Modulation of epithelial sodium channel in human alveolar epithelial cells by lipoxin A4 through AhR-cAMP-dependent pathway

Bihuan Cheng ,Sheng-Rong Zhang,Liuzhan Pan ,Benji Wang ,Binyu Ying ,Guoliang Lin ,Shifang Ding

doi:10.4314/tjpr.v16i8.5

Abstract

Purpose: To investigate the effect of lipoxin A4 (LXA4) on the expressions of protein and mRNA of alveolar epithelial sodium channel (ENaC) in normal and lipopolysaccharide (LPS)-stimulated A549 cells. Methods: A549 cell-lines were randomized into 11 groups ( N = 8) and treated. EnaC level was evaluated by Western blot. Total RNA was extracted and reverse-transcribed and then levels of ENaC mRNA, cGMP and cAMP in the cells were determined. Results: LXA4 (10 -7 mol/L) increased the expressions of α-subunit of ENaC relative to LPS group. In addition, LXA4 significantly up-regulated the expression of mRNAs of α, β and γ subunits of ENaC ( p < 0.01). The level of cAMP was increased in LXA4 group, but significantly reduced in LPS group relative to control group ( p < 0.05). However, treatment with LXA4 annulled the increased cAMP concentration, compared with LPS group ( p < 0.05) Conclusion: These results show that LXA4 influences ENaC up-regulation in normal and LPS stimulated A549 alveolar epithelial cells. Keywords: Acute lung injury, Alveolar epithelial sodium channel, Lipoxin A4, AhR, cAMP, cGMP

Highlights

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) lead to respiratory failure and pulmonary edema [1]
Dose-dependency of lipoxin A4 (LXA4) regulated ENaC expression in A549 cells stimulated with LPS
In subsequent experiments, the expression of ENaC in A549 cells treated with LPS was assessed using 10-7 M LXA4

Summary

INTRODUCTION

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS) lead to respiratory failure and pulmonary edema [1]. Earlier studies showed that alveolar epithelial sodium channel (ENaC) and Na-K-ATPase play critical roles in reducing edema in ALI/ARDS [3,4]. The present study aims to test the hypothesis that LXA4 increases ENaC protein and mRNA expressions in normal and LPS-stimulated A549 cells. It investigated LXA4 receptors, ALX, AhR, cysLRT1 and cysLRT2 to determine the key receptors involved in up-regulation of ENaC function in A549 cells, as well as the levels of cAMP and cGMP in A549 cells. Levels of ENaC mRNA in A549 cells were determined with SYBR Prime Script Kit on the CFX96 Real-Time PCR Detection System (BioRad, Hercules, CA, USA). A probability of p < 0.05 was considered significant

RESULTS

DISCUSSION

Conflict of Interest

CONCLUSION