OC 67.2 - Platelet factor 4 aggregates bacteria, modulating bacterial clearance, coagulopathy and the inflammatory milieu, improving sepsis outcomes

Tuesday, June 25, 2024

15:00 – 15:15 ICT

Room: 210 A-D

Presenting Author(s)

Anh Ngo, PhD

Research Post Doc Fellow
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States

Co-Author(s)

Hyunjun Kim

Research Technician
The Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
Veronica Bochenek, BA

Research Technician
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
NL

Nate Levine

Researcher
University of Pennsylvania
Philadelphia, Pennsylvania, United States
SY

Santosh Yadav

Research Associate
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
Amrita Sarkar, PhD

Research Associate Scientist II
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
LR

Lubica Rauova

Associate Professor
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
AK

Anna Kowalska

Researcher
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
Mortimer Poncz, MD

Professor of Pediatrics
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States
Kandace Gollomp

Attending Physician
Children's Hospital of Philadelphia
Philadelphia, Pennsylvania, United States

Background: During sepsis, platelets release high concentrations of platelet factor 4 (PF4), a positively-charged chemokine that strongly crosslinks polyanions including proteins in the bacterial cell wall. We previously showed that PF4 expression improves outcomes in murine sepsis models, but the precise mechanism(s) by which PF4 achieves this protective effect remains unclear.

Aims: Define how PF4 accelerates bacterial clearance to improve sepsis outcomes.

Methods: Human umbilical vein endothelial cells (HUVECs) were incubated with Escherichia (E) coli ± PF4 (0-25ug/mL) for 5h. Cell lysates were cultured on agar plates, and colony forming units (CFUs) were quantified to assess endothelial antimicrobial activity. To assess bacterial clearance in vivo, wildtype (WT) and PF4-/- mice were infused with fluorescent E coli (8x1011 CFU) and subjected to lung confocal intravital imaging. 2h post E coli infusion, liver and spleen residual CFUs were quantified. To investigate effects of PF4 in polymicrobial sepsis, cecal contents (cecal slurry, CS) from WT donor mice were pre-treated with dextrose or PF4 (100ug/mL) prior to intravenous infusion into WT or PF4-/- mice. Animals were evaluated for sepsis severity for 96h, plasma inflammatory cytokine levels were quantified, and survival analysis was performed.

Results: PF4 increased E coli uptake by HUVECs at 2h but reduced intracellular E coli proliferation from 3-5h (Fig. 1). Following E coli infusion, PF4-/- mice had increased platelet accumulation in the pulmonary microvasculature and reduced bacterial clearance (Fig. 2). In the CS sepsis model, PF4 enhanced bacterial clearance, decreased inflammatory cytokine levels, and prevented mortality.

Conclusion(s): In vitro, PF4 crosslinks E coli, initially enhancing bacterial uptake by endothelial cells, but prevents subsequent intracellular proliferation. In vivo, PF4 promotes bacterial clearance while dampening coagulopathy and inflammation, leading to improved survival in polymicrobial sepsis. These studies suggest that PF4-bacteria interaction may be an important host defense mechanism, meriting further translational studies in sepsis.