Published September 15, 2023 - More info
Protease-activated receptor 4 (PAR4) (gene F2RL3) harbors a functional dimorphism, rs773902 A/G (encoding Thr120/Ala120, respectively) and is associated with greater platelet aggregation. The A allele frequency is more common in Black individuals, and Black individuals have a higher incidence of ischemic stroke than White individuals. However, it is not known whether the A allele is responsible for worse stroke outcomes. To directly test the in vivo effect of this variant on stroke, we generated mice in which F2rl3 was replaced by F2RL3, thereby expressing human PAR4 (hPAR4) with either Thr120 or Ala120. Compared with hPAR4 Ala120 mice, hPAR4 Thr120 mice had worse stroke outcomes, mediated in part by enhanced platelet activation and platelet-neutrophil interactions. Analyses of 7,620 Black subjects with 487 incident ischemic strokes demonstrated the AA genotype was a risk for incident ischemic stroke and worse functional outcomes. In humanized mice, ticagrelor with or without aspirin improved stroke outcomes in hPAR4 Ala120 mice, but not in hPAR4 Thr120 mice. P selectin blockade improved stroke outcomes and reduced platelet-neutrophil interactions in hPAR4 Thr120 mice. Our results may explain some of the racial disparity in stroke and support the need for studies of nonstandard antiplatelet therapies for patients expressing PAR4 Thr120.
Frederik Denorme, Nicole D. Armstrong, Michelle L. Stoller, Irina Portier, Emilia A. Tugolukova, Rikki M. Tanner, Emilie Montenont, Seema Bhatlekar, Mark Cody, John L. Rustad, Abigail Ajanel, Neal D. Tolley, Darian C. Murray, Julie L. Boyle, Marvin T. Nieman, Steven E. McKenzie, Christian Con Yost, Leslie A. Lange, Mary Cushman, Marguerite R. Irvin, Paul F. Bray, Robert A. Campbell
It has been over three decades since the thrombin receptor was cloned and its mechanism of action revealed (1). We now know that thrombin-activated G-protein coupled receptors (PARs) are a group of proteins that play a role in various physiological processes, including stroke. When thrombin, a key enzyme involved in blood clotting, binds to PARs, it triggers a cascade of cellular events. PAR-1 and PAR-4 are the two main thrombin-activated receptors relevant to stroke.In stroke, the activation of PARs by thrombin can contribute to protective and detrimental effects (2). On one hand, PAR activation can promote neuroprotection, tissue repair, and angiogenesis that may aid in stroke recovery. On the other hand, excessive PAR activation can lead to inflammation, blood-brain barrier disruption, and neuronal damage, worsening stroke outcomes. Notably, PAR-1 activation can exacerbate brain injury in stroke by promoting inflammation and blood-brain barrier dysfunction, while PAR-1 blockade has potential therapeutic benefits in reducing stroke-related damage and improving outcomes.As for PAR-4, its role in stroke is less well-defined. PAR-4 activation may contribute to platelet activation and thrombosis, which could worsen stroke outcomes. However, further investigations are needed to fully understand the impact of PAR-4 in stroke pathophysiology.
In this issue of the JCI, Denorme and colleagues (3) investigated the potential contribution of a known PAR-4 genetic variant with high prevalence in patients with African ancestry to ischemic stroke (IS). The F2RL3 gene that encodes PAR harbors a functional genetic variant, rs773902 A/G (encoding Thr120/Ala120, respectively) and is associated with augmented platelet aggregation (4). The A allele frequency is more common in Black individuals, and Black individuals have a higher incidence of ischemic stroke than White individuals. To examine this subject, the authors engineered an elegant model of human PAR4 (hPAR4) that expressed the normal allele and the variant in mice using a CRISPR-Cas9 gene editing approach. The major findings were notable — mice carrying the human PAR4 risk allele had worse stroke outcomes, which correlated with more platelet-neutrophil aggregates and neutrophil extracellular traps (NETs) in brain tissue than mice with the lower-risk allele. Moreover, mice carrying the high-risk allele responded well to pharmacological PAR4 inhibition and to P-selectin blockade in terms of stroke outcomes. However, the mice did not respond well to contemporary anti-platelet therapies, which instead promoted cerebral hemorrhage in the stroke model. Finally, human population data indicated that Black individuals carrying the high-risk allele were more prone to ischemic stroke and worse neurological outcomes, and human platelets carrying the high-risk allele induced greater NET formation ex vivo. These findings are important because they expand upon the current understanding of the PAR4 Ala120Thr variant in vivo and its implications for stroke outcomes. They also provide unexpected and valuable pharmacogenomic hints into therapeutic approaches that may provide better outcomes for stroke patients carrying the risk variant (Thr120) than traditional anti-platelet therapies.
Conflict of interest: DEV is the CSO of Zoe Biosciences and owns stock in the company.
Copyright: © 2023, Vaughan. This is an open access article published under the terms of the Creative Commons Attribution 4.0 International License.
Reference information: J Clin Invest. 2023;133(18):e173932. https://doi.org/10.1172/JCI173932.
See the related article at The predominant PAR4 variant in individuals of African ancestry worsens murine and human stroke outcomes.