OC 41.2 - Coagulation factor XII-independent contribution of kallikrein to contact pathway-mediated thrombin generation in mouse whole blood

Monday, June 26, 2023

15:00 – 15:15 ET

Room: 516

Session Available As: Live-Stream and On-Demand

Co-Author(s)

Jun Wan, PhD (he/him/his)

Research Associate
Cyrus Tang Hematology Center, Soochow University
Suzhou, Jiangsu, China (People's Republic)
RK

Rohan Kathuri

Undergraduate
University of North Carolina at Chapel Hill
Chapel Hill, North Carolina, United States
SD

Sophia Dhrolia

Undergraduate
University of North Carolina at Chapel Hill
Chapel Hill, North Carolina, United States
Mark Roest, PhD (he/him/his)

Clinical Director
Department of Platelet pathophysiology, Synapse Research Institute, Maastricht, the Netherlands
Maastricht, Limburg, Netherlands
Alisa S. Wolberg, PhD

Professor
Univerisity of North Carolina at Chapel Hill
Chapel Hill, North Carolina, United States
Rafal Pawlinski, PhD

Professor of Medicine
Division of Hematology and Blood Research Center, University of North Carolina at Chapel Hill
Chapel Hill, North Carolina, United States
PB

Pavan K. Bendapudi

Research Assistant Professor
Division of Hemostasis and Thrombosis, Beth Israel Deaconess Medical Center, Boston, MA
Boston, Massachusetts, United States
Nigel Mackman, PhD

Professor
Univerisity of North Carolina at Chapel Hill
Chapel Hill, North Carolina, United States

Presenting Author(s)

Steven P. Grover, PhD

Research Assistant Professor
Division of Hematology and Blood Research Center, University of North Carolina, Chapel Hill
Chapel Hill, North Carolina, United States

Background: Kallikrein is a reciprocal activator of coagulation factor (F)XII in the contact pathway of coagulation. In addition, kallikrein can directly activate FIX, bypassing FXII and FXI. The contribution of kallikrein-mediated activation of FIX to thrombin generation in complex biological systems, such as whole blood, has not been clear.

Aims: We sought to determine the relative contribution of FXII-dependent and -independent activation of coagulation in mouse whole blood.

Methods: Silica-initiated thrombin generation was evaluated in whole blood from mice deficient for kallikrein, FXII, FXI, FIX, or expressing low levels of tissue factor and compared to wildtype controls. Whole blood thrombin generation was also evaluated in the presence or absence of the kallikrein inhibitor berotralstat and/or the FXIIa inhibitor garadacimab.

Results: Compared to controls, FXII and FXI deficiency only partially reduced silica-initiated whole blood thrombin generation. In contrast, FIX deficiency completely abrogated silica-initiated whole blood thrombin generation. This differential phenotype was not observed in silica-initiated plasma thrombin generation. These findings suggest that contact pathway activation supports thrombin generation in a FIX-dependent, but partially FXII- and FXI-independent, manner in whole blood. Inhibition of kallikrein significantly reduced silica-initiated thrombin generation in FXII-deficient whole blood. Conversely, inhibition of FXIIa significantly reduced silica-initiated thrombin generation in kallikrein-deficient whole blood. These findings indicate that kaillikrein can facilitate contact pathway-mediated thrombin generation independent of FXII. Interestingly, in wildtype whole blood, no additive effect of FXIIa and kallikrein inhibition was observed. When FXIIa and kallikrein were both inhibited silica-initiated thrombin generation was further reduced in whole blood from mice expressing low levels of tissue factor compared to controls.

Conclusion(s): Here we demonstrate that kallikrein can support contact pathway-mediated thrombin generation independent of FXII in murine whole blood. These findings are consistent with a contribution of kallikrein-mediated activation of FIX to contact pathway-mediated thrombin generation in the complex milieu of whole blood.

Learning Objectives:

Upon completion, participant will be able to describe the experimental setup and data acquisition process for the mouse whole blood thrombin generation assay.
Upon completion, participant will be able to evaluate results from mouse whole blood thrombin generation assays.
Upon completion, participant will be able to evalaute the contribution of alternate coagulation reactions to mouse whole blood thrombin generation in experiments using pharmacologic and genetic interventions.