Noah Ziemba
Vascular Medicine Institute
Pulmonary arterial hypertension (PAH) is a type of cardiovascular disease that results in right heart failure and eventually death. One of the major pathways involved in this dysfunction is the nitric oxide (NO) pathway. NO is naturally produced in the vascular endothelium of the body where it holds multiple vasoregulatory characteristics. PAH has been found to result in decreased availability and responsiveness to NO. Because of this, NO has been targeted as a therapy for PAH.
NO can be reduced endogenously from nitrite through multiple mechanisms. One of these mechanisms is the molybdenum enzyme mARC2. The goal of my project was to characterize the function of mARC2 in catalyzing the reduction of nitrite to NO. The function of mARC2 is studied in vivo by observing the effect of mARC2 knockout in mice.
During the experiments, I assisted in caring for rodent specimen. A variety of tissues were excised from the mice, including liver, heart, and fat. From these tissue samples, I prepared tissue lysates with a Dounce homogenizer. The concentration of these lysates was determined through a BSA assay and linear regression analysis. With the lysates prepared, I then analyzed the levels of nitrite in each sample.
The amount of nitrite in the lysate is measured using a nitric oxide analyzer (NOA). A stoichiometrically known reaction is used to reduce nitrite to NO in the lysate. The NO is then reacted with ozone which produces a detectable photon. This signal can be recorded over time and analyzed to determine the amount of nitrite in the lysate. I collected samples for heart and liver tissue for multiple subjects throughout the duration of the project.
The ability to analyze my prepared tissues was hindered by the onset of the coronavirus and inability to work in person. As a result, I do not unfortunately have results from this project at the time. When it is safe to work in the lab again, I hope to continue this project.