Inducible lung epithelial resistance requires multisource reactive oxygen species generation to protect against bacterial infections.

Hayden H Ware,Carson T Kirkpatrick,Jezreel Pantaleón García,Sarah Syed,Alexander D Kontoyiannis,Shradha Wali,James M Tour,Scott E Evans,William K A Sikkema,Miguel Leiva Juarez,Vikram V Kulkarni,Yongxing Wang,Samithamby Jeyaseelan

doi:10.1371/journal.pone.0208216

Abstract

Pneumonia remains a global health threat, in part due to expanding categories of susceptible individuals and increasing prevalence of antibiotic resistant pathogens. However, therapeutic stimulation of the lungs’ mucosal defenses by inhaled exposure to a synergistic combination of Toll-like receptor (TLR) agonists known as Pam2-ODN promotes mouse survival of pneumonia caused by a wide array of pathogens. This inducible resistance to pneumonia relies on intact lung epithelial TLR signaling, and inducible protection against viral pathogens has recently been shown to require increased production of epithelial reactive oxygen species (ROS) from multiple epithelial ROS generators. To determine whether similar mechanisms contribute to inducible antibacterial responses, the current work investigates the role of ROS in therapeutically-stimulated protection against Pseudomonas aerugnosa challenges. Inhaled Pam2-ODN treatment one day before infection prevented hemorrhagic lung cytotoxicity and mouse death in a manner that correlated with reduction in bacterial burden. The bacterial killing effect of Pam2-ODN was recapitulated in isolated mouse and human lung epithelial cells, and the protection correlated with inducible epithelial generation of ROS. Scavenging or targeted blockade of ROS production from either dual oxidase or mitochondrial sources resulted in near complete loss of Pam2-ODN-induced bacterial killing, whereas deficiency of induced antimicrobial peptides had little effect. These findings support a central role for multisource epithelial ROS in inducible resistance against a bacterial pathogen and provide mechanistic insights into means to protect vulnerable patients against lethal infections.

Highlights

Lower respiratory tract infections remain the leading cause of premature death and disability among both otherwise healthy and immunosuppressed people worldwide [1,2,3,4,5]
We have previously reported that a single nebulized treatment with Pam2-ODN results in improved survival of otherwise lethal pneumonias, including those caused by P. aeruginosa [16, 17, 20, 21]
We found that the protection afforded by Pam2-ODN (Fig 1A) is associated with reduced bacterial burden immediately after infection, whether assessing burden by serial dilution culture of whole lung homogenates or culture of bronchoalveolar lavage (BAL) fluid (Fig 1B), suggesting that a Pam2-ODN-induced antimicrobial environment existed at the time of infection

Summary

Introduction

Lower respiratory tract infections remain the leading cause of premature death and disability among both otherwise healthy and immunosuppressed people worldwide [1,2,3,4,5]. In an effort to address the persisting threat of pneumonia to vulnerable populations, our laboratory has developed a program focused on manipulating the intrinsic antimicrobial capacity of the host to prevent pneumonia in susceptible populations in order to avert these catastrophic outcomes. Based on this program, we have reported that the mucosal defenses of the lungs can be stimulated to protect mice against a wide array of otherwise lethal pneumonias, including those caused by antibiotic-resistant bacteria [16,17,18,19].

Methods

Results

Conclusion