin coating has no effect on the complement activation that occurs on gas surfaces in extracorporeal circuits.
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| 2. |
- Cabric, Sanja, et al.
(författare)
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A new method for incorporating functional heparin onto the surface of islets of Langerhans
- 2008
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Ingår i: Tissue Engineering. Part C, Methods. - : Mary Ann Liebert Inc. - 1937-3384 .- 1937-3392. ; 14:2, s. 141-147
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Tidskriftsartikel (refereegranskat)abstract
- A novel technique is described to conjugate macromolecular heparin complexes to cell surfaces. The method is based on the dual properties of avidin-expressing binding sites for both biotin and a macromolecular complex of heparin. A quartz crystal microbalance with dissipation monitoring (QCM-D) revealed sequential binding of biotin, avidin, and heparin complexes. Large particle flow cytometry confirmed functional integrity. Confocal microscopy of the heparinized islets showed evenly distributed fluorescence. An in vitro Chandler loop model demonstrated that the biocompatibility of the new method is comparable to the previous method used on artificial materials with regard to coagulation and antithrombin uptake. The technique presented allows human islets of Langerhans to successfully be covered with functional heparin as a means to reduce instant blood-mediated inflammatory reactions induced by the innate immune system.
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| 3. |
- Cabric, Sanja, et al.
(författare)
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Anchoring of vascular endothelial growth factor to surface-immobilized heparin on pancreatic islets : implications for stimulating islet angiogenesis
- 2010
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Ingår i: Tissue engineering. Part A. - : Mary Ann Liebert Inc. - 1937-3341 .- 1937-335X. ; 16:3, s. 961-970
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Tidskriftsartikel (refereegranskat)abstract
- In pancreatic islet transplantation, early revascularization is necessary for long-term graft function. We have shown in in vitro and in vivo models that modification with surface-attached heparin protects the islets from acute attack by the innate immune system of the blood following intraportal islet transplantation. In this study, we have investigated the ability of an immobilized conjugate composed of heparin to bind the angiogenic growth factor vascular endothelial growth factor-A (VEGF-A) as a means of attracting endothelial cells (ECs) to induce angiogenesis and revascularization. We analyzed the capacity of VEGF-A to bind to immobilized heparin and how this affected the proliferation and adherence of ECs to both artificial glass surfaces and islets. Quartz crystal microbalance with dissipation monitoring and slot-blot demonstrated the binding of VEGF-A to heparin-coated surfaces upon which ECs showed protein-dependent proliferation. Also, ECs cultured on heparin-coated glass surfaces exhibited effects upon focal contacts. Heparinized islets combined with VEGF-A demonstrated unaffected insulin release. Further, covering islets with heparin also increased the adhesion of ECs to the islet surface. Immobilized heparin on the islet surface may be a useful anchor molecule for achieving complete coverage of islets with angiogenic growth factors, ultimately improving islet revascularization and engraftment in pancreatic islet transplantation.
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| 4. |
- Cabric, Sanja, et al.
(författare)
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Islet Surface Heparinization Prevents the Instant-Blood Mediated Inflammatory Reaction in Islet Transplantation
- 2007
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 56:8, s. 2008-2015
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Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE—In clinical islet transplantation, the instant blood-mediated inflammatory reaction (IBMIR) is a major factor contributing to the poor initial engraftment of the islets. This reaction is triggered by tissue factor and monocyte chemoattractant protein (MCP)-1, expressed by the transplanted pancreatic islets when the islets come in contact with blood in the portal vein. All currently identified systemic inhibitors of the IBMIR are associated with a significantly increased risk of bleeding or other side effects. To avoid systemic treatment, the aim of the present study was to render the islet graft blood biocompatible by applying a continuous heparin coating to the islet surface.RESEARCH DESIGN AND METHODS—A biotin/avidin technique was used to conjugate preformed heparin complexes to the surface of pancreatic islets. This endothelial-like coating was achieved by conjugating barely 40 IU heparin per full-size clinical islet transplant.RESULTS—Both in an in vitro loop model and in an allogeneic porcine model of clinical islet transplantation, this heparin coating provided protection against the IBMIR. Culturing heparinized islets for 24 h did not affect insulin release after glucose challenge, and heparin-coated islets cured diabetic mice in a manner similar to untreated islets.CONCLUSIONS—This novel pretreatment procedure prevents intraportal thrombosis and efficiently inhibits the IBMIR without increasing the bleeding risk and, unlike other pretreatment procedures (e.g., gene therapy), without inducing acute or chronic toxicity in the islets.
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| 5. |
- Kristensen, Emma, et al.
(författare)
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Heparin coating durability on artificial heart valves studied by XPS and antithrombin binding capacity
- 2006
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Ingår i: Colloids and Surfaces B. - : Elsevier BV. - 0927-7765 .- 1873-4367. ; 49:1, s. 1-7
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Tidskriftsartikel (refereegranskat)abstract
- The durability and functionality of a heparin coating on artificial heart valve leaflets were evaluated with X-ray photoelectron spectroscopy (XPS) and by the coatings’ capacity to bind antithrombin. Current methods for accelerated life-time testing are based on exposing leaflets to water solutions. In this paper a method is explored, in which heart valve leaflets were exposed to a continuous high shear rate (4 L/min) of human citrated plasma. It was found that the heparin coating was stable and wear resistant enough to still be present after 3 weeks and to have about the same antithrombin uptake as coatings not exposed to circulating plasma. It was, however, partly destroyed by the test as found using XPS. We suggest that heparin chains from the upper layer of heparin have been torn off from the carrier chain, in combination with loss of heparin conjugate and plasma deposition in patches. This study showed that XPS provides additional information to biological measurements such as antithrombin uptake. XPS is therefore a valuable technique not only to characterize biomaterials but also to evaluate the effect of a performance test.
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| 6. |
- Leijon, Jonas, et al.
(författare)
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Attachment of macromolecular heparin conjugate to gelatin scaffolds improves endothelial cell infiltration
- 2013
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Ingår i: Tissue Engineering. Parts A, B and C. - 2152-4947 .- 2152-4955. ; 19:11-12, s. 1336-1348
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Tidskriftsartikel (refereegranskat)abstract
- Long-term survival of implanted cells requires oxygen and nutrients, the need for which is met by vasculari- zation of the implant. The use of scaffolds with surface-attached heparin as anchoring points for angiogenic growth factors has been reported to improve this process. We examined the potential role of surface modification of gelatin scaffolds in promoting endothelial cell infiltration by using a unique macromolecular conjugate of heparin as a coating. Compared to other heparin coatings, this surface modification provides flexible heparin chains, representing a new concept in heparin conjugation. In vitro cell infiltration of scaffolds was assessed using a three-dimensional model in which the novel heparin surface, without growth factors, showed a 2.5-fold increase in the number of infiltrating endothelial cells when compared to control scaffolds. No additional improvement was achieved by adding growth factors (vascular endothelial growth factor and/or fibroblast growth factor-2) to the scaffold. In vivo experiments confirmed these results and also showed that the addition of angiogenic growth factors did not significantly increase the endothelial cell infiltration but increased the number of inflammatory cells in the implanted scaffolds. The endothelial cell-stimulating ability of the heparin surface alone, combined with its growth factor-binding capacity, renders it an interesting candidate surface treatment to create a prevascularized site prepared for implantation of cells and tissues, in particular those sensitive to inflammation but in need of supportive revascularization, such as pancreatic islets of Langerhans.
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| 7. |
- Sanchez, Javier, et al.
(författare)
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Surface-adsorbed fibrinogen and fibrin may activate the contact activation system
- 2008
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Ingår i: Thrombosis Research. - : Elsevier BV. - 0049-3848 .- 1879-2472. ; 122:2, s. 257-263
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Tidskriftsartikel (refereegranskat)abstract
- INTRODUCTION: This study was designed to investigate whether fibrinogen, soluble desAA-fibrin, and insoluble desAABB-fibrin are able to induce clotting by triggering the plasma contact activation system when adsorbed to polystyrene. MATERIALS AND METHODS: The above-mentioned substances were individually prepared on polystyrene meshwork squares, and then exposed to a purified FXII solution or non-calcium containing plasma (citrated and dialyzed normal pooled plasma) in polystyrene cuvettes coated with surface-immobilized heparin, to completely block contact activation and the coagulation mechanism that might be induced by the cuvette surfaces. Sodium glass beads were used as the reference material. RESULTS: On exposure to purified FXII solution and plasma, all the tested materials adsorbed and activated FXII to varying degrees. This activation led to the formation of FXIa in the exposed plasma, with the highest activation occurring upon exposure to glass, desAA-fibrin and desAABB-fibrin and the lowest upon exposure to fibrinogen-adsorbed or unmodified polystyrene meshwork squares. Following recalcification, in cuvettes with surface-immobilized heparin, a spectrophotometric assay showed that the surface-exposed plasma aliquots clotted within 5 min after contact with glass, within 10 to 15 min after contact with the two forms of fibrin, and somewhat longer after contact with adsorbed fibrinogen. The longest lag phase, close to 20 min, occurred in plasma exposed to unmodified polystyrene meshwork. Whole blood deposited in surface heparinized cuvettes directly from the cubital vein did not clot during the observation time (2 h). CONCLUSIONS: These results indicate that domains induced by conformational changes in adsorbed fibrinogen and fibrin are capable of activating adsorbed proenzymes and that various forms of fibrin are considerably stronger activators of the contact activation system than are adsorbed fibrinogen or a polystyrene meshwork. The delayed coagulation in plasma exposed to the unmodified polystyrene meshwork can be explained by a two-step process: first, adsorption of fibrinogen, and second, activation of FXII. Under our experimental conditions, the adsorption and activation of FXII on fibrinogen and fibrin seems to be an important mechanism for triggering coagulation.
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| 8. |
- Thorslund, Sara, et al.
(författare)
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A PDMS-based disposable microfluidic sensor for CD4+ lymphotic counting
- 2008
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Ingår i: Biomedical microdevices (Print). - : Springer Science and Business Media LLC. - 1387-2176 .- 1572-8781. ; 10:6, s. 851-857
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Tidskriftsartikel (refereegranskat)abstract
- A refined sensor for CD4+ lymphocyte count was developed and evaluated by comparison to flow cytometry. The micropillar structured sensor surface was cast in PDMS polymer and surface modified to gain biocompatibility and CD4-cell capturing properties. The sensor works by pure capillary action and sample filling and rinsing is performed without external equipment. Whole blood samples showed acceptable agreement (79%) with flow cytometry, however when diluting the blood in PBS buffer we discovered that a larger number of cells were drawn into the sensor microchannel compared to the initial sample, explained by enhanced shear-induced cell migration. Using plasma or PBS with glycerol or albumin additives as diluting media greatly influenced this cell behavior, showing the importance of controlling the dilution media when working with devices based on capillary filling. The sensors need to be further tested with blood samples with lower CD4-counts (<500 cells/μl), which are clinically relevant.
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| 9. |
- Thorslund, Sara, et al.
(författare)
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Bioactivated PDMS microchannel evaluated as sensor for human CD4+ cells : The concept of a point-of-care method for HIV monitoring
- 2007
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Ingår i: Sensors and actuators. B, Chemical. - : Elsevier BV. - 0925-4005 .- 1873-3077. ; 123:2, s. 847-855
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Tidskriftsartikel (refereegranskat)abstract
- Up to today, the number of CD4(+) lymphocytes remains the most important biological marker to determine the clinical stage of an HIV-infection. Analysis by flow cytometry, the standard method used today, is unsuitable in many developing countries, because of high costs involved and practical inconveniences. We here present the concept of an inexpensive PDMS-based point-of-care device for CD4(-)count. A simple fluorescence microscope for stained leucocytes counting is the only detection equipment needed. The biosensor surface consists of an initial heparin-based coating that adds hydrophilicity and thromboresistance to the PDMS material. The specific capturing chemistry is based on an avidin/biotin-antibody surface architecture. Pure capillary forces draw whole blood, as well as rinsing buffer, into the biosensor channel, minimizing the need of external equipment. Detection of the captured cells was performed by fluorescence imaging of HOECHST (stains cell nuclei) and CD3-FITC signals. It was shown that the non-specific adsorption of CD4(-) leucocytes was minimal to none. and the detection could therefore be done by only counting the easy identifiable HOECHST+ cells. Characterization of the biosensor coating process was additionally performed with the quartz crystal microbalance-dissipation technique.
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| 10. |
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