Cigarette Smoke Impairs A2A Adenosine Receptor Mediated Wound Repair through Up-regulation of Duox-1 Expression

Zhi Tian,Kusum Kharbanda,Jendayi Dixon,Todd A Wyatt,Nicole Traphagen,Narasaiah Kolliputi,Diane S Allen-Gipson,Hui Zhang,Samantha Simet Chadwick

doi:10.1038/srep44405

Abstract

Cigarette smoke (CS) exposure and intrinsic factors such as the NADPH oxidases produce high levels of reactive oxygen species (ROS), ensuing inflammatory tissue injury. We previously demonstrated that CS-generated ROS, particularly hydrogen peroxide (H2O2), impaired adenosine stimulated wound repair. We hypothesized that CS exposure modulates expression of Dual oxidase 1 (Duox-1), a NADPH oxidases known to generate H2O2. To test this hypothesis, we used human bronchial epithelial cell line Nuli-1 and C57BL/6 mice. Cells were treated with 5% CS extract (CSE) for various periods of time, and mice were exposed to whole body CS for six weeks. Both CSE and CS treatment induced increased expression of Duox-1, and silencing of Doux-1 improved the rate of cell wound repair induced by CSE treatment. Nuli-1 cells pretreated with thapsigargin but not calcium ionophore exhibited increased Duox-1 mRNA expression. CSE treatment stimulated PKCα activation, which was effectively blocked by pretreatment with diphenylene iodonium, a NADPH oxidase inhibitor. Compared to control, lungs from CS-exposed mice showed a significant increase in PKCα activity and Duox-1 expression. Collectively, the data demonstrated that CS exposure upregulates expression of Duox-1 protein. This further leads to H2O2 production and PKCα activation, inhibiting A2AAR-stimulated wound repair.

Highlights

We previously demonstrated that Cigarette smoke (CS)-generated reactive oxygen species (ROS), H2O2, is implicated in blunting ADO-mediated wound repair of airway epithelial cells[1,2]
We recently demonstrated that CS exposure blunts ADO-mediated activation of cAMP-dependent Protein Kinase A (PKA) and this was facilitated by the robust activation of protein kinase C (PKC) signaling[2]
Pre-exposure of A2B Adenosine Receptor (A2BAR) knock-out Nuli-1 cells stimulated with CGS21680 to CS extract (CSE) or H2O2, decreased transepithelial resistance (TEER) compared to non-pre-exposed cells implying that A2AAR but not A2BAR is the target of inhibitory effect

Summary

Introduction

We previously demonstrated that CS-generated ROS, H2O2, is implicated in blunting ADO-mediated wound repair of airway epithelial cells[1,2]. The objective of this study was to determine whether CS exposure of airway epithelial cells activates signaling pathways associated the Duox-1 and 2 generation of H2O2. We recently demonstrated that CS exposure blunts ADO-mediated activation of cAMP-dependent Protein Kinase A (PKA) and this was facilitated by the robust activation of PKC signaling[2]. Duox-1 and/or 2, and the subsequent generation of H2O2 activates PKC, which modulates ADO’s airway epithelial cell repair and recovery

Objectives

Methods

Results

Conclusion