PAR4-Mediated PI3K/Akt and RhoA/ROCK Signaling Pathways Are Essential for Thrombin-Induced Morphological Changes in MEG-01 Cells

Yunkyung Heo, Hyejin Jeon, Wan Namkung

Research output: Contribution to journalArticlepeer-review

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 languageEnglish
Article number776
JournalInternational journal of molecular sciences
Volume23
Issue number2
DOIs
Publication statusPublished - 2022 Jan 1

Bibliographical note

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).

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).

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

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