Thesis
Design of a cone and plate co-culture device to investigate the effects of fluid shear stress and free fatty acids on endothelial nitric oxide production
Master of Science (M.S.), Drexel University
Jun 2016
DOI:
https://doi.org/10.17918/etd-6838
Abstract
Metabolic syndrome and obesity in particular are significant cardiovascular disease risk factors. Excessive central adipose tissue is strongly linked to insulin resistance, which may contribute to systemic and adipose tissue vascular inflammation. Insulin resistant and obese individuals have increased levels of circulating free fatty acids, which inhibit insulin-induced production of atheroprotective nitric oxide by endothelial cells in vitro. Decreased endothelial nitric oxide can further lead to chronic inflammation of the vasculature and adipose tissue. Furthermore, insulin resistant and obese individuals display reduced adipose tissue blood flow. Thus the FFA-induced reduction in endothelial nitric oxide production may further decrease shear stress-induced endothelial nitric oxide production and exacerbate adipose tissue inflammation and metabolic disease. The objective of this thesis is to elucidate the effects of FFAs on endothelial cell response to fluid shear stress. I hypothesize that shear stress leads to endothelial nitric oxide synthase phosphorylation in the presence of FFAs. If proved true, this would suggest that restoring adipose tissue blood flow may abrogate adipose tissue inflammation and metabolic syndrome. Since existing in vitro fluid shear stress testing devices cannot test the effects of shear stress on endothelial cells in a 3D co-culture system with adipocytes, this thesis first presents a new cone and plate flow device with a 3D hydrogel co-culture component. The cone and plate flow system successfully created laminar fluid shear stress in both acute (5 minute) and chronic (24 hour) bovine aortic endothelial cells, demonstrated by increased endothelial nitric oxide synthase phosphorylation and cell alignment, respectively. This thesis then presents a method to incorporate a gelatin co-culture component; however the gelatin gel requires further study since it is not adequately flat nor have cells been incorporated. FFAs were demonstrated to inhibit insulin-induced endothelial nitric oxide synthase phosphorylation in static culture. Finally, preliminary data demonstrated that 5 minutes of flow at 20 dynes/cm2 was able to increase eNOS phosphorylation in samples treated with FFA incubation in 2 of 3 sample groups tested. This thesis has provided design of a new in vitro flow system that could allow testing of co-cultures previously not possible. The results presented here support further examination of the role of shear stress and FFAs in vascular disease. Future work will include the improvement of gelatin molding techniques with finer tolerance features, as well as developing methods to suspend the cells with the gel. Additional experiments should be conducted with FFAs in flow. Furthermore, the cone and plate device offers opportunities for work with other cell types and co-cultures including endothelial, adipocyte, smooth muscle, and macrophages.
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Details
- Title
- Design of a cone and plate co-culture device to investigate the effects of fluid shear stress and free fatty acids on endothelial nitric oxide production
- Creators
- Nicholas W. Houriet - DU
- Contributors
- Alisa Morss Clyne (Advisor) - Drexel University (1970-)
- Awarding Institution
- Drexel University
- Degree Awarded
- Master of Science (M.S.)
- Publisher
- Drexel University; Philadelphia, Pennsylvania
- Number of pages
- xiii, 147 pages
- Resource Type
- Thesis
- Language
- English
- Academic Unit
- School of Biomedical Engineering, Science, and Health Systems (1997-2026); Drexel University
- Other Identifier
- 6838; 991014632328904721