Rachel R. Wade
| Rachel R. Wade |
Rachel R. Wade, Kathleen Boesze-Battaglia, PhD, Taewan Kim, DDS, Samuel Spadlin
Basic & Translational Sciences, University of Pennsylvania, School of Dental Medicine
Introduction
Oxidative stress results when there is an imbalance between reactive oxygen species (ROS) and antioxidants. These reactive oxygen species are created to kill invading microbes, but can also damage nearby tissue. ROS are thought to play a role in many inflammatory diseases. Cytolethal distending toxin (Cdt) is a toxin that is often associated with pathogens that involve persistent infection along with intense inflammatory response. The hypothesis of this experiment is that the Cdt toxin will increase the amount of ROS and oxidative stress and that the oxidative stress will be mitigated by LGM2605. LGM2605 is an MPO inhibitor. Myeloperoxidase (MPO) is an enzyme that catalyzes the formation of reactive oxygen intermediates. More specifically, MPO using hydrogen peroxide and halides, catalyzes the formation of hypochlorous acid (HOCl). The experiment in this project will attempt to find MPO at the translational level and mRNA expression levels, and the activity of ROS species in THP-1 cells with the cdt toxin. These studies aim to answer the question: Does Cytolethal Distending Toxin (Cdt) produced by produced by Aggregatibacter actinomycetemcomitans (Aa) effect the amount of phagocytosis-associated oxidative stress? Can the oxidative stress be mitigated with synthetic antioxidants (LGM2605)?
Methods
The methods that will be utilized to conduct this experiment are Western Blot, qPCR, and Flow cytometry. The experiment will use THP-1 cells. The Western Blot will be conducted with human/mouse MPO antibody to find translation levels of MPO protein. qPCR will be conducted to find mRNA- expression of MPO. Flow cytometry will be conducted to assess ROS activity.
Results
Our hypothesis predicts that Cdt will increase phagocytosis associated oxidative stress and enhance MPO activity. Pretreatment of cells with LGM2605 is expected to mitigate this effect. Studies are currently under analysis with detail to follow.
Conclusion
Conclusions about this project cannot be made until more data is available.