Research in the Sheffield laboratory focuses on plasma, the liquid component of the blood. Plasma components regulate hemostasis, and plasma is the only product of whole blood processing that can be readily frozen, thawed and transfused.
Understanding the biological functions of plasma components and their efficacy in the regulation of hemostasis can provide insight towards the development of better transfusion practice and more efficient utilization of donated plasma.
Plasma is usually transfused in patients to prevent or stop excessive bleeding. We investigate the use of transfusable plasma as a biological medication by testing its ability to support in vitro clotting, or to suppress bleeding, in mouse models. We compare these results with those obtained with proposed alternatives to transfusable plasma, such as prothrombin complex concentrates (PCC) – a plasma protein product. To facilitate this work, we have created a mouse model of global plasma protein deficiency to test the role of various plasma components in regulating coagulation. Our results led us to propose that high fibrinogen levels, not FVIII levels, in the transfused plasma, more reliably reduce bleeding. We also investigate the ability of plasma or PCC to reverse the anticoagulant properties of ‘blood-thinning’ drugs. Such intervention is necessary when a patient on blood thinning medication requires emergency surgery. Our research in this area has shown that, for some oral anticoagulants, PCC is superior to plasma transfusion to achieve reversal. For other anticoagulants, neither plasma nor PCCs were effective. These findings correlate well with clinical studies and provide further evidence on the best use of plasma and plasma protein products. We have also investigated a potential antidote.
Eltringham-Smith LJ, Lei X, Reheman A, Lambourne MD, Pryzdial EL, Ni H, Sheffield WP: The fibrinogen but not the Factor VIII content of transfused plasma determines its effectiveness at reducing bleeding in coagulopathic mice. Transfusion 2015; 55:1040-1050.
Sheffield WP, Lambourne MD, Eltringham-Smith LJ, Bhakta V, Arnold DM, Crowther MA: γT -S195A thrombin reduces the anticoagulant effects of dabigatran in vitro and in vivo. J Thromb Haemost 2014; 12:1110–1115.
Lambourne MD, Eltringham‐Smith LJ, Gataiance S, Arnold DM, Crowther MA, Sheffield WP: Prothrombin complex concentrates reduce blood loss in murine coagulopathy induced by warfarin, but not in that induced by dabigatran etexilate. J. Thromb Haemost 2012; 10:1830–1840.
Plasma Manufacturing and Quality Assessment
Our research supports the provision of high quality plasma and plasma products for Canadian patients.
Our scientific studies provide the blood system with evidence upon which more rational regulations and/or improved processes can be based.
We seek to better understand the quality of stored plasma products for transfusion, and the impact of the manufacturing and storing process on these products. Recently, we have investigated levels of FVIII and seven other factors in frozen plasma, and we have demonstrated that thawed cryosupernatant plasma can be refrigerated for up to five days (compared with current regulation of 24 hours) prior to transfusion without compromising the quality or the safety of the thawed product.
Bhakta V, Jenkins C, Ramirez‐Arcos S, Sheffield WP: Stability of relevant plasma protein activities in cryosupernatant plasma units during refrigerated storage for up to 5 days postthaw. Transfusion 2014; 54:418–425.
Sheffield WP, Bhakta V, Talbot K, Pryzdial ELG, Jenkins C: Quality of frozen transfusable plasma prepared from whole blood donations in Canada: An update. Transfus Apher Sci 2013; 49:440–446.
Sheffield WP, Bhakta V, Jenkins C, Devine DV: Conversion to the buffy coat method and quality of frozen plasma derived from whole blood donations in Canada. Transfusion 2012; 50:1043–1049.
Plasma Proteins and Recombinant Engineered Plasma Proteins
Inspired by the properties of natural plasma proteins and using recombinant DNA technology, our group is developing novel plasma proteins to target specific therapeutic applications.
There are as yet unmet therapeutic needs in the areas of coagulation, and our research efforts aim to advance discovery in this field through the optimization of naturally occurring plasma proteins. Working in this area also gives us expertise with which to appraise new commercial plasma protein products to inform decisions on whether Canadian Blood Services should distribute them to Canadian hospitals.
We alter the structure of plasma proteins to modify their procoagulant or anticoagulant properties, or their circulatory lifetime. Ongoing projects include: 1) Structure/function studies of the serine protease inhibitor (serpin) family members alpha-1-proteinase inhibitor, antithrombin, heparin cofactor II, and alpha-2-antiplasmin; 2) Fate of mutant albumin molecules in vivo in rabbits and mice, and efficacy in animal models of hemorrhage or thrombosis; 3) Gene fusion approaches to altering the clearance or distribution of plasma proteins (e.g. factors VII and VIII).
Sheffield WP, Eltringham-Smith LJ, Gataiance S, Bhakta V: A plasmin-activatable thrombin inhibitor reduces experimental thrombosis and assists experimental thrombolysis in murine models. J Thromb Thrombolysis 2015; 39:443-51.
Scott BM, Matochko WL, Gierczak RF, Bhakta V, Derda R, Sheffield WP: Phage display of the serpin alpha-1 proteinase inhibitor randomized at consecutive residues in the reactive centre loop and biopanned with or without thrombin. PLoS One 2014; 9:e84491.
Bhakta V, Gierczak RF, Sheffield WP: Expression screening of bacterial libraries of recombinant alpha-1 proteinase inhibitor variants for candidates with thrombin inhibitory capacity. J Biotech 2013; 168:373–381.