MiR-203a-3p regulates TGF-β1-induced epithelial-mesenchymal transition (EMT) in asthma by regulating Smad3 pathway through SIX1.

Qi Fan,Yu Jian

doi:10.1042/bsr20192645

Abstract

Asthma is a common chronic airway disease with increasing prevalence. MicroRNAs act as vital regulators in cell progressions and have been identified to play crucial roles in asthma. The objective of the present study is to clarify the molecular mechanism of miR-203a-3p in the development of asthma. The expression of miR-203a-3p and Sine oculis homeobox homolog 1 (SIX1) were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The protein levels of SIX1, fibronectin, E-cadherin, vimentin, phosphorylated-drosophila mothers against decapentaplegic 3 (p-Smad3) and Smad3 were measured by Western blot. The interaction between miR-203a-3p and SIX1 was confirmed by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay. MiR-203a-3p was down-regulated and SIX1 was up-regulated in asthma serums, respectively. Transforming growth factor-β1 (TGF-β1) treatment induced the reduction of miR-203a-3p and the enhancement of SIX1 in BEAS-2B and 16HBE cells in a time-dependent manner. Subsequently, functional experiments showed the promotion of epithelial–mesenchymal transition (EMT) induced by TGF-β1 treatment could be reversed by miR-203a-3p re-expression or SIX1 deletion in BEAS-2B and 16HBE cells. SIX1 was identified as a target of miR-203a-3p and negatively regulated by miR-203a-3p. Then rescue assay indicated that overexpressed miR-203a-3p ameliorated TGF-β1 induced EMT by regulating SIX1 in BEAS-2B and 16HBE cells. Moreover, miR-203a-3p/SIX1 axis regulated TGF-β1 mediated EMT process in bronchial epithelial cells through phosphorylating Smad3. These results demonstrated that MiR-203a-3p modulated TGF-β1-induced EMT in asthma by regulating Smad3 pathway through targeting SIX1.

Highlights

Asthma, as a common chronic airway disease, affects approximately 300 million people in the world [1]
Western blot analysis determined that Sine oculis homeobox homolog 1 (SIX1) level was significantly enhanced by Transforming growth factor-β1 (TGF-β1) treatment in BEAS-2B and 16HBE cells in a time-dependent manner (Figure 2B)
The epithelial–mesenchymal transition (EMT)-related proteins were measured and we found that the level of fibronectin and vimentin were up-regulated by TGF-β1 in BEAS-2B (P < 0.0001, P < 0.0001) and 16HBE (P < 0.0001, P < 0.0001) cells, while these up-regulation were sharply inhibited by miR-203a-3p re-expression (P < 0.0001, P < 0.0001 in BEAS-2B cells and P < 0.0001, P < 0.0001 in 16HBE cells)

Summary

Introduction

As a common chronic airway disease, affects approximately 300 million people in the world [1]. Asthma is still a major problem to public health, and the prevalence of asthma is increasing in developing countries and undeveloped countries because of late diagnosis and poor treatment [2]. Asthma patients often represent chronic airway inflammation and remodeling reactions [8]. Up to now there are no feasible methods to restore airway remodeling. The main treatment for asthma is inhaling corticosteroids. A considerable number of patients are still incurable despite inhalation of high concentrations of corticosteroids [9]. A better understanding of the molecular mechanism in asthma is essential for the treatment and prevention of asthma

Objectives

Methods

Results

Conclusion