Generation of a human foetal pancreatic organoid model of pancreas development

  • Michael Elsy

Student thesis: Master of Philosophy


Introduction: Diabetes mellitus (DM) represents a significant global health and economic burden, and its incidence is increasing. Although injectable insulin is the gold standard for Type 1 DM management, it confers life-long dependence and hypoglycaemia may instead occur. Therefore, restoration or regeneration of damaged insulin-producing beta-cells could provide a cure. Despite their immense potential, human pluripotent stem cell (PSC)-derived beta-cells currently do not have the appropriate functionality or maturation capability compared to bona fide beta-cells. A greater understanding of beta-cell specification during early human development is required for safe transplantation of PSC-derived beta-cells. Organoid systems refer to clusters of multiple cell types in 3-dimensions (3D) that can self-organise, allowing mimicry of the cell-cell contacts and signalling events of the native tissue environment. Human foetal pancreatic organoids are emerging as exciting models for pancreas development, enhancing our understanding of hPSC-derived beta-cell differentiation protocols, and providing opportunities for disease modelling. Aims and Objectives: This project aims to establish and maintain a 3D organoid model of human pancreas development using foetal tissue. Light sheet fluorescence microscopy (LSFM) will be optimised as a method of imaging key pancreatic transcription factors (TFs) throughout pancreas morphogenesis. Methods: Human foetal pancreata were dissected and dissociated into single-cell suspension before incubation with Matrigel. Organoids were