A mouse model of the protease-activated receptor 4 Pro310Leu variant has reduced platelet reactivity
Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved..
BACKGROUND: Protease-activated receptor 4 (PAR4) mediates thrombin signaling on platelets and other cells. Our recent structural studies demonstrated that a single nucleotide polymorphism in extracellular loop 3 and PAR4-P310L (rs2227376) leads to a hyporeactive receptor.
OBJECTIVES: The goal of this study was to determine how the hyporeactive PAR4 variant in extracellular loop 3 impacts platelet function in vivo using a novel knock-in mouse model (PAR4-322L).
METHODS: A point mutation was introduced into the PAR4 gene F2rl3 via CRISPR/Cas9 to create PAR4-P322L, the mouse homolog to human PAR4-P310L. Platelet response to PAR4 activation peptide (AYPGKF), thrombin, ADP, and convulxin was monitored by αIIbβ3 integrin activation and P-selectin translocation using flow cytometry or platelet aggregation. In vivo responses were determined by the tail bleeding assay and the ferric chloride-induced carotid artery injury model.
RESULTS: PAR4-P/L and PAR4-L/L platelets had a reduced response to AYPGKF and thrombin measured by P-selectin translocation or αIIbβ3 activation. The response to ADP and convulxin was unchanged among genotypes. In addition, both PAR4-P/L and PAR4-L/L platelets showed a reduced response to thrombin in aggregation studies. There was an increase in the tail bleeding time for PAR4-L/L mice. The PAR4-P/L and PAR4-L/L mice both showed an extended time to arterial thrombosis.
CONCLUSION: PAR4-322L significantly reduced platelet responsiveness to AYPGKF and thrombin, which is in agreement with our previous structural and cell signaling studies. In addition, PAR4-322L had prolonged arterial thrombosis time. Our mouse model provides a foundation to further evaluate the role of PAR4 in other pathophysiological contexts.
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| Medienart: |
E-Artikel |
| Erscheinungsjahr: |
2024 |
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| Erschienen: |
2024 |
| Enthalten in: |
Zur Gesamtaufnahme - year:2024 |
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| Enthalten in: |
Journal of thrombosis and haemostasis : JTH - (2024) vom: 19. März |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Han, Xu [VerfasserIn] |
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Animal model |
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| Anmerkungen: |
Date Revised 06.04.2024 published: Print-Electronic UpdateOf: bioRxiv. 2023 Dec 01;:. - PMID 38077081 Citation Status Publisher |
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| doi: |
10.1016/j.jtha.2024.03.004 |
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| PPN (Katalog-ID): |
NLM369979796 |
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| 100 | 1 | |a Han, Xu |e verfasserin |4 aut | |
| 245 | 1 | 2 | |a A mouse model of the protease-activated receptor 4 Pro310Leu variant has reduced platelet reactivity |
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| 500 | |a Date Revised 06.04.2024 | ||
| 500 | |a published: Print-Electronic | ||
| 500 | |a UpdateOf: bioRxiv. 2023 Dec 01;:. - PMID 38077081 | ||
| 500 | |a Citation Status Publisher | ||
| 520 | |a Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved. | ||
| 520 | |a BACKGROUND: Protease-activated receptor 4 (PAR4) mediates thrombin signaling on platelets and other cells. Our recent structural studies demonstrated that a single nucleotide polymorphism in extracellular loop 3 and PAR4-P310L (rs2227376) leads to a hyporeactive receptor | ||
| 520 | |a OBJECTIVES: The goal of this study was to determine how the hyporeactive PAR4 variant in extracellular loop 3 impacts platelet function in vivo using a novel knock-in mouse model (PAR4-322L) | ||
| 520 | |a METHODS: A point mutation was introduced into the PAR4 gene F2rl3 via CRISPR/Cas9 to create PAR4-P322L, the mouse homolog to human PAR4-P310L. Platelet response to PAR4 activation peptide (AYPGKF), thrombin, ADP, and convulxin was monitored by αIIbβ3 integrin activation and P-selectin translocation using flow cytometry or platelet aggregation. In vivo responses were determined by the tail bleeding assay and the ferric chloride-induced carotid artery injury model | ||
| 520 | |a RESULTS: PAR4-P/L and PAR4-L/L platelets had a reduced response to AYPGKF and thrombin measured by P-selectin translocation or αIIbβ3 activation. The response to ADP and convulxin was unchanged among genotypes. In addition, both PAR4-P/L and PAR4-L/L platelets showed a reduced response to thrombin in aggregation studies. There was an increase in the tail bleeding time for PAR4-L/L mice. The PAR4-P/L and PAR4-L/L mice both showed an extended time to arterial thrombosis | ||
| 520 | |a CONCLUSION: PAR4-322L significantly reduced platelet responsiveness to AYPGKF and thrombin, which is in agreement with our previous structural and cell signaling studies. In addition, PAR4-322L had prolonged arterial thrombosis time. Our mouse model provides a foundation to further evaluate the role of PAR4 in other pathophysiological contexts | ||
| 650 | 4 | |a Journal Article | |
| 650 | 4 | |a animal model | |
| 650 | 4 | |a blood platelets | |
| 650 | 4 | |a protease-activated receptor 4 | |
| 650 | 4 | |a single nucleotide polymorphisms | |
| 650 | 4 | |a thrombin receptor | |
| 700 | 1 | |a Knauss, Elizabeth A |e verfasserin |4 aut | |
| 700 | 1 | |a Fuente, Maria de la |e verfasserin |4 aut | |
| 700 | 1 | |a Li, Wei |e verfasserin |4 aut | |
| 700 | 1 | |a Conlon, Ronald A |e verfasserin |4 aut | |
| 700 | 1 | |a LePage, David F |e verfasserin |4 aut | |
| 700 | 1 | |a Jiang, Weihong |e verfasserin |4 aut | |
| 700 | 1 | |a Renna, Stephanie A |e verfasserin |4 aut | |
| 700 | 1 | |a McKenzie, Steven E |e verfasserin |4 aut | |
| 700 | 1 | |a Nieman, Marvin T |e verfasserin |4 aut | |
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