Long-lived lung megakaryocytes contribute to platelet recovery in thrombocytopenia models
Alison C. Livada, … , James Palis, Craig N. Morrell
Alison C. Livada, … , James Palis, Craig N. Morrell
Published September 20, 2024
Citation Information: J Clin Invest. 2024;134(22):e181111. https://doi.org/10.1172/JCI181111.
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Research Article Hematology

Long-lived lung megakaryocytes contribute to platelet recovery in thrombocytopenia models

Abstract

Lung megakaryocytes (Mks) are largely extravascular with an immune phenotype (1). Because bone marrow (BM) Mks are short lived, it has been assumed that extravascular lung Mks are constantly “seeded” from the BM. To investigate lung Mk origins and how origin affects their functions, we developed methods to specifically label lung Mks using CFSE dye and biotin delivered via the oropharyngeal route. Labeled lung Mks were present for up to 4 months, while BM Mks had a lifespan of less than 1 week. In a parabiosis model, lung Mks were partially replaced over 1 month from a circulating source. Unlike tissue-resident macrophages, using MDS1-Cre-ERT2 TdTomato mice, we found that lung Mks arose from hematopoietic stem cells. However, studies with FlkSwitch mTmG mice showed that lung Mks were derived from a Flt3-independent lineage that did not go through a multipotent progenitor. CFSE labeling to track lung Mk–derived platelets showed that approximately 10% of circulating platelets were derived from lung-resident Mks at steady state, but in sterile thrombocytopenia this was doubled (~20%). Lung-derived platelets were similarly increased in a malaria infection model (Plasmodium yoelii) typified by thrombocytopenia. These studies indicate that lung Mks arise from a Flt3– BM source, are long-lived, and contribute more platelets during thrombocytopenia.

Authors

Alison C. Livada, Kathleen E. McGrath, Michael W. Malloy, Chen Li, Sara K. Ture, Paul D. Kingsley, Anne D. Koniski, Leah A. Vit, Katherine E. Nolan, Deanne Mickelsen, Grace E. Monette, Preeti Maurya, James Palis, Craig N. Morrell

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Figure 3

Lung-resident Mks produce platelets.

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Lung-resident Mks produce platelets. (A) Mice were simultaneously given ...
(A) Mice were simultaneously given CFSE o.p. and anti-CD42c platelet-labeling antibody i.p. Lung-derived platelets were quantified as CFSE+CD42c– platelets by flow cytometry in representative plots. SSC, side scatter. (B) Results showed that 2%–5% of total platelets were lung derived (n = 9–12 from 3 independent experiments), and when normalized to CFSE+ Mks (n = 5; results are representative of 2 independent experiments), a maximum of approximately 10% of the platelets were lung-resident, Mk-derived. (C) CFSE data were validated by biotin delivered o.p. and by quantification of streptavidin-binding platelets (Plts) (n = 4; results are representative of 2 independent experiments). (D) CFSE+ lung-derived and CFSE– platelets were agonist stimulated, and platelet activation was quantified by flow cytometry for CD62P surface expression. Lung-derived platelets and BM-derived platelets responded similarly to thrombin and U46619 (n = 4 per group; results are representative of 2 independent experiments). (E) Isolated platelets from mice treated with CFSE o.p. were stimulated with PAM3CSK4 (PAM) (10, 50, or 100 μg/mL) or Tyrode’s or with loxoribine (Loxo) (250, 400, or 500 μg/mL) or 1:1 DMSO/H2O, and CD62P surface expression was compared in either CFSE+ or CFSE– platelets (n = 4 per group; results are representative of 2 independent experiments). Data indicate the mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, by (D and E) multiple t tests with Holm-Šidák multiple-comparison correction.