Date of Award

2016

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Pharmacology and Experimental Therapeutics

Abstract

Cardiac myosin binding protein-C (cMyBP-C) is a thick filament-associated protein that has been suggested to regulate cardiac contraction via its amino terminal (N’) region. Following ischemic injury to the heart, cMyBP-C is cleaved into a predominant N’ fragment consisting of domains C0 through C1 and the first 17 residues of the M-domain that is referred to as C0-C1f. However, the necessity of the N’-C0-C1f region of cMyBP-C in regulating cardiac function in vivo has not been elucidated. I hypothesized that the N’-C0-C1f region of cMyBP-C is critical for normal cardiac function in vivo. To test this hypothesis, transgenic (TG) mice with 81 ± 2.2% expression of a truncated cMyBP-C missing the N’-C0-C1f region (cMyBP-C110kDa) were generated and characterized in comparison to their non-transgenic (NTG) littermates between 3- and 8-months of age. Echocardiography at 3- and 6-months of age showed a significant reduction in percent fractional shortening (FS) in cMyBP-C110kDa hearts compared to NTG hearts at both time-points indicating progressive cardiac dysfunction in cMyBP-C110kDa animals. I further observed cardiac enlargement, determined by whole-heart confocal imaging, and an elevation in cardiac pathological hypertrophy markers, determined by quantitative real-time PCR (qPCR) and RNA-Seq, in cMyBP-C110kDa compared to NTG animals. Histopathological and second harmonic generation (SHG) analyses on myocardial sections indicated increased cardiac fibrosis (p < 0.0001) and increased sarcomere area (p < 0.01) in cMyBP-C110kDa hearts compared to NTG hearts. Crucially, immunofluorescence analysis of isolated cardiac myocytes from cMyBP-C110kDa and NTG hearts revealed that the cMyBP-C110kDa TG protein localized properly at the C-zone within the cardiac sarcomere. Intriguingly, increased phosphorylation of the cMyBP-C110kDa TG protein within the M domain was observed in cMyBP-C110kDa myofilament fractions compared to those from NTG controls. Finally, using isolated, permeabilized papillary muscle fibers from cMyBP-C110kDa and NTG hearts, I determined that a significant elevation in maximum force (p < 0.01) and rate of tension redevelopment (ktr) (p < 0.05) were produced from cMyBP-C110kDa fibers compared NTG fibers. Based upon this data, I conclude that the N’-C0-C1f region of cMyBP-C regulates cardiac contractility and is necessary for maintaining normal cardiac function in vivo.

Creative Commons License

Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License.

Download

Included in

Physiology Commons

Share

COinS