Training undergraduate and graduate students is the cornerstone of my research program at UTT. Students are exposed to a multidisciplinary research environment utilizing the principals of molecular biology, biochemistry, pharmacology, microbiology, analytical techniques, and a nanoparticle drug delivery system. The theoretical knowledge, critical thinking, and practical skills gained by students may encourage them to consider careers in biomedical research, medicine, and allied health careers. In fact, three of my previous UTT students, Aby Green, Brandon Beddingfield, and Adam Canion, started their postgraduate education at the University of Washington (Microbiology Dept), Tulane (Immunology), and UTMBG medical school in the Fall 2010. While others assumed positions in local food industry (Brookshire’s) and research associate positions in Biomedical Research Department at UTHSCT. Currently, 5 pre-med undergraduate and 3 graduate students are conducting research in my lab at UTT.
My research projects have been supported by such funding agencies as National Institute of Health, the American Heart Association, and the American Lung Association. Our findings, populated in 37 peer-reviewed articles and 5 book chapters, have provided new information about how Pseudomonas infections injure the lungs and introduced novel therapeutics for patients with severe lung injuries due to bacterial infections and inflammation.
To accelerate my research efforts, I collaborate and published with the following scientists and physicians:
Drs. Subramanian and Mahdavi, UTT
Drs. R. Amaro, A. Komissarov, G. Florova, and S. Idell; UTHSCT, Tyler, TX
Dr. Mantell, St. John's University College of Pharmacy, Queens, NY.
Dr. Ed Miller, North Shore-LIJ Health System, Manhasset, NY
Dr. R. Hancock, The university of British Colombia, CA
Dr. A. Omri, Laurentian University, Ontario, Canada.
Current Research Students Projects at UTT
I) Pathogenesis of Lung Injury in Pseudomonas aeruginosa
P. aeruginosa is responsible for 12% of hospital related urinary tract infections, 8% of surgical wound infections, 12% of blood infections, and 14% of pneumonias. Pseudomonas-related infections are serious diseases that affect children as well as elders due to their immature or weaken immune systems. In addition, almost all the children with cystic fibrosis carry this organism in their lungs by their 10th birthday. The recurrent Pseudomonas infections, in turn, worsen their pulmonary functions. Infections with this bacterium are the cause of death in over three-quarters of children with CF. We would like to determine the molecular mechanisms by which Pseudomonas causes infection and inflammation.
Omar Castillo (Graduate Student)
Role of Pseudomonas aeruginosa elastase and its role in the disruption of epithelial tight junctions.
The pathogenesis of Pseudomonas is the result of numerous pathogen-host interactions. I am investigating cell signaling pathways and their role in tight junction disruption. Our lab has previously shown that Pseudomonas elastase (PE) activates the epidermal growth factor receptor (EGFR). Activation of this receptor leads to numerous intracellular signaling activity, including activation of the mitogen-activated pathway kinase (MAPK). The aim of this project is to investigate the mechanism of PE-induced activation of the MAPK pathway in human lung epithelia and the role of the EGFR-Ras-Raf-MEK-ERK1/2 pathway in the infection process. The expectation of this project is to decrease the risk of P. aeruginosa infection by inhibition of bacterial secreted proteins or by blocking cell signaling receptors.
Kourteny Neal (Graduate Student)
Cytokine gene and protein expression in human lung fibroblast exposed to Pseudomonas aeruginosa elastase
For my Master’s Thesis research project I am investigating cell signaling mechanisms of inflammation in the human lung fibroblast. The organism of interest is Pseudomonas aeruginosa (PA) and particularly one of its virulence factors known as elastase (PE). I am seeking to understand how PE affects signaling cascades involved in proinflammatory cytokine release by the human lung fibroblast. The cytokines under investigation are IL1A, IL1B, IL2, IL4, IL6, IL8, IL10, IL12, IL17A, IFNγ, TNFα, GM-CSF. In order to thoroughly evaluate these cytokines for both protein and gene expressions, enzyme-linked immunosorbent assays (ELISA) and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), and specific signal transduction inhibitors are utilized. My research will contribute to drug discovery efforts to suppress Pseudomonas-induced inflammation.
Brandon Beddingfield (Graduate Student)
The Role of PA4514 in the Pathogenesis of Pseudomonas aeruginosa
Cystic Fibrosis is the most common genetic disease that affects about 1 in every 2,500 Caucasian births. CF is characterized by mutations in the CFTR gene that regulates electrolyte transport across cellular membranes. These mutations result in a defective chloride transport that ultimately leads to an overly viscous mucous that impedes the normal airway function. The lungs then become colonized with microorganisms, P. aeruginosa in particular, that results in decline of lung function and ultimately death.
In the CF lung, iron is more plentiful than in normal lung tissue. It has been shown that lung epithelial cells in CF release iron in significantly higher amounts than non-CF cells. Strains of P. aeruginosa deficient in genes for the production of pyoverdines make defective biofilms, showing that iron plays an important role in the pathogenesis of disease.
Exactly which iron receptors on the outer membrane of P. aeruginosa are important, and to what extent, are not known. PA4514 is homologous for the piuA gene, which encodes an outer membrane iron uptake receptor protein. My goals in this project include inactivating the PA4514 gene from PA01 through insertional inactivation and determining its importance in the pathogenesis of P. aeruginosa in vitro.
Lauren Thomas, Samamtha Steinley and Douglas Vaughan (Undergrad)
II) Nanoparticle Delivery System Project
A significant rise in multi-drug resistant strains of P. aeruginosa calls for intense research effort for developing novel antibiotics. Hence, we are investigating the safety and effectiveness of a new antibiotic delivery system that we have developed in collaboration with Dr. Omri at the Laurentian University, Canada (Patent pending). Our goal is to encapsulate anti-Pseudomonas drugs in specially formulated liposomes in order to enhance antibacterial properties of the existing drugs whilst reducing their toxicity.
