Considerations for using isolated cell systems to understand cardiac metabolism and biology
Introduction
Metabolism is an area of traditional biochemistry that, until recently, was considered a relatively mature field. The backbone of metabolic knowledge reflects the success of our scientific approaches and technological advances. New methods for understanding complex system behavior have further propelled the scientific community’s quest to understand metabolism, not only by increasing the breadth of exploration, but also by increasing the throughput and analysis of metabolic data. In addition, the development of cell models of cardiovascular disease, such as induced pluripotent stem cells (iPSCs), have expanded interest in understanding the metabolic properties of cultured cells, which could be used to predict disease or to devise actionable interventions. Experimental systems that integrate cell models with techniques for measuring metabolic activity have been used to generate volumes of data that must be carefully weighed for elements of rigor and for their meaningfulness for understanding in vivo biology. Ultimately, the value of data from isolated cell systems depends upon their usefulness to contribute to coherent explanations of biological phenomena. In this review, we discuss methodological considerations for measuring cardiomyocyte cell metabolism.
Section snippets
How does the heart’s metabolic machinery process substrates?
A general goal of many metabolic studies is to understand how living systems regulate the production of useable energy. Because the heart has an extremely high energetic demand, it is a model organ for understanding how cells control ATP levels (reviewed in [1]). Large quantities of ATP are required to maintain cardiac ion homeostasis and contractile function, and this ATP demand is met via catabolism of a variety of circulating hydrocarbon substrates [[2], [3], [4], [5], [6], [7], [8]]. For
Respirometry using isolated mitochondria
Purified mitochondrial fractions can be used to understand specific aspects of mitochondrial biology, including respiratory efficiency and capacity [[16], [17], [18]]. Typically, respirometry uses oxygen electrode- or fluorophore-based systems to measure the consumption of O2 as an index of catabolic activity in isolated mitochondria, cells, tissues, or whole organisms. Because the vast majority of O2 in most cells is consumed at cytochrome c oxidase, the rate of O2 consumption is used to
Considerations for other metabolic assays in cultured cardiomyocytes
Other techniques in the investigator’s metabolic toolbox, discussed briefly below, complement XF data and provide additional information on catabolic pathway flux, redox state, and biosynthetic pathway activity (e.g., see: [88,89]).
Summary
Sensible application of metabolic assays can facilitate understanding of how metabolic changes influence cardiomyocyte biology. Initial experiments to optimize cell number, substrate media composition, and pharmacological compound concentrations are critical for deriving reproducible and interpretable results. Moreover, complementary metabolic assays can provide additional rigor and depth to experimental results and interpretation. Considerations for the advantages and limitations of each
Acknowledgements
This work was supported in part by grants from the NIH (HL130174, HL147844, ES028268, HL078825, GM127607, HL154663).
Disclosures
None.
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