Heme oxygenase-1 protects corexit 9500A-induced respiratory epithelial injury across species.

Fu Jun Li,R Douglas Watson,Subhashini Bolisetty,Ranu Surolia,Ryan N Duggal,Stephen Watts,Hitesh Batra,Suman Karki,Veena B Antony,Mickie Powell,Anupam Agarwal,Octavio M Oliva,Tejaswini Kulkarni,Zheng Wang,Victor J Thannickal,Ferenc Gallyas

doi:10.1371/journal.pone.0122275

Abstract

The effects of Corexit 9500A (CE) on respiratory epithelial surfaces of terrestrial mammals and marine animals are largely unknown. This study investigated the role of CE-induced heme oxygenase-1 (HO-1), a cytoprotective enzyme with anti-apoptotic and antioxidant activity, in human bronchial airway epithelium and the gills of exposed aquatic animals. We evaluated CE-mediated alterations in human airway epithelial cells, mice lungs and gills from zebrafish and blue crabs. Our results demonstrated that CE induced an increase in gill epithelial edema and human epithelial monolayer permeability, suggesting an acute injury caused by CE exposure. CE induced the expression of HO-1 as well as C-reactive protein (CRP) and NADPH oxidase 4 (NOX4), which are associated with ROS production. Importantly, CE induced caspase-3 activation and subsequent apoptosis of epithelial cells. The expression of the intercellular junctional proteins, such as tight junction proteins occludin, zonula occludens (ZO-1), ZO-2 and adherens junctional proteins E-cadherin and Focal Adhesion Kinase (FAK), were remarkably inhibited by CE, suggesting that these proteins are involved in CE-induced increased permeability and subsequent apoptosis. The cytoskeletal protein F-actin was also disrupted by CE. Treatment with carbon monoxide releasing molecule-2 (CORM-2) significantly inhibited CE-induced ROS production, while the addition of HO-1 inhibitor, significantly increased CE-induced ROS production and apoptosis, suggesting a protective role of HO-1 or its reaction product, CO, in CE-induced apoptosis. Using HO-1 knockout mice, we further demonstrated that HO-1 protected against CE-induced inflammation and cellular apoptosis and corrected CE-mediated inhibition of E-cadherin and FAK. These observations suggest that CE activates CRP and NOX4-mediated ROS production, alters permeability by inhibition of junctional proteins, and leads to caspase-3 dependent apoptosis of epithelial cells, while HO-1 and its reaction products protect against oxidative stress and apoptosis.

Highlights

The Deepwater Horizon oil spill following a well head blowout emitted 205.8 million gallons of crude oil before getting capped three months later [1]
We addressed these two responses in epithelial cells exposed to Corexit 9500A (CE) in vitro and in vivo
We showed that CE-induced apoptosis is caspase-3 dependent and that reactive oxygen species (ROS) play an important role in this pathway (Fig. 9)

Summary

Introduction

The Deepwater Horizon oil spill following a well head blowout emitted 205.8 million gallons of crude oil before getting capped three months later [1]. A total of 1.84 million gallons of the dispersant was sprayed on the surface and released subsea [3,4]. A study carried out at Louisiana State University found that the 50%-lethal-concentration (LC50) of Louisiana Sweet Crude oil in killifish was decreased more than eleven times when dispersed by CE [5], a more recent report from the Georgia Institute of Technology and Universidad Autonoma de Aguascalientes (UAA) showed that mixing the dispersant with oil increased the toxicity of the mixture up to 52 times when compared with oil alone [6]. Despite the large volume of CE used in remediation, the effects on the respiratory epithelium of humans and gills of aquatic animals such as fish and crabs exposed to this dispersant are largely unknown

Methods

Results

Conclusion