The mechanisms by which platelets accumulate in the lung in the setting of inflammation are not completely understood. At the Thursday Platelets and Inflammation Oral Session, Simon Cleary, Ph.D. presented a study of platelet responses in the mouse lung microcirculation. He demonstrated that:
- Bacterial lipopolysaccharides (LPS) significantly increased platelet numbers and CD41 expression of both neutrophil-associated and non-neutrophil-associated platelets as measured in lung sections.
- In a murine living lung model, more adherent platelets were present in the pulmonary microvasculature following LPS inhalation than in controls, as measured by intravital video microscopy.
This study identified both neutrophil-independent and neutrophil-associated platelet activation following LPS inhalation.
This study was designed to measure platelet activation and adhesion in the mouse lung microcirculation following LPS inhalation, using immunohistochemistry and intravital microscopy.
Frozen lung sections were collected from PBS control mice (n=6) or mice treated with 5 mg/kg LPS (n=7, O55:B5, i.n., +48h). Immunohistochemistry and image analysis were used to measure the number of CD41+ platelets, activation status (platelet CD41 expression), and platelet/neutrophil interactions by evaluating platelet association with neutrophil elastase. LPS increased platelet number (60%) in lung sections and CD41 expression on platelets (38%) on both platelets associated with neutrophils and platelets not associated with neutrophils.
Using the same LPS challenge protocol, platelets in the lungs of Pf4-cre×mTmG mice (n=4, cells not expressing Pf4 are tomato fluorescent protein+, platelets expressing Pf4 are green fluorescent protein+) were imaged using a thoracic window and multiphoton microscopy in order to observe the effect of LPS on platelet adhesion in the living murine lung. Frame-to-frame tracking of intravital video microscopy revealed a significantly increased number of adherent platelets per frame in the pulmonary microvasculature following LPS inhalation (47%).
As reported here, LPS-induced platelet activation and adhesion in the murine pulmonary circulation can be measured using two imaging technologies. Spatial patterns of platelet activation are supportive of the existence of both neutrophil-independent and neutrophil-associated platelet activation following inhalation of LPS.