Kidney & Blood Pressure Research (Mar 2022)

Requirement of Na+/H+ exchanger NHE1 for vasopressin-induced osteogenic signaling and calcification in human aortic smooth muscle cells

  • Xuexue Zhu,
  • Ke Ma,
  • Kuo Zhou,
  • Xia Pan,
  • Jibin Liu,
  • Bernd Nürnberg,
  • Ioana Alesutan,
  • Jakob Völkl,
  • Florian Lang

DOI
https://doi.org/10.1159/000524050

Abstract

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Background/Aims: Vasopressin is a powerful stimulator of vascular calcification, augmenting osteogenic signaling in vascular smooth muscle cells (VSMCs) including up-regulation of the transcription factors core-binding factor α-1 (CBFA1), msh homeobox 2 (MSX2), and SRY-Box 9 (SOX9), as well as of tissue-nonspecific alkaline phosphatase (ALPL). Vasopressin-induced osteogenic signaling and calcification require the serum- & glucocorticoid-inducible kinase 1 (SGK1). Known effects of SGK1 include up-regulation of Na+/H+ exchanger 1 (NHE1). NHE1 further participates in the regulation of reactive oxygen species (ROS). NHE1 has been shown to participate in the orchestration of bone mineralization. The present study, thus, explored whether vasopressin modifies NHE1 expression and ROS generation, as well as whether pharmacological inhibition of NHE1 disrupts vasopressin-induced osteogenic signaling and calcification in VSMCs. Methods: Human aortic smooth muscle cells (HAoSMCs) were treated with vasopressin in the absence or presence of SGK1 siRNA, SGK1 inhibitor GSK-650394, and NHE1 blocker cariporide. Transcript levels were determined by using qRT-PCR, protein abundance by western blotting, ROS generation with 2’,7’-dichlorofluorescein diacetate (DCFDA) fluorescence, as well as ALP activity and calcium content by using colorimetric assays. Results: Vasopressin significantly enhanced the NHE1 transcript and protein levels in HAoSMCs, effects significantly blunted by SGK1 inhibition with GSK-650394 or SGK1 siRNA. Vasopressin increased ROS accumulation, an effect significantly blocked by NHE1 inhibitor cariporide. Vasopressin further significantly increased osteogenic markers CBFA1, MSX2, SOX9, and ALPL transcript levels, as well as ALP activity and calcium content in HAoSMCs, all effects significantly blunted by SGK1 silencing or in the presence of GSK-650394 and cariporide. Conclusion: Vasopressin stimulates NHE1 expression and ROS generation, an effect dependent on SGK1 and required for vasopressin-induced stimulation of osteogenic signaling and calcification of VSMCs.