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  3. Reduced intracellular chloride concentration impairs angiogenesis by inhibiting oxidative stress-mediated VEGFR2 activation

Reduced intracellular chloride concentration impairs angiogenesis by inhibiting oxidative stress-mediated VEGFR2 activation

  • Acta Pharmacol Sin. 2021 Apr;42(4):560-572. doi: 10.1038/s41401-020-0458-7.
Kai Li  # 1,  Ying-Ying Liu  # 1,  Xiao-Fei Lv  # 1,  Zhuo-Miao Lin 1,  Ting-Ting Zhang 1,  Fei-Ran Zhang 1,  Jia-Wei Guo 1,  Yu Hong 1,  Xiu Liu 1,  Xiao-Chun Lin 1,  Jia-Guo Zhou 2 1 3 4 5 6,  Qian-Qian Wu 7 8 9,  Si-Jia Liang 10 11,  Jin-Yan Shang 12 13
Affiliations

Affiliations

  • 1 Department of Pharmacology, Cardiac and Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510080, China.
  • 2 Program of Kidney and Cardiovascular Disease, The Fifth Affiliated Hospital, Sun Yat-Sen University, Zhuhai, 519000, China.
  • 3 Guangdong Province Key Laboratory of Brain Function and Disease, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510080, China.
  • 4 Department of Cardiology, Sun Yat-sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, 510080, China.
  • 5 Department of Physiology, Key Laboratory of Cardiovascular disease, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou, 511436, China.
  • 6 Guangzhou Institute of Cardiovascular Disease, The Second Affiliated Hospital, Guangzhou Medical University, Guangzhou, 511436, China.
  • 7 Key Laboratory of Metabolic Cardiovascular Diseases Research of National Health Commission, Ningxia Medical University, Yinchuan, 750004, China.
  • 8 Ningxia Key Laboratory of Vascular Injury and Repair Research, Ningxia Medical University, Yinchuan, 750004, China.
  • 9 School of Basic Medical Sciences, Ningxia Medical University, Yinchuan, 750004, China.
  • 10 Program of Kidney and Cardiovascular Disease, The Fifth Affiliated Hospital, Sun Yat-Sen University, Zhuhai, 519000, China. [email protected].
  • 11 Department of Pharmacology, Cardiac and Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510080, China. [email protected].
  • 12 Program of Kidney and Cardiovascular Disease, The Fifth Affiliated Hospital, Sun Yat-Sen University, Zhuhai, 519000, China. [email protected].
  • 13 Department of Pharmacology, Cardiac and Cerebral Vascular Research Center, Zhongshan School of Medicine, Sun Yat-Sen University, Guangzhou, 510080, China. [email protected].
  • # Contributed equally.
PMID: 32694758 DOI: 10.1038/s41401-020-0458-7
Abstract

Chloride (Cl-) homeostasis is of great significance in cardiovascular system. Serum Cl- level is inversely associated with the mortality of patients with heart failure. Considering the importance of angiogenesis in the progress of heart failure, this study aims to investigate whether and how reduced intracellular Cl- concentration ([Cl-]i) affects angiogenesis. Human umbilical endothelial cells (HUVECs) were treated with normal Cl- medium or low Cl- medium. We showed that reduction of [Cl-]i (from 33.2 to 16.18 mM) inhibited HUVEC proliferation, migration, Cytoskeleton reorganization, tube formation, and subsequently suppressed angiogenesis under basal condition, and VEGF stimulation or hypoxia treatment. Moreover, VEGF-induced NADPH-mediated Reactive Oxygen Species (ROS) generation and VEGFR2/KDR/Flk-1 axis activation were markedly attenuated in low Cl- medium. We revealed that lowering [Cl-]i inhibited the expression of the membrane-bound catalytic subunits of NADPH, i.e., p22phox and Nox2, and blunted the translocation of cytosolic regulatory subunits p47phox and p67phox, thereby restricting NADPH Oxidase complex formation and activation. Furthermore, reduced [Cl-]i enhanced ROS-associated protein tyrosine Phosphatase 1B (PTP1B) activity and increased the interaction of VEGFR2/KDR/Flk-1 and PTP1B. Pharmacological inhibition of PTP1B reversed the effect of lowering [Cl-]i on VEGFR2/KDR/Flk-1 phosphorylation and angiogenesis. In mouse hind limb ischemia model, blockade of Cl- efflux using Cl- channel inhibitors DIDS or DCPIB (10 mg/kg, i.m., every other day for 2 weeks) significantly enhanced blood flow recovery and new capillaries formation. In conclusion, decrease of [Cl-]i suppresses angiogenesis via inhibiting oxidase stress-mediated VEGFR2/KDR/Flk-1 signaling activation by preventing NADPH Oxidase complex formation and promoting VEGFR2/KDR/Flk-1/PTP1B association, suggesting that modulation of [Cl-]i may be a novel therapeutic avenue for the treatment of angiogenic dysfunction-associated diseases.

Keywords

PTP1B; PTP1B inhibitor II; VEGFR2; angiogenesis; chloride; oxidative stress.

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