OC 75.3 - Platelet generation from circulating megakaryocytes is triggered in the lung vasculature

Wednesday, July 13, 2022

10:45 AM – 11:00 AM

Location: ExCel Center, ICC Capital Suite Room 10&11

Presentation Format: In-Person

Co-Author(s)

Xiaojuan Zhao, Ph.D

Senior Research Associate
University of Bristol
Bristol, England, United Kingdom
DA

Dominic Alibhai

LM Specialist
University of Bristol
Bristol, England, United Kingdom
TW

Tony Walsh

Senior Research Associate
University of Bristol
Bristol, England, United Kingdom
NT

Nathalie Tarassova

Ph.D student
University of Bristol
Bristol, England, United Kingdom
SB

Semra Birol

Research Associate
University of Bristol
Bristol, England, United Kingdom
CW

Christopher Williams

Senior Research Associate
University of Bristol
Bristol, England, United Kingdom
CN

Chris Neal

EM Specialist
University of Bristol
Bristol, England, United Kingdom
EA

Elizabeth Aitken

Lab technician
University of Bristol
Bristol, England, United Kingdom
Amie K. Waller, PhD

Postdoctoral researcher
University of Cambridge
Cambridge, England, United Kingdom
JB

Jose Ballester-Beltran

Research Associate
University of Cambridge
Cambridge, England, United Kingdom
PG

Peter Gunning

Professor
University of New South Wales
Syndey, New South Wales, Australia
EH

Edna Hardeman

Professor
University of New South Wales
Sydney, New South Wales, Australia
Ejaife O. Agbani, BPharm, MSc, PhD

Assistant Professor
University of Calgary
Calgary, Alberta, Canada
IH

Ingeborg Hers, PhD

Principal Investigator
University of Bristol
Bristol, England, United Kingdom
Cedric Ghevaert, MD PhD

Professor of Transfusion Medicine
Wellcome-MRC Cambridge Stem Cell Institute, University of Cambridge, Jeffrey Cheah Biomedical Centre
Cambridge, England, United Kingdom

Presenting Author(s)

AP

Alastair W. Poole

Principal Investigator
School of Physiology, Pharmacology and Neuroscience/University of Bristol
Bristol, England, United Kingdom

Background: Platelets are derived from megakaryocytes (MKs), although the generation process is unclear. Only small numbers of platelets have been produced in systems outside the body, where bone marrow and lung are proposed as sites of platelet generation . The release of platelets requires cytoskeletal re-organisation involving actin. Tropomyosin TPM4 has a role in platelet formation .

Aims: To investigate the mechanisms of platelet generation from MKs in the lung using a novel ex vivo mouse model

Methods: We established an ex vivo mouse heart-lung model (Fig. 1A) for perfusion of murine MKs. MKs and their derivates were imaged by confocal microscopy. The numbers of generated platelets (GPs) in the perfusate or retained in the lung were counted by FACS and two-photon microscopy, respectively. Morphological and functional features of GPs were determined by FACS, transmission electron microscopy and in vitro thrombosis assays. A microfluidic chamber mimicking the lung vasculature system was designed to explore the mechanism of platelet generation in the lung (Fig. 1B). MKs from TPM4-/- mice were introduced to study whether TPM4 is required for platelet generation in the lung.

Results: MKs can pass multiple times through the lung vasculature, leading to the generation of physiological levels of functional platelets (approximately 1,000~4,000 per MK), through a process involving nuclear marginalization and enucleation, prior to TPM4-dependent fragmentation. GPs demonstrated morphological and functional features comparable to control platelets. Air ventilation and healthy pulmonary endothelial cells play critical roles in platelet generation.

Conclusion(s): The ex vivo mouse heart-lung model allows the detailed study of platelet generation, and has enabled us to show that MKs can repeatedly passage through the lung vasculature under air ventilation, leading to enucleation and TPM4-dependent fragmentation to generate platelets. The findings add to our understanding of why the lung is a primary site for platelet generation.