Increased expression of microRNA‐29b attenuates function of Ca2+‐activated K+ channels in human PASMC from idiopathic PAH patients
Authors: Babicheva Aleksandra , Ayon Ramon Jose , Makino Ayako , Yuan Jason X.‐J. 
Published: June 2021

Introduction

Pulmonary artery smooth muscle cells (PASMC) isolated from patients with idiopathic pulmonary arterial hypertension (IPAH) exhibit a more depolarized membrane potential in comparison to PASMC from failed donor lung tissue. The development of sustained pulmonary vasoconstriction and vascular remodeling, depend, in part on the depolarized membrane potential. Changes in activity and/or expression of large‐conductance Ca2+‐activated K+ (BKCa) channels have a direct impact on the resting membrane potential of PASMC. Inhibition of BKCa channels is involved in the pathophysiology of IPAH by causing membrane depolarization and vasoconstriction due to elevated [K+]i that promotes enhanced Ca2+ influx through voltage‐gated Ca2+ channels. However, the underlying mechanisms responsible for downregulation of BKCa channels in IPAH‐PASMC remains unknown. MicroRNAs (miRNAs) serve as important post‐transcriptional regulators of gene expression by inhibiting translation and stability of mRNAs. We hypothesized that upregulation of miRNA‐29b inhibits BKCa channels in PASMC isolated from IPAH patients.

Methods

PASMC from healthy subjects and patients with IPAH were used for this study. K+ currents, recorded using the conventional whole‐cell configuration of the patch clamp technique, were elicited by a series of depolarizing steps from a holding potential of −80 mV for 500 ms. The cell‐attached configuration of the patch clamp technique was used to measure single‐channel currents from BKCa channels. The activity of BKCa channels was also determined in normal PASMC overexpressed miRNA‐29b, and IPAH‐PASMC transfected with miRNA‐29b inhibitor. mRNA and protein of BKCa channels were quantified by qPCR and Western blot, respectively.

Results

Upregulated miRNA‐29b correlated with the decreased activity of BKCa channel in PASMC from IPAH patients. Expression of the pore‐forming subunit α (KCNMA1) is higher (7.9845±1.49, n=4, P<0.001) while that of the auxiliary subunit β1 (KCNMB1) is lower (0.1175±0.05, n=4, P<0.001) in IPAH‐PASMC compared to normal cells. Overexpression of miRNA‐29b in normal PASMC significantly attenuates BKCa channel current density (15.01±1.76 pA/pF, n=10) when compared to controls (100.18±8.53 pA/pF, n=10) at +80mV and downregulates KCNMB1. Inhibition of miRNA‐29b rescued BKCa currents (127.16±18.50 pA/pF, n=5) in IPAH‐PASMCs compared to negative controls (5.81±0.68 pA/pF, n=3) at +80mV and upregulates KCNMB1.

Conclusion

Upregulated miRNA‐29b results in posttranscriptional downregulation of KCNMB1 which leads to attenuation of BKCa channel activity in IPAH‐PASMC.

Support or Funding Information

This work was supported in parts by grants from the National Heart, Lung, and Blood Institute of the National Institutes of Health (HL125208, HL126609, and HL135807).

This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.