Klotho Modulates Pro-Fibrotic Activities in Human Atrial Fibroblasts through Inhibition of Phospholipase C Signaling and Suppression of Store-Operated Calcium Entry

  • Biomedicines. 2022 Jul 1;10(7):1574. doi: 10.3390/biomedicines10071574.
Yuan Hung  1  2 ,  Cheng-Chih Chung  3  4 ,  Yao-Chang Chen  5 ,  Yu-Hsun Kao  6  7 ,  Wei-Shiang Lin  1  2 ,  Shih-Ann Chen  8  9 ,  Yi-Jen Chen  3  4  6
Affiliations
  • 1. Graduate Institute of Medical Sciences, National Defense Medical Center, Taipei 11490, Taiwan.
  • 2. Division of Cardiology, Department of Internal Medicine, Tri-Service General Hospital, National Defense Medical Center, Taipei 11490, Taiwan.
  • 3. Division of Cardiology, Department of Internal Medicine, School of Medicine, College of Medicine, Taipei Medical University, Taipei 11031, Taiwan.
  • 4. Division of Cardiovascular Medicine, Department of Internal Medicine, Wan Fang Hospital, Taipei Medical University, Taipei 11696, Taiwan.
  • 5. Department of Biomedical Engineering, National Defense Medical Center, Taipei 11490, Taiwan.
  • 6. Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei 11031, Taiwan.
  • 7. Department of Medical Education and Research, Wan Fang Hospital, Taipei Medical University, Taipei 11696, Taiwan.
  • 8. Faculty of Medicine and Institute of Clinical Medicine, National Yang-Ming Chiao Tung University, Taipei 112304, Taiwan.
  • 9. Cardiovascular Center, Taichung Veterans General Hospital, Taichung 40705, Taiwan.
PMID: 35884879 DOI: 10.3390/biomedicines10071574
Abstract

Background: Atrial fibroblasts activation causes atrial fibrosis, which is one major pathophysiological contributor to atrial fibrillation (AF) genesis. Klotho is a pleiotropic protein with remarkable cardiovascular effects, including anti-inflammatory, anti-oxidative, and anti-apoptotic effects. This study investigated whether Klotho can modulate the activity of human atrial fibroblasts and provides an anti-fibrotic effect.

Methods: Cell migration assay and proliferation assay were used to investigate fibrogenesis activities in single human atrial fibroblasts with or without treatment of Klotho (10 and 100 pM, 48 h). Calcium fluorescence imaging, the whole-cell patch-clamp, and Western blotting were performed in human atrial fibroblasts treated with and without Klotho (100 pM, 48 h) to evaluate the store-operated calcium entry (SOCE), transient receptor potential (TRP) currents, and downstream signaling.

Results: High dose of Klotho (100 pM, 48 h) significantly reduced the migration of human atrial fibroblasts without alternating their proliferation; in addition, treatment of Klotho (100 pM, 48 h) also decreased SOCE and TRP currents. In the presence of BI-749327 (a selective canonical TRP 6 channel inhibitor, 1 μM, 48 h), Klotho (100 pM, 48 h) could not inhibit fibroblast migration nor suppress the TRP currents. Klotho-treated fibroblasts (100 pM, 48 h) had lower phosphorylated Phospholipase C (PLC) (p-PLCβ3 Ser537) expression than the control. The PLC inhibitor, U73122 (1 μM, 48 h), reduced the migration, decreased SOCE and TRP currents, and lowered p-PLCβ3 in atrial fibroblasts, similar to Klotho. In the presence of the U73122 (1 μM, 48 h), Klotho (100 pM, 48 h) could not further modulate the migration and Collagen synthesis nor suppress the TRP currents in human atrial fibroblasts.

Conclusions: Klotho inhibited pro-fibrotic activities and SOCE by inhibiting the PLC signaling and suppressing the TRP currents, which may provide a novel insight into atrial fibrosis and arrhythmogenesis.

Keywords
Klotho; atrial fibrillation; fibroblast; transient receptor potential channels.
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