Abstract
Thrombin stimulates platelets via a dual receptor system of protease-activated receptors (PARs): PAR1 and PAR4. PAR1 activation induces a rapid and transient signal associated with the initiation of platelet aggregation, whereas PAR4 activation results in a prolonged signal, required for later phases, that regulates the stable formation of thrombus. In this study, we observed differential signaling pathways for thrombin-induced PAR1 and PAR4 activation in a human megakaryoblastic leukemia cell line, MEG-01. Interestingly, thrombin induced both calcium signaling and morphological changes in MEG-01 cells via the activation of PAR1 and PAR4, and these intracellular events were very similar to those observed in platelets shown in previous studies. We developed a novel image-based assay to quantitatively measure the morphological changes in living cells, and observed the underlying mechanism for PAR1-and PAR4-mediated morphological changes in MEG-01 cells. Selective inhibition of PAR1 and PAR4 by vorapaxar and BMS-986120, respectively, showed that thrombin-induced morphological changes were primarily mediated by PAR4 activation. Treatment of a set of kinase inhibitors and 2-aminoethoxydiphenyl borate (2-APB) revealed that thrombin-mediated morphological changes were primarily regulated by calcium-independent pathways and PAR4 activation-induced PI3K/Akt and RhoA/ROCK signaling pathways in MEG-01 cells. These results indicate the importance of PAR4-mediated signaling pathways in thrombin-induced morphological changes in MEG-01 cells and provide a useful in vitro cellular model for platelet research.
Original language | English |
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Article number | 776 |
Journal | International journal of molecular sciences |
Volume | 23 |
Issue number | 2 |
DOIs | |
Publication status | Published - 2022 Jan 1 |
Bibliographical note
Funding Information:This research was funded bythe National ResearchFoundation of Korea (NRF-2018R1A6A1A03023718) and the Ministry of Education (MOE, Korea) through the fostering project of ‘Yonsei University and Industry Cooperation Complex’ supervised by the Korea Institute for Advancement of Technology (KIAT).
Funding Information:
Funding: This research was funded by the National Research Foundation of Korea (NRF-2018R1A6A1A03023718) and the Ministry of Education (MOE, Korea) through the fostering project of ‘Yonsei University and Industry Cooperation Complex’ supervised by the Korea Institute for Advancement of Technology (KIAT).
Publisher Copyright:
© 2022 by the authors. Licensee MDPI, Basel, Switzerland.
All Science Journal Classification (ASJC) codes
- Catalysis
- Molecular Biology
- Spectroscopy
- Computer Science Applications
- Physical and Theoretical Chemistry
- Organic Chemistry
- Inorganic Chemistry