Samantha Steinley (Undergrad)
Liposomal Antibiotics to Combat Multidrug Resistant P. aeruginosa
Pseudomonas aeruginosa accounts for approximately 95% of deaths in patients with cystic fibrosis. This opportunistic pathogen is responsible for nosocomial pneumonia, chronic respiratory infection in patients with cystic fibrosis, and acute pulmonary infection in immune-compromised individuals. Pseudomonas infections have long been treated with aminoglycosides. However, these drugs are not effective against certain strains of Pseudomonas that have developed resistance due to decreased membrane permeability. The prevalence of multidrug resistant strains appears to be increasing significantly with a mortality rate greater than 31%. A nanoparticle delivery system that could reduce antibiotic toxicity and provide effective therapeutic modality is of great importance and could be provided by encapsulating antibiotics currently used to treat Pseudomonas infections inside liposomes.
Liposomes are emerging as a preferred drug carrier to combat antibiotic resistant strains due to their ability to fuse with the prokaryotic cell membrane as the endpoint of their carrier function. Liposomes are artificial vesicles composed of a phospholipid bilayer with an aqueous center that contains, in this case, aminoglycosides such as tobramycin, gentamicin, and amikacin. They reduce the toxicity of the antibiotics, protect the drug from unwanted metabolic breakdown, and alter the pharmacokinetics of the active agent.
Through this project we investigate the efficacy of this alternate method in drug delivery, more specifically, we study its effects on microbial adhesion which is accepted as a prerequisite to P. aeruginosa colonization and infection. Using A549 pulmonary epithelial cells, we quantify P. aeruginosa adhesions to host cells using electron microscopy and colony forming unit (CFU) assay. Our ultimate goal is to safely eliminate bacterial infections caused by organism resistance to conventional drugs. We hypothesize that the use of liposomal antibiotics as opposed to conventional methods will augment the existing antibiotics ability to efficiently inhibit the adherence of P. aeruginosa to human lung cells and therefore decrease susceptibility to infection.
Douglas Vaughan (Undergrad)
Application of Liposomal Antibiotics to Reduce Bacterial Adherence to Lung Epithelia
There is a substantial growth in antibiotic resistant bacterial strains, which has increased the need for improved treatment options. The most promising method of antibiotic resistance being decreased outer membrane permeability. Liposomes are an effective drug carrier in antibiotic resistant strain combat due to their ability to abolish or reduce the toxicity of the encapsulated antibiotics, target the desired anatomic site, protect the drug from unwanted metabolic breakdown, and alter the pharmacokinetics of the active agent.
We have developed innovative methods of encapsulating common antibiotics in liposomes against antibiotic resistant strains. These strains contain Pseudomonas aeruginosa, which is an opportunistic, Gram-negative bacterium responsible for infections among immunocompromised individuals and is a leading cause of acute nosocomial pneumonia and chronic respiratory infection in cystic fibrosis patients.
We are investigating the efficacy of the liposomal amikacin in preventing bacterial adhesion and colonization in an in-vitro model of bacterial infection and inflammation. Using A549 pulmonary epithelial cells, we quantify P. aeruginosa adhesions to host cells using electron microscopy and colony forming unit assay. Our ultimate goal is to safely eliminate bacterial infections caused by organism resistant to conventional drugs in animal and human populations.
Anti-inflammatory Effects of Erythromycin on Human Pulmonary Epithelial Cells
Inflammation in the host is due to the production of cytokines and chemokines including Interleukin-8 (IL-8), one of the major mediators of the inflammatory response. IL-8 is a chemotaxic chemokine that induces neutrophils to leave the bloodstream and enter into the surrounding tissue. Inflammation is necessary in the body to a certain extent but too much inflammation can be harmful and even fatal. Erythromycin (EM) and its derivatives have been used to treat chronic inflammation of upper and lower respiratory tract such as diffuse panbronchiolitis (DPB), bronchial asthma, and chronic sinusitis. Although the mechanism of anti-inflammatory property of erythromycin is not fully understood, EM inhibits the production of proinflammatory cytokine such as IL-6 and IL-8 via inhibition of nuclear transcription factors.
The purpose of my project is to compare the anti-inflammatory effects of our novel liposomal formulations to free erythromycin. My hypothesis is that the liposomal form of erythromycin is superior to the free drug in terms of their anti-inflammatory effects.Open Project
Free and liposomal-entrapped erythromycin derived immuno-modulation in human alveolar cells
Pulmonary inflammation kills millions of people across the globe each year. It is estimated that chronic obstructive pulmonary disease (COPD) will be the third leading cause of death in the world by the year 2020. COPD is caused by a number of different stimuli but is always characterized in the same manner: excessive inflammation of the airways. Steroids are often prescribed for reducing pulmonary inflammation, but their efficacy has long been questions and the side effects are troubling so alternative therapies are currently in demand.
We are investigating the efficacy of both free and liposomal erythromycin in reducing inflammation in human alveolar cells. Using human A549 cells we model inflammation and the immuno-modulatory effects of erythromycin. We use an ELISA to determine exact concentrations of various mediators of inflammation including IL-8 and TNF-α. Our ultimate goal is to provide more options for relieving inflammation in the pulmonary system.
In Summary, our research could lead to new treatments for patients prone to severe lung injuries due to bacterial infections, including those with immunosuppression, cancer patients, burn victims and individuals with cystic fibrosis.