OC 78.2 - Activation of coagulation factor XII by bacterial lipopolysaccharides

Wednesday, June 28, 2023

10:30 – 10:45 ET

Room: 515

Session Available As: On-Demand

Presenting Author(s)

Andre L. Lira da Silva, PhD (he/him/his)

Postdoctoral Scholar
OREGON HEALTH & SCIENCE UNIVERSITY
Portland, Oregon (EUA), Oregon, United States

Co-Author(s)

Cristina Puy, PhD

Research Assistant Professor
Oregon Health & Science University
Portland, Oregon, United States
Joseph E. Aslan, PhD

Associate Professor
Oregon Health & Science University
Portland, Oregon, United States
AS

Alvin Schmaier

Professor
Division of Hematology and Oncology Department of Medicine Case Western Reserve University
Cleveland, Ohio, United States
FL

Florea Lupu, PhD

Professor, Cardiovascular Biology
Oklahoma Medical Research Fundation
Oklahoma city, Oklahoma, United States
David Gailani, MD

Professor
Vanderbilt University Medical Center
Nashville, Tennessee, United States
ET

Erik I. Tucker

Chief Executive Officer
Aronora, Inc.
Portland, Oregon, United States
CL

Christina U. Lorentz

Director of Clinical Operations
Aronora, Inc
Portland, Oregon, United States
Owen J.T McCarty, PhD, FAHA

Professor and Chair of Biomedical Engineering
Oregon Health & Science University
Portland, Oregon, United States

Background: Lipopolysaccharides (LPS) released to the blood flow during infection are known to contribute to the development of disseminated intravascular coagulation (DIC). Since LPS molecules are strongly phosphorylated, they may activate FXII thus contributing to the DIC initiated by contact activation. Little is known about the direct activation of FXII by LPS.

Aims: Investigate the mechanisms underlying the activation of FXII by select LPS chemotypes.

Methods: The physical characteristics of LPS aggregates−formed by smooth (O111:B4 or O26:B6) or rough (Rd2) chemotypes of E.coli−were determined by dynamic light scattering, zeta potential (ZP), and fluorescence spectroscopy. Chromogenic substrate and plasma clotting assays were used to study the effects of LPS on FXII activation and FXIIa activity.

Results: O111:B4, O26:B6, and Rd2 produced aggregates with hydrodynamic diameters of 123±38, 213±46, 201±42nm; and ZP of −0.414.2±0.9, −23.0±2.1, −55.0±2.1mV, respectively. Using pyrene fluorescence, it was determined that the microenvironment within aggregates is most hydrophobic with Rd2. LPS showed procoagulant activity reducing the clotting time from 555±6 seconds to 234±12 for O111:B4, 208±8 for O26:B6, and 280±21 for Rd2. FXII was not activated by O111:B4 or Rd2 but was activated by O26:B6. O26:B6 showed a bell-shaped dose-response curve, suggesting a template mechanism for FXII autoactivation. The amidolytic activity of FXIIa was not affected in the presence of O111:B4, which was corroborated by the calculated values of Vmax and KM. In contrast, FXIIa amidolytic activity was increased in the presence of Rd2 and O26:B6, showing a sigmoidal kinetics curve, indicating a positive allosteric modulation. However, at higher concentrations, FXIIa activity decreased in the presence of O26:B6, which could be a result of protein structural modification and/or steric effects.

Conclusion(s): These investigations show that LPS chemotypes specifically impact the activation and activity of FXII, and as such, may differentially respond to therapeutics targeting contact pathway activation.

Learning Objectives:

Compare the effects of distinct lipopolysaccharides on the regulation of factor XII activity.
Conclude that physicochemical properties of lipopolysaccharides may differ between strains or species, and that difference may be relevant with regard to biological effects.
Apply methods that will illustrate how lipopolysaccharides interact with proteins and regulates its function.