Browsing by Subject "cardiomyopathy"
Now showing 1 - 2 of 2
- Results Per Page
- Sort Options
Item Differential Gene Expression Profiling in a Small Animal Model of Progressively Pacing-Induced Heart Failure(2006-06-01) Selby, Donald Evan; Stephen R. Grant; Patricia A. Gwirtz; Dan DimitrijevichDonald Evan Selby, Differential Gene Expression Profiling in a Small Animal Model of Progressively Pacing-Induced Heart Failure. Doctor of Philosophy (Biomedical Sciences), July 2006, 235 pp, 4 tables, 35 illustrations, references, 328 titles. Pacing induced tachycardia (PIT), in mammals, is known to cause a change from normal heart function to early left ventricular dysfunction. Progression to heart failure in experimental animals, such as dogs, pigs, and sheep, takes place in a relatively short period of time compared to the disease development observed in humans. Due to the cost and nature of using such animals, there is a need for a small animal model of PIT, which would delineate the etiology of the disease state by impairing the systolic function. The mode of action of overpacing inducement of cardiomyopathy, as the data suggests, may be through a sarcomere stretch sensor and its length-dependent signaling mechanism. In this study, an internal electrical-overpacing of an isogenic rabbit strain over a 52-day period was used to initiate a pathology consistent with human CHF. The data presented demonstrated that PIT causes alterations in the systolic ability of the heart, observed as reduced fractional shortening of the heart. This is seen in changes of the message pool population for proteins of the contractile architecture. Initially the heart is being paced rapidly and therefore there is insufficient time to get blood into the chamber. Thus, the data suggests that a mechanical stretch sensor is the process by which overpacing the heart leads to changes in gene expression which ultimately cause a compounding cellular condition which exists during heart failure. The data shows that there are gene isoform ratio changes that occur as the disease develops these include changes in differential expression of cardiac titin alternative splicing isoforms. The data suggests that there is also isoform switching occurring with alternative splicing of the gene encoding for SERCA2a, the probe 1587641_at shows a moderate decrease in expression and using BLAST for this probe this sequence is homologous to an alternative splicing variant of SERCA2a of the rabbit accession number J04703. The data shows that ferritin heavy chain also has an alternative splicing variant that are differentially regulated, this dysregulation of the isoform ratio may be linked to ADAMSTS1, a disintegrin and metalloproteinase isoform 1, which is seen to be downregulated in the data, these play a role in negative regulation of cellular proliferation. In addition to these detected isoform changes in the ratios of alternative splice variants changes are seen in genes linked to sarcomere integrity such as dystrophin probe 1582958_at is significantly increased in its expression, also integrin beta-1 probe 1584175)at shows a marginal increase in expression. The protease calpain probe 1604384)at, which uses a substrate the aforementioned integrin, dystrophin, and titin is also significantly upregulated in the data. Interestingly calpastatin, probe 1591603_at the inhibitor of calpain is marginally increased in its expression. Only recently has titin become to be appreciated as the protein that is responsible for the Frank Starling law as it undergoes an isoform ratio change as heart failure develops. These changes are initially caused by changes in ion concentration and stress upon the contractile proteins but as seen in the study, leads to altered gene expression. In this model, these gene alterations lead to diastolic dysfunction and the compounded problems constitute heart failure. This work shows that heart failure induced by over-pacing creates physical demands upon the framework of the heart and these physical stresses are transmitted through mechanical sensors leading to differential expression of the message pools for proteins involved in the way the heart contracts, and fills upon relaxation which ultimately ends in a heart that can do neither, thus leading to death.Item Studies in Molecular Mechanisms of Skeletal Muscle Contraction: Applications to Transgenic Mice with Inherited Cardiomyopathies(2013-05-01) Midde, Krishna; Julian BorejdoMuscular contraction impacts virtually every physiological function in a human body. Nearly 50% of the body weight is contributed by muscle. Skeletal muscle contraction is responsible for performing every day general functions such as moving, grabbing things and lifting weights. Cardiac muscle contraction is responsible for blood circulation in the body and smooth muscle contraction is involved in the contraction of hollow organs such as lungs, stomach and kidneys. It also maintains body temperature. Central to these events is the transformation of energy derived by hydrolysis of ATP to mechanical force. The force generating steps of muscle contraction is thought to result from the interaction of actin and myosin proteins. Although much of the general knowledge of the mechanism of contraction has been known for over 50 years, emerging advanced techniques have identified some of the key intermediate steps and regulating parameters. My doctoral research involves utilizing one such high resolution technique – single molecule fluorescence spectroscopy. I have used it to discern the motion and conformation of myosin cross-bridges in ex-vivo muscle. When studying the dynamic behavior of actin and myosin it is essential to reduce the number of molecules under observation because 1. The local concentration of actin and myosin is dense and the information obtained by averaging trillions of molecules doesn’t reflect the true character of the process. 2. The trajectory of an enzyme catalyzed reaction cannot be followed and the associated kinetics of actomyosin interaction is lost. 3. The situation becomes worse when studying mutations in sarcomeric proteins with low expression. 4. Heterogeneity between neighboring sarcomeres can be reduced. An important goal of muscle research is to measure the rate of the power stroke. Therefore, part of my thesis is focused on characterizing the pre- and post- power stroke states of muscle contraction. While the extent of force generated during muscle contraction is proportional to the extent of Ca2+ released into the muscle, some of the recent studies have shown that the phosphorylation of the regulatory light chain (RLC) of myosin also modulates contraction. Considering the importance of the phosphorylation of RLC, I investigated the distribution of orientation and kinetics of myosin cross-bridges in phosphorylated and de-phosphorylated muscle. Lastly, I have applied our fluorescence polarization technique with single molecule sensitivity to unravel the deranged contractile properties of muscle in people afflicted with Familial Hypertrophic Cardiomyopathy (FHC) disease. Some of the significant conclusions drawn from my project include evidence for the existence of distinct pre- and post- power stroke states of myosin cross-bridges during contraction in Ex Vivo muscle, Regulatory Light Chain phosphorylation disturbs cross-bridge organization and enhances the power stroke state of contraction and FHC induced by mutations in Troponin-T protein impairs myosin cross-bridge interaction with actin and alters cross-bridge kinetics. Clinically, drugs can be developed to modulate power stroke and enhance muscle performance in myopathies. Site targeted small molecules (peptides) can now be screened to correct for hypo-contractile or hyper-contractile properties associated with FHC. Our technique may also serve as a diagnosis tool for early identification of FHC disease. Finally increasing the basal ATPase activity of resting muscle by RLC phosphorylation is of therapeutic importance in treating individuals with Obesity, Type II Diabetes and Metabolic syndrome.