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Article
In Vitro Interaction of Pseudomonas aeruginosa with Human Middle Ear Epithelial Cells
PLOS ONE
  • Rahul Mittal, University of Miami
  • M'hamed Grati, University of Miami
  • Robert Gerring, University of Miami
  • Patricia Blackwelder, University of Miami
  • Denise Yan, University of Miami
  • Jian-Dong Li, Georgia State University - Atlanta
  • Xue Zhong Liu, University of Miami
Document Type
Article
Publication Date
3-1-2014
Keywords
  • Pseudomonas aeruginosa,
  • Actins,
  • Bacterial Pathogens,
  • Host Cells,
  • Middle Ear,
  • Microfilaments,
  • Epithelial Cells,
  • Microtubules
Peer Reviewed
1
Abstract

Background: Otitis media (OM) is an inflammation of the middle ear which can be acute or chronic. Acute OM is caused by Streptococcus pneumoniae, Haemophilus influenzae, and Moraxella catarrhalis whereas Pseudomonas aeruginosa is a leading cause of chronic suppurative otitis media (CSOM). CSOM is a chronic inflammatory disorder of the middle ear characterized by infection and discharge. The survivors often suffer from hearing loss and neurological sequelae. However, no information is available regarding the interaction of P. aeruginosa with human middle ear epithelial cells (HMEECs).

Methodology and Findings: In the present investigation, we demonstrate that P. aeruginosa is able to enter and survive inside HMEECs via an uptake mechanism that is dependent on microtubule and actin microfilaments. The actin microfilament disrupting agent as well as microtubule inhibitors exhibited significant decrease in invasion of HMEECs by P. aeruginosa. Confocal microscopy demonstrated F-actin condensation associated with bacterial entry. This recruitment of Factin was transient and returned to normal distribution after bacterial internalization. Scanning electron microscopy demonstrated the presence of bacteria on the surface of HMEECs, and transmission electron microscopy confirmed the internalization of P. aeruginosa located in the plasma membrane-bound vacuoles. We observed a significant decrease in cell invasion of OprF mutant compared to the wild-type strain. P. aeruginosa induced cytotoxicity, as demonstrated by the determination of lactate dehydrogenase levels in culture supernatants of infected HMEECs and by a fluorescent dye-based assay. Interestingly, OprF mutant showed little cell damage compared to wild-type P. aeruginosa.

Conclusions and Significance: This study deciphered the key events in the interaction of P. aeruginosa with HMEECs in vitro and highlighted the role of bacterial outer membrane protein, OprF, in this process. Understanding the molecular mechanisms in the pathogenesis of CSOM will help in identifying novel targets to design effective therapeutic strategies and to prevent hearing loss.

Comments

e91885

DOI
10.1371/journal.pone.0091885
Citation Information
Rahul Mittal, M'hamed Grati, Robert Gerring, Patricia Blackwelder, et al.. "In Vitro Interaction of Pseudomonas aeruginosa with Human Middle Ear Epithelial Cells" PLOS ONE Vol. 9 Iss. 3 (2014) p. 1 - 11 ISSN: 1932-6203
Available at: http://works.bepress.com/patricia-blackwelder/18/