Publication Date

2009

Document Type

Thesis

Committee Members

Norma Adragna (Committee Member), Mark Anstadt (Advisor), Mark Anstadt (Committee Co-chair), David Cool (Committee Co-chair), Lawrence Prochaska (Committee Member)

Degree Name

Master of Science (MS)

Abstract

Cardiovascular disease accounts for more than 40% of all deaths in the United States (AHA-2004 report). A non blood contacting ventricular assist device (VAD) can be used to treat heart failure without the complications that arise from blood contacting VADs. This study used cellular markers of heart failure as indicators of heart function in an attempt to assess if direct mechanical ventricular actuation (DMVA) support lessened the impact of heart failure in rabbits. Cell signaling proteins were monitored using enzyme activity measurements and quantitative immunoblotting with antibodies against intrinsic and extrinsic apoptotic pathways during heart failure with and without DMVA support.

New Zealand White rabbits were treated as sham or heart failure, with or without DMVA assistance. Animals had heart failure induced by esmolol, with or without DMVA support for 30, 60, or 120 minutes. At all time points, animals were recovered for 30 minutes after which the hearts were excised. Tissue extracts were prepared and measurements were made using the following groups; acute model with sham and 30 minute groups or an extended support/failure model with 60 minute and 120 minute groups. Cell extracts from left and right ventricles were used for immunoblot analysis to determine protein levels for the following heart failure markers; the extrinsic apoptotic pathway comprised of tumor necrosis factor receptor (TNFR) and caspase-8, the intrinsic apoptotic pathway comprised of caspase-9, cytochrome-C, or the stress related marker of heat shock protein-70 (Hsp70). Enzymatic activity was used to monitor the extrinsic pathway with caspase-8, or markers of stress with superoxide dismutase (SOD), and matrix metalloproteinase-9 (MMP-9) in all heart extracts.

In the left ventricle acute model there was a rise in both 30 minute groups compared to the sham group for all the markers, except in the caspase-8 enzymatic assay where both groups showed less activity. The extended support/failure model showed that the use of DMVA during heart failure significantly attenuated the increase of protein content for TNFR, Hsp70, and caspase-8 at the 120 minute time point. The caspase-8 enzymatic assay showed a significant decrease in 120 minute DMVA assisted group compared to the heart failure group. For every marker in the extended support/failure model there was an increase in all the markers as the duration of the experiment increased.

The right ventricle acute model exhibited an increase in both 30 minute groups for TNFR content. A significant rise in SOD levels for the 30 minute heart failure compared to both the sham and 30 DMVA groups. MMP-9 levels for both 30 minute groups rose significantly. In the extended support/failure model, the right ventricle showed a significant increase of both groups at 120 minutes when compared to the 60 minute heart failure group in TNFR and Hsp70 protein content. In the caspase-8 enzyme assay the 120 minute DMVA group had a significant rise when compared to the 60 minute heart failure group.

DMVA provided evidence of the ability to attenuate the increase of pro-apoptotic signaling during heart failure. Evidence of this was mostly observed in the left ventricle samples, often significantly different. Both ventricles exhibited trends of attenuation by DMVA in many markers that did not differ significantly. It should also be noted that at no time point did DMVA appear to have a negative effect on the heart during failure. The validity of the heart failure model being used was also confirmed by the continuing increase in activation of known markers over time.

Page Count

179

Department or Program

Department of Pharmacology and Toxicology

Year Degree Awarded

2010

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