Barcoding tools to track development and function of stem cell constructs for biomedicine
Doctor of Philosophy
Biological regeneration has long been explored as a breakthrough modality for human therapy. Degenerative diseases, a majority of which are a related to ageing, manifest as a wide variety of pathologies: cardiovascular diseases, diabetes and Alzheimer to name a few. Consequently, the idea of a panacea in the form of a biological therapeutic that can grow and restore a debilitated bodily function has been much investigated. Outwardly, the premise has seemed remarkably simple. Scientists have long observed lizards growing back whole tails after decapitation and studied the remarkable regeneration potential of Planaria. Yet increasing work suggests processes including complex cellular growth dynamics, patterns of gene expression and clonal expansion of cells drives regeneration and analogously, biological tissue development. Improved understanding of developmental biology can thus improve the function and persistence of regenerative therapies. Our lab has previously developed in-vitro tools that allow the study of biological development. This work extends on that body by developing two distinct tool sets. Firstly, we develop a scheme for tracing large-scale clonal dynamics conveniently using a scalable lineage tracing method. We demonstrate the capacity of this system to accurately detect cell lineages based on a system of inheritable genetic barcoding. Our system can deconvolute lineage mixtures, track growth dynamics and consequently probe biological phenomena. Importantly, our process preserves spatial integrity of the sample and thus quantifies both lineage dynamics and captures their positional information. Since differential patterns of gene expression is another important component of regeneration and development, we develop a method to measure gene expression non-destructively, with the capacity to allow multiplexed monitoring of multiple targets even while in-vivo. As cellular and regenerative therapies move through primate and clinical trials, such a tool can serve the critical role of augmenting available biomarkers for monitoring therapy and predicting clinical outcomes thereby improving regulatory clearance. Taken together, our work adds to the body of knowledge that seeks to better understand tissue genesis and its re-engineering to create new therapeutic modalities.
Regenerative medicine; bioengineering; cell therapy; synthetic biology