Exploring human-specific intestinal development through great ape organoids and organ atlases

Kilik, Umut. Exploring human-specific intestinal development through great ape organoids and organ atlases. 2023, Doctoral Thesis, University of Basel, Faculty of Science.

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Official URL: https://edoc.unibas.ch/96299/

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Diet, microbiota, and other environmental exposures, together with metabolic requirements of a larger brain, place the intestinal epithelium as a nexus for evolutionary change in the human lineage. We know that dietary innovations through cooking and agriculture have had profound changes in recent human evolution, with lactase persistence into adulthood being one of the most striking examples of positive selection in humans. Genome comparisons between humans and the other great apes have identified genetic changes that could underlie our uniquely human phenotypes. However, very few studies have functionally linked this genetic change to modern human phenotypes.
The ability to generate human intestine tissue from pluripotent stem cells offers significant opportunities for studying human intestine development and modeling disease. However, certain unresolved questions persist about the cell states within this system. This thesis aims to reveal the emerging cell states and types in developing human intestinal organoids (HIO) and tissues. Additionally, we aim to explore differences in intestine development between humans and chimpanzees at both cellular and genomic levels, utilizing a combined approach with stem cell-derived intestinal organoids and tissue atlases. This integrated strategy allows for a quantitative assessment of evolutionary changes in intestinal cell types between the two species and facilitates subsequent functional investigations in controlled culture environments. Understanding human-specific intestinal phenotypes is essential for understanding digestive disorders in modern human populations and uncovering potential disease susceptibilities.
In the first part, we use single-cell RNA sequencing to reconstruct the entire differentiation process of HIO development. We also profile the transcriptome of multiple developing human organs with single-cell resolution to quantify the specificity of HIO cell fate acquisition. By interrogating HIO development and comparing it to developing human endodermal primary tissues, we identify epithelium-mesenchyme interactions, transcriptional regulators involved in cell fate specification, and features of stem cell maturation in the intestine. We also identify off-target populations that emerge in the HIOs. Next, we follow up on the resource by analyzing HIOs generated from stem cells that harbor a deletion of intestinal master regulator CDX2. We find that the epithelium acquires a more anterior identity, and the mesenchyme is also disrupted and acquires lung-like features. Last but not least, we characterize organ-specific developing mesenchyme features and benchmark niche interaction inspired multiple culture conditions of intestinal organoids, including subepithelial fibroblast-derived NRG1 that enhances intestinal stem cell maturation in vitro.
In the second part, we wanted to use organoids to study the evolutionary history of gene regulation in the human intestinal epithelium. We first reconstruct the gene regulatory network of the developing human small intestine using primary tissue and HIOs. We use comparative genomics to determine the deepest ancestry of human intestinal gene regulatory repertoires, identifying periods of change related to developmental, metabolic, and immune processes. We find that epithelial and immune cells of the developing intestine have more recent adaptations, with absorptive enterocytes showing the fastest rate of evolutionary change within the intestine. We then establish chimpanzee intestinal organoids to identify human-specific features in the gut using single-cell RNA and ATAC sequencing. Comparative and functional genomic analyses highlighted genic and regulatory elements with human-specific signatures and activity specific to distinct cell types/states. In addition, we establish human, marmoset, and mouse adult-stem cell-derived enteroids to define the features that persist into adulthood and could be modeled in 3D culture environments to test the human evo-devo hypotheses. Finally, we establish the first multiomic primate intestine reference atlas to systematically describe human-specific features and determine which genic and regulatory features of the human intestinal cell types persist into adulthood.
Collectively, we performed deep characterization of developing human intestinal organoids (HIOs) derived from induced pluripotent stem cells (iPSCs), highlighted features that could be modeled based on primary tissue counterparts, and improved the existing models. Establishing chimpanzee intestinal organoids allowed us to pinpoint developmental differences during intestine development at single-cell resolution and provide the first quantitative assessment of the evolutionary change between the human and chimpanzee gut. This work paved the way for functional testing of the role of genes and regulatory elements in controlled culture environments. Altogether, this thesis provides an atlas of human-specific intestinal features and suggests a new inroad to use intestinal organoids from our closest living relatives to explore the human condition.
Advisors:Camp, Jarrett Grayson and Doetsch, Fiona
Committee Members:Liberali, Prisca and Wells, James
Faculties and Departments:09 Associated Institutions > Friedrich Miescher Institut FMI > Quantitative Biology > Cellular heterogeneity during collective cell behavior (Liberali)
UniBasel Contributors:Doetsch, Fiona
Item Type:Thesis
Thesis Subtype:Doctoral Thesis
Thesis no:15305
Thesis status:Complete
Number of Pages:172
Identification Number:
  • urn: urn:nbn:ch:bel-bau-diss153051
edoc DOI:
Last Modified:03 Apr 2024 04:30
Deposited On:02 Apr 2024 08:19

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