Development of a method to examine shear stress on cardiomyocytes
Biomedical Engineering Committee
Master of Science
Ritter, Arthur B.
Reisman, Stanley S.
Rockland, Ronald H.
Renin Angiotensin System (RAS)
Cardiac hypertrophy is the remodeling or enlargement of the heart exhibited by individuals suffering from heart disease. Cardiac hypertrophy involves two mechanisms, mechanical stresses and humoral changes in the cardiac tissue. One such humoral change involves the Renin Angiotensin System (RAS). The primary fimction of RAS is maintenance of hemodynamics of the body mediated through an effector protein, Angiotensin 11. However, RAS has been shown to play a significant role in the pathophysiology of the heart; contributing to hypertension, cardiac hypertrophy and myocardial infarction. RAS acts through binding of Angiotensin 11 at its receptors. The two forms of Angiotensin 11 receptors, designated type one (ATI) and type two (AT2), have been shown to have different functions and distribution in various body tissues. Receptors for both ATI and AT2 are expressed on cardiomyocytes. The level of expression of each receptor type has been shown to have important implication. It has been suggested that the receptors may have cooperative yet opposing effects when bound by Angiotensin 11. Thus, the level of expression may drive the system to or away from hypertrophy.
The current research developed techniques to allow examination of the hypothesis that in cardiomyocytes, exposure to increasing shear stress causes upregulation of the ATI and AT2 receptors. The isolated rat cardiomyocytes were exposed to shear stress and methods were developed to measure the change in ATI and AT2 cellular expression resulting from the application of shear stress. The techniques required to accomplish this task including isolation of rat cardiomyocytes, flow chamber (shear stress) apparatus and use, handling of cardiomyocytes and molecular science techniques (RT-PCR) are described here in detail. The ability to track gene regulation of ATI and AT2 receptors though RT-PCR techniques is demonstrated. The ability to attach cardiomyocytes to a culture dish surface in order to expose them to shear stress is also shown. However, to take full advantage of the flow chamber, stronger cell attachment to culture dish is required. Future research will be directed to achieving stronger adhesion between cardiomyocytes and the cell culture surface. Recent reports have suggested use of various culture surfaces and adhesion molecules including laminin, elastin, cell culture plastic, and fibronectin. Use of serum and serum free medium has also been suggested.
njit-etd2002-030 (49 pages ~ 2,681 KB pdf)
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Created March 26, 2003