Title:
Metabolic and Bioelectric Crosstalk in Directed Differentiation and Spatial Patterning of iPSC-Derived Cardiomyocytes
Metabolic and Bioelectric Crosstalk in Directed Differentiation and Spatial Patterning of iPSC-Derived Cardiomyocytes
Author(s)
Norfleet, Dennis Andre
Advisor(s)
Kemp, Melissa L.
Editor(s)
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Abstract
The goal of multi-cellular engineered living systems is the design and manufacturing of multicellular
systems with novel form or function using engineering design principles. Induced
pluripotent stem cells represent an excellent tool to enable actualization of these design goals
because of their intrinsic pluripotent capacity and recapitulation of various embryogenesis and
organogenesis processes. The objective of this research was to investigate through computational
modeling how molecular components of bioelectric and metabolic systems alter multicellular
bioelectric patterning and cell metabolic flux dynamics, and to extend system understanding to
guide emergent morphogenic outcomes via external modulation of the culturing environment.
The central hypothesis of this work was that specific media compositions can alter molecular
components of bioelectric and metabolic multicellular systems in a predictable manner, leading
to desired morphologies, cell phenotypes, and novel functionalities. In the first study, a multiscale
bioelectric computational model describing human iPSC tissue-scale membrane voltage
potentials (Vmem) was developed to understand unexplored patterning outcomes when various
molecular components of the bioelectric system are altered by culture media. Model simulations
accurately predicted multicellular Vmem patterns when one or more molecular components were
altered, as quantitatively confirmed by a machine learning-based quantitative image pattern
similarity analysis. In the second modeling analysis, a genome-scale computational model of the
human metabolic network was expanded with additional descriptors to investigate how induced
pluripotent stem cells reroute metabolic fluxes and achieved cell growth objectives during
cardiomyocyte differentiation under various culture media compositions. This framework
integrated transcriptomic, thermodynamic, kinetic, and proteomic and novel fluxome constraints
including transport exchange between the cytosol and extracellular environment. From a
comparative analysis across multiple published studies and our own experimental validations, we
observed that the combination of novel and previous model constraints was required to replicate
experimental media-induced changes in metabolic network dynamics during pluripotency and
hiPSC-cardiomyocyte (hiPSC-CM) differentiation. We extended this study to a novel media
supplementation condition of glutamine and ascorbic acid and found that experimental
extracellular flux assays supported the model-predicted improvements to metabolic respiration of
iPSC-derived cardiomyocyte progenitor cells. In summary, these results collectively validate the
potential for model-guided media design of engineered living systems using understanding of
bioelectric and metabolic systems properties.
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Date Issued
2023-05-08
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Resource Type
Text
Resource Subtype
Dissertation