Proteomic analysis of heart failure : insights into myofibril assembly and regulation
Stanley, Brian Allan
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Heart failure (HF) is a prevalent disease in society which is associated with decreased cardiac output. This thesis describes the proteomic analysis of cardiac tissue obtained from HF patients and a transgenic animal model of HF. Initial experiments optimized one proteomic technology, 2-dimensional gel electrophoresis (2-DE), to maximize the number of proteins which could be observed / resolved from human cardiac tissue. Protein abundance changes in cardiac tissue between normal patients and those with a diagnosis of ischemic cardiomyopathy were determined by performing 2-DE and identifying proteins by mass spectrometry. HF patients had a reduced abundance of proteins involved in energy production and the sarcomere. Sarcomeres contain the myofilament subproteome consisting of thick and thin filaments with the thick filaments primarily myosin. Thick and thin filament undergo Ca2+ induced ATP hydrolysis to form crossbridge cycles, resulting in muscle contraction. An assembly chaperone for myosin, UNC-45B, was found to be increased in HF patients. Western blot analysis confirmed that the abundance of UNC-45B was increased in different etiologies of heart failure. Follow up physiological measurements demonstrated that UNC-45B is most likely a protein necessary for transcriptional control of the α-isoform of myosin heavy chain. In a second proteomics study, abundance changes occurring in a pacing induced model of HF in wild-type (WT) and transgenic (TG) rabbits with increased expression of the α-isoform of myosin heavy chain were examined. WT and TG rabbits had a different response in their myofilament and intermediate filament abundance changes following induction of HF. TG rabbits had a decreased abundance of heat shock proteins and non-sarcomeric associated desmin. As well, TG rabbits had an increased ratio of actin:myosin heavy chain and UNC-45B suggesting an altered ratio of thick to thin filaments. In conclusion, an altered abundance of contractile proteins, regulated in part by UNC-45B, may be one cause of the contractile dysfunction which occurs in HF.