Date of Award
Doctor of Philosophy (PhD)
Bruce Z. Gao
Cardiomyopathy, a disease of the heart muscle, is characterized by structural and functional abnormalities of the myocardium, in the absence of coronary artery disease, hypertension, valvular disease or congenital heart disease. In 2020, there were approximately 6.11 million prevalent cases of cardiomyopathy and myocarditis, and 0.37 million related deaths worldwide. Many affected individuals are asymptomatic, and chronically treated patients are at risk of heart failure. Within the myocardium, cardiomyocytes reside in a complex and dynamic extracellular matrix (ECM), consisting of basement membrane and interstitial matrix. The interactions between cardiomyocytes and myocardial ECM play a critical role in maintaining cardiac geometry and function throughout cardiac development and in adult hearts. Evidence from patient and animal models has demonstrated that abnormal alterations of myocardial ECM composition and nanostructure are pathogenic and associated with cardiomyopathy. Understanding how the structural changes of myocardial ECM affect cardiomyocyte function requires knowledge of pericellular structures. A critical barrier is the lack of effective tools that allow for in situ characterization and assessment of cardiomyocyte microenvironment at the tissue level. In my doctoral research, multimodal stimulated emission depletion (STED) microscopy was developed to characterize structural features of cardiomyocyte microenvironment in the pericellular domain at the tissue level. A multimodal STED microscope was constructed, consisting of two-color confocal fluorescence, two-color STED, and two-channel second harmonic generation (SHG) imaging modalities. This microscope also offers routine sample examination tools, including brightfield microscopy and widefield fluorescence microscopy. Using standard preparation procedures developed for one microscopy in another may introduce imaging artifacts, compromise image quality, and lead to inaccurate characterization. Sample preparation strategies were developed and optimized to balance image quality when conducting multi-scale and multi-aspect characterization using multimodal STED microscopy. The power of multimodal STED microscopy for in-depth characterization of myocardial structures was demonstrated on murine myocardial tissue samples. SHG imaging achieved label-free examination of fibrillar collagen in interstitial matrix. SHG- and autofluorescence-facilitated multiplexed imaging enabled the interpretation of protein distribution in 3D. Super-resolution STED imaging revealed pericellular basement membrane structures in the myocardium. Moreover, meaningful measurements retrieved from acquired images, such as sarcomere length and capillary density, enabled quantitative assessment of myocardial structures. In summary, multimodal STED microscopy provides a versatile approach for structural characterization of myocardial ECM, holding the promise for uncovering the molecular mechanisms underlying cardiomyopathy.
Zhang, Zhao, "Structural Characterization of Cardiomyocyte Microenvironment in Murine Myocardium by Multimodal Stimulated Emission Depletion Microscopy" (2023). All Dissertations. 3470.
Author ORCID Identifier
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