Molecular Mechanisms Regulating Shear Stress-Induced Nitric Oxide Production in Endothelial Cells

Friday, May 6, 2016

9:00 AM-11:00 AM

BIOMED PhD Research Proposal

Title:
Elucidating the Molecular Mechanisms Regulating Shear Stress-Induced Nitric Oxide Production in Endothelial Cells

Speaker:
Tenderano Muzorewa, PhD Candidate, School of Biomedical Engineering, Science and Health Systems

Advisor:
Ken Barbee, PhD, Professor and Interim Director, School of Biomedical Engineering, Science and Health Systems

Abstract:
Nitric Oxide (NO), a key regulator of vascular homeostasis, is produced by the endothelium in response to agonists and hemodynamic forces. Impaired NO production is the hallmark of endothelial dysfunction, which is a precursor to and a complication of several cardiovascular diseases, affecting up to one third of the US population. The goal of this study is to investigate the signaling mechanisms regulating flow-induced NO production in endothelial cells in order to shed light on the mode of impairment of a pathological endothelium.


Recent work found shear stress-induced NO production to depend on adenosine triphosphate (ATP) signalling and store-operated channel entry (SOCE), mediated by a complex interplay between Protein Kinase C (PKC) and SOCs in caveolae microdomains. In the proposed study, the identity and roles of the specific PKC isozymes and SOCs involved in this pathway will be investigated. The approach is to apply ATP or shear stress to bovine aortic endothelial cells, then measure the NO and calcium response following pharmacological and genetic interference with different PKC isozymes and calcium channels. The NO response will be measured by evaluating endothelial Nitric Oxide synthase (eNOS) activity at the protein level and by real-time detection of NO using an electrode. A fluorescent dye will be employed for assaying calcium signalling while immunofluorescent staining is used to monitor the intensity and localization of activated eNOS with respect to caveolae and calcium channels following ATP stimulation.


The findings of the proposed study would elucidate the role of molecular players involved in regulation of shear-induced NO production, as well as their interplay and the consequences of their microdomain compartmentalisation. This will in turn inform the treatment of endothelial dysfunction, for which there is no gold standard.

Contact Information

Ken Barbee
215-895-1335
barbee@drexel.edu