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Joshi Dissertation final.pdf (5.45 MB)

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Micro-engineering of embryonic stem cells niche to regulate neural cell differentiation

Joshi, Ramila, Joshi
http://rave.ohiolink.edu/etdc/view?acc_num=akron1544029342969082

Abstract Details

2018, Doctor of Philosophy, University of Akron, Biomedical Engineering.
Neurodegenerative diseases that are caused by deterioration of nerve cells in the brain and spinal cord affect more than 6 million Americans and cost nearly 0.8 trillion dollars annually in patient care. With a growing number of elderly population, the statistics are expected to worsen as there is currently no cure for these disorders. Modern medicines are at best palliative and only manage the symptoms. Therapeutic interventions to deliver functional neural cells to the ravaged tissue are essential to restore lost tissue functions. The use of stem cell-derived neural cells is a promising strategy for cell replacement therapies of neurodegenerative diseases. Embryonic stem cells (ESCs) are promising cell sources for therapeutic uses including cell replacement therapy of neural tissues. This is because ESCs have unlimited self-renewal and proliferation capabilities and the ability to differentiate into various neural cells. Nevertheless, despite significant investment and research, therapeutic uses of ESCs for neural cell replacement has been largely unsuccessful. Low and inconsistent yield of neural cells from ESCs and lack of a complete understanding of molecular mechanisms of neural differentiation of ESCs are major obstacles against clinical uses of ESC-based therapies. A cohort of cell surface bound and soluble factors, interactions of ESCs with their neighboring cells and extracellular matrix proteins, and various epigenetic factors may act synergistically to drive differentiation of stem cells. While most of current research is centered on functionalizing specific biomolecules on scaffolds and tuning the matrix stiffness, or altering media compositions to gain a better control over the neural differentiation of stem cells, the role of niche-mediated factors is less understood. In this study, we showed that intrinsic niche parameters such as stem cell colony size and interspacing between the two colonies can significantly impact the differentiation efficiency of stem cells toward neural cell lineages. Using our contact-free cell micro-printing technology, we generated ESC colonies of defined size and interspacing on stromal cells. Stromal cells-ESCs interactions and self-regulatory signaling of ESCs drive neurogenesis of ESCs. Our results showed that the neural differentiation capacity disproportionately enhances with increase in the colony size. Our multi-step statistical analysis approach using a combination of hierarchical clustering, change point detection, and multiple t-tests pinpointed specific sets of genes that underlie colony size-mediated neural differentiation of stem cells. We also established that, optimally interspacing ESC colonies further enhances neural differentiation of ESCs, significantly greater than that of individual colonies. Our computational model of spatial distribution of soluble factors of cells in interspaced colony pairs showed that the enhanced neural differentiation was due to the presence of stable concentration gradients of soluble signaling factors between neighboring colonies. Our results indicate that culturing ESCs in colony pairs with a defined interspacing distance increases the efficiency of deriving neural cells from ESCs. Additionally, we elucidated the self-regulatory role of ESCs in their neural cell differentiation. We supplemented conditioned media from differentiating ESCs to the ESCs-stromal cells co-cultures and quantitatively evaluated neural differentiation of ESCs. Our statistical tools segregated various growth and trophic factors produced by ESCs and stromal cells in the co-culture. Through this study, we showed the potential of modulating embryonic stem cells differentiation to the cells of neural lineages by using a simple engineering strategy to alter two niche parameters, i.e., size and the interspacing of stem cell colonies, without any additional differentiation-inducing chemicals. This study provided a model for quantitative studies of molecular mechanisms that regulate neurogenesis of stem cells. Future efforts to identify major soluble factors that drive and regulate the neural differentiation process in the ESCs-stromal cells co-culture can be a major step toward developing novel protocols to differentiate stem cells using novel media compositions and without using feeder stromal cells to help transition the finding to clinical applications. Our micro-engineering approach to improve the yield of differentiating ESCs to specific types of nervous system cells may make a major impact on stem cell research for treating neurodegenerative diseases.
Hossein Tavana (Advisor)
Marnie Saunders (Committee Member)
Nic Leipzig (Committee Member)
Yang Yun (Committee Member)
Sailaja Paruchuri (Committee Member)
242 p.
Biomedical Engineering; Biomedical Research; Engineering; Neurobiology
colony size, colony interspacing, embryonic stem cells, neural differentiation, stromal cells coculture, self induced neural differentiation, change point detection, stem cell secreted factors, niche engineering, stem cell microenvironment

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Citations

  • Joshi, Joshi, R. (2018). Micro-engineering of embryonic stem cells niche to regulate neural cell differentiation [Doctoral dissertation, University of Akron]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=akron1544029342969082

    APA Style (7th edition)

  • Joshi, Joshi, Ramila. Micro-engineering of embryonic stem cells niche to regulate neural cell differentiation . 2018. University of Akron, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=akron1544029342969082.

    MLA Style (8th edition)

  • Joshi, Joshi, Ramila. "Micro-engineering of embryonic stem cells niche to regulate neural cell differentiation ." Doctoral dissertation, University of Akron, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=akron1544029342969082

    Chicago Manual of Style (17th edition)

akron1544029342969082
201
© 2018, all rights reserved.
This open access ETD is published by University of Akron and OhioLINK.