DOC PREVIEW
FVII Dependent Coagulation Activation

This preview shows page 1-2-3 out of 8 pages.

Save
View full document
View full document
Premium Document
Do you want full access? Go Premium and unlock all 8 pages.
Access to all documents
Download any document
Ad free experience
View full document
Premium Document
Do you want full access? Go Premium and unlock all 8 pages.
Access to all documents
Download any document
Ad free experience
View full document
Premium Document
Do you want full access? Go Premium and unlock all 8 pages.
Access to all documents
Download any document
Ad free experience
Premium Document
Do you want full access? Go Premium and unlock all 8 pages.
Access to all documents
Download any document
Ad free experience

Unformatted text preview:

FVII Dependent Coagulation Activation in Citrated Plasma byPolymer HydrogelsBrendan J. Casey,†Adam M. Behrens,†John R. Hess,‡Zhongjun J. Wu,§Bartley P. Griffith,§and Peter Kofinas*,†Fischell Department of Bioengineering, University of Maryland, 2330 Jeong H. Kim Engineering Building,College Park, Maryland 20742, United States, Department of Pathology, University of Maryland School ofMedicine, University of Maryland Medical Center, Blood Bank N2W50a, Baltimore,Maryland 21201, United States, and Department of Surgery, University of Maryland School of Medicine, MedicalSchool Teaching Facility Building Room 434F, 10 South Pine Street, Baltimore, Maryland 21201, United StatesReceived May 31, 2010; Revised Manuscript Received October 19, 2010Polymer hydrogels containing positively charged functional groups were used to investigate the critical materialand biological components of FVII activation and subsequent fibrin formation in citrated plasma. A FVIIa ELISAconfirmed the ability of the polymer to induce FVII activation and provided insight into the material parameterswhich were influential in this activation. Experiments utilizing coagulation factor depleted and inhibited plasmasindicated that FVII, FX, FII, and FI are all vital to the process outlining the general mechanism of fibrin formationfrom the onset of FVII activation. Dynamic mechanical analysis and swelling experiments were used to establisha critical correlation between polymer microstructure and FVII activation.IntroductionBlood coagulation is a complex process involving a seriesor “cascade” of enzyme activation reactions. At each stage aprecursor protein (zymogen) is converted to an active proteaseby cleavage of one or more peptide bonds in the precursormolecule. Activation of the coagulation cascade is believed tooccur through two main pathways: the contact activationpathway and/or the tissue factor pathway. These two pathwaysconverge in the activation of a final common pathway, whichin turn results in the formation of a cross-linked fibrin network.1,2The activation and sustainment of the tissue factor pathwayinvolves the coordination of a multitude of enzymes andcofactors including factor VII (FVII), calcium, tissue factorpathway inhibitor (TFPI), tissue factor (TF), and a phospholipidsurface.3Human coagulation FVII is a single chain, glycoproteinwhich circulates in normal human blood, and is the mainactivator of the tissue factor pathway.3,4Cleavage of a singlepeptide bond results in a structural change that activates thezymogen, transforming it into a potent vitamin K-dependentserine protease.5This structural change also allows for theprotein to effectively bind to its cofactor, TF, in the presenceof calcium.6,7The distribution of TF is carefully coordinatedin a hemostatic layer surrounding the vascular endothelium.8-12Upon disruption of the endothelium TF forms a stable complexwith activated FVII (FVIIa) within blood, and this complex (TF-FVIIa) is capable of directly initiating the common pathwayvia the activation of FX or indirectly through the activation ofFIX, which in turn is able to activate FX.13TF is most effectiveas a procoagulant when incorporated into a phospholipidmembrane, a surface which is typically provided by platelets.14-16TFPI is the main inhibitor of the tissue factor pathway, exertingits inhibitory action on the FVIIa-TF-FX complex.17There has been considerable research into the effects ofcharged surfaces on the coagulation cascade. The ability ofnegatively charged surfaces, such as glass or the mineral kaolin,to initiate the contact activation pathway in blood or plasmavia activation of FXII has been known for over 50 years.18-20Recently, research has shown that negatively charged poly-phosphate species are capable of enhancing fibrin clot structurein buffer solutions.21It had previously been discovered byBjourn et al.,22that highly positively charged materials havethe ability to activate FVII. Later research by Pederson et al.23explained the ability of these positively charged surfaces toactivate FVII via an autoactivation theory. Other research groupshave shown that amine containing polymers such as poly(lysine)are capable of enhancing the activation of various coagulationproteins, including FVII, FX, and FII in various buffers.23-29Although the ability of charged surfaces to induce specificcoagulation factor activation is well-known, the roles thatspecific material properties have in the activation is not wellunderstood. Our research helps to establish a better understand-ing of the effects that material properties, specifically theinterplay of electrostatic charge and microstructure, have onFVII dependent coagulation activation. A better understandingof the way in which specific material properties affect the body’sblood clotting response is vital to the development of next-generation blood contacting biomaterials.Materials and MethodsMaterials. Acrylamide, N,N′-methylenebisacrylamide (BIS), N,N,N′,N′-tetramethylethylenediamine (TEMED), 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (HEPES), ammonium persulfate (APS), andhuman fibrinogen (FI) were purchased from Sigma-Aldrich (Milwaukee,WI). N-(3-Aminopropyl)methacrylamide hydrochloride (APM) waspurchased from Polysciences (Warrington, PA). N-(2-Hydroxyethyl)-methacrylamide (HEM) was purchased from Monomer-Polymer (Tre-vose, PA). Sodium hydroxide (NaOH), sodium phosphate monobasic,sodium chloride (NaCl), potassium chloride (KCl), and hydrochloricacid (HCl) were purchased from Mallinckrodt Baker (Phillipsburg, NJ).* To whom correspondence should be addressed. E-mail: [email protected].†Fischell Department of Bioengineering, University of Maryland.‡Department of Pathology, University of Maryland School of Medicine.§Department of Surgery, University of Maryland School of Medicine.Biomacromolecules 2010, 11, 3248–3255324810.1021/bm101147w  2010 American Chemical SocietyPublished on Web 11/08/2010Dextrose (anhydrous, ACS grade) was purchased from EMD Chemicals(Madison, WI). Deionized water (DI water) was obtained using aMillipore Super-Q water system (Billerica, MA). Plastic-capped glassvials were purchased from VWR Scientific (West Chester, PA). 24-Well cell culture plates were purchased from Corning Life Science(Corning, NY).Normal, human source plasma (pooled, sterile filtered, 4% (w/v)sodium citrate), and FI deficient plasma (pooled, sterile filtered, 4%(w/v) sodium citrate) were purchased from Vital


FVII Dependent Coagulation Activation

Download FVII Dependent Coagulation Activation
Our administrator received your request to download this document. We will send you the file to your email shortly.
Loading Unlocking...
Login

Join to view FVII Dependent Coagulation Activation and access 3M+ class-specific study document.

or
We will never post anything without your permission.
Don't have an account?
Sign Up

Join to view FVII Dependent Coagulation Activation 2 2 and access 3M+ class-specific study document.

or

By creating an account you agree to our Privacy Policy and Terms Of Use

Already a member?