Medical Student Research Fellowship for Summer 2008

Mentor: Danielle Robertson, O.D., Ph.D. & Dwight Cavanagh, M.D., Ph.D.
Department: Ophthalmology
Room number: E7.234
Mail Code: 9057
Phone number: 214-648-3416
E-mail: Danielle.robertson@utsouthwestern.edu

Project 1

Project title: Inflammation-induced lipid raft expression

Human subjects IRB approved project number (where applicable):

Animal subjects IRB approved project number (where applicable):0550-07-03-1

Project Type (patient-based research, animal-based research, or basic research; this characterization is only to permit a general classification for grouping similar types of projects)
Animal-based

Brief Description of Project:
Microbial keratitis is a potentially blinding disease and remains the most significant complication of contact lens wear. Pseudomonas aeruginosa (PA), a pathogenic gram-negative organism, has been identified as the leading causative agent in contact lens-related microbial keratitis. Recently, PA has been shown to invade corneal epithelial cells following contact lens wear through cholesterol enriched domains in the plasma membrane known as lipid rafts. Previous studies evaluating the pathogenesis of MK have shown that IFN gamma is upregulated in MK. In addition, IFN gamma has been implicated in the formation of lipid rafts in intestinal epithelia. The purpose of this project is to establish a potential role for IFN gamma in lipid raft formation in the corneal epithelium. In the first part of this project, IFN gamma upregulation in the tear film following contact lens wear will be examined. To accomplish this, New Zealand white rabbits, the standard animal model for contact lens corneal studies, will be used. Tear samples from contact lens wearing rabbits will be collected immediately upon lens removal and tested for IFN gamma using an ELISA. In the second part of this project, the effect of IFN gamma on lipid raft formation will be examined in vitro using a telomerase-immortalized human corneal epithelial cell line (hTCEpi). Raft formation will be detected by beta cholera toxin labeling and visualized using the laser scanning confocal microscope. The results of this study will provide valuable insight into an inflammatory regulated mechanism of microbial keratitis in contact lens wearers.

Project 2

Project title: Pseudomonas Biofilm Formation and Microbial Infection in the Rabbit Cornea

Human subjects IRB approved project number (where applicable):

Animal subjects IRB approved project number (where applicable): 2008-0012, pending

Project Type (patient-based research, animal-based research, or basic research; this characterization is only to permit a general classification for grouping similar types of projects)
Animal-based

Brief Description of Project:
Pseudomonas aeruginosa (PA) is the primary causative agent in contact lens- related microbial infections of the cornea. Repeated exposure to PA can lead to biofilm development, conferring a protective advantage to PA from antibiotics as well as the immune system, thereby placing the patient at an increased risk for a potentially blinding infection. In cystic fibrosis patients, it has been shown that continued neutrophil accumulation in the airway leads to an enhancement in biofilm formation, which appears to be regulated by the formation of surface polymers consisting of DNA and actin released from necrotic neutrophils. The focus of this project is to evaluate the use of an investigational compound that blocks neutrophil enhanced biofilm formation as a potential therapeutic treatment to eliminate risk of PA infection in the rabbit cornea. We have recently shown that the formation of lipid rafts mediates PA invasion in corneal epithelial cells. For all experiments, contact lens wearing rabbits will be exposed to PA with and without the use of the investigational compound. The endpoints for this study, raft formation and PA internalization, will be assessed by direct visualization using the laser confocal microscope. The results of this project could potentially lead to the development of a new therapeutic modality that can be used to prevent and treat corneal infection.