Supervisor: Dr. Chris Kennedy
Award: KRESCENT Summer Studentship
Institution: University of Ottawa/The Ottawa Hospital Research Institute
Year: 2025
 
Project Title: Role of Proximal Tubule NOX5 in Angiotensin II–Induced Hypertensive Renal Injury

Biography:
Michael is a student at the University of Ottawa, studying Translational Molecular Medicine (entering his 4th year). His passions include serving his community, long distance running, and travelling.  He has previously done research in the SEA-PHAGES program and at the Neural Regeneration Laboratory.  

Lay Summary:
My work is on the NADPH oxidase enzyme 5 (NOX5). NOX5 is a transmembrane enzyme that has been identified as a significant contributor to renal ROS (Reactive Oxidative Species) in humans, namely superoxide. Previous studies have shown that NOX5 is induced in the proximal tubule of individuals with hypertension. It is also an enzyme that is responsive to angiotensin II signaling. NOX5 expression and activation might therefore contribute to cellular injury associated with inflammation and/or fibrosis that occur in renal tissue as hypertensive kidney disease progresses. In order to better understand the functioning of NOX5, two hypotheses will be tested.

The first hypothesis states that transgenic human NOX5 expression in the proximal tubule of mice confers injury exacerbation and increased blood pressure. To test hypothesis 1, a line of mice with inducible NOX5 expression in proximal tubule will be subjected to angiotensin II-induced hypertension for 4 weeks. Pax8 rtTA mice are crossed with a line of NOX5 transgenic mice to generate NOX5PT mice. These mice are fed a chow containing doxycycline for 2 weeks prior to implantation of osmotic pumps to deliver angiotensin II. Blood pressure is measured by tail cuff plethysmography weekly. Blood sampling and spot urine collection are performed one week prior to pump installation and at the end point. Plasma is obtained for creatinine and BUN determination. Urine is assayed for albumin:creatinine and for large EV (extracellular vesicle) quantification. Kidneys are collected at endpoint for histology, NOX5 immunohistochemistry, and kidney injury marker assessment (mRNA/protein).

The second hypothesis states that NOX5 drives proinflammatory gene expression and extracellular vesicle formation in human proximal tubular epithelial cells. To test this, a human proximal tubule epithelial cell line is cultured and treated with a concentration range of angiotensin II (0-500 nM) for 24 and 48-hour times. Media is collected for EV quantification and mtDNA analysis. Cells are also harvested for bulk RNAseq studies. Furthermore, NOX5 expression can be manipulated by siRNA-mediated knockdown or adenoviral-mediated transduction.
Knowing the answers to these questions will give us crucial insights into the interplay between NOX5 and Angiotensin II.