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Thank you for attending THSNA 2026. The virtual meeting is now closed.
Thank you for attending THSNA 2026. The virtual meeting is now closed.
Presentation Details
| Uptake of circulating DNA by platelets promotes lung injury in Sickle Cell Disease(THSNA �Travel Awardee) FNU Manikanta1, Tomasz W kaminski1, Ayyanar Sivanantham1, Nicholas Swendrowski1, Joshua Field1, 2, Shane Howe1, Prithu Sundd1, 2. 1Versiti Blood Research Institute, Milwaukee, WI, USA.2Medical College of Wisconsin, Milwaukee, WI, USA |
Abstract
Background: Sickle Cell Disease (SCD) is the most common monogenic disorder affecting people of African ancestry. Acute chest syndrome (ACS), a type of lung injury is a major cause of respiratory failure among SCD patients hospitalized with vaso-occlusive pain episodes (VOEs). Lung vaso-occlusion by neutrophil-platelet-erythrocyte aggregates and development of thrombocytopenia contributes to development of ACS. Platelet activation promotes the formation of neutrophil-platelet aggregates, suggesting that therapies designed to attenuate platelet activation may prevent ACS in SCD. Circulating DNA, a potent damage associated molecular patterns (DAMPs) molecules is abundant in the plasma of SCD patients and further upregulated during ACS. However, how circulating DNA promotes platelets activation and development of ACS in SCD remains unknown. Platelets express nucleic acid receptor toll-like receptor 9 (TLR9) on their surface enabling direct recognition and binding of circulating DNA. However, whether and how recognition of circulating DNA by surface TLR9 on platelets promotes development of ACS in SCD remains unknown. Objectives: To determine whether uptake of circulating DNA by surface TLR9 leads to platelet activation and development of ACS in SCD. Methods: Townes knock-in humanized control (AS) and SCD (SS) mice were (IV) intravenously administered oxy-hemoglobin (oxy-Hb) ± TLR9 blocking Ab to induce vaso-occlusive crisis. Use of intravital (in vivo) multi-photon-excitation fluorescence microscopy of intact lung in live SCD mice was used to assess lung vaso-occlusion. Proteomic analysis of platelets was conducted using mass spectrometry and data were analyzed with Perseus. Gene set enrichment analysis (GSEA) was performed to identify enriched pathways and biological processes. Imaging-Flow-Cytometry, standard flow cytometry and DNA quantification study to assess the uptake of circulating DNA by SCD mice platelets ± TLR9 blocking Ab or inhibitor. Results: IV challenge with oxy-Hb led to the occlusion of pulmonary arterioles by neutrophil-platelet aggregates in SCD but not control mice. Phospho-proteomic analysis revealed the upregulation of platelet activation pathway in IV oxy-Hb challenged SCD but not control mice platelets. Imaging-Flow-Cytometry, standard flow cytometry and DNA quantification results show that increased levels of circulating DNA uptake by platelets of IV oxy-Hb challenged SCD than control mice. Interestingly, anti-TLR9 function blocking Ab pretreatment reduced the uptake of circulating DNA by platelets in IV oxy-Hb challenged mice. Finally, intravital lung microscopy in SCD mice revealed that development of lung vaso-occlusion (neutrophil-platelet aggregation) was significantly abolished following pretreatment with function blocking TLR9 Ab. Conclusion: Our preliminary findings demonstrate for the first time that surface TLR9-dependent uptake of circulating DNA by platelets leads to platelet activation, lung vaso-occlusion by neutrophil-platelet aggregates and development of ACS in SCD. Our findings also suggest the targeting TLR9 signaling in platelets could be a potential therapy for ACS. Currently, experiments are in progress to identify the TLR9-dependent downstream signaling driving platelet activation in SCD.
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No part of this publication may be reproduced, distributed, or transmitted in any form or by any means, including photocopying, recording, or other electronic or mechanical methods, without the prior written permission of the author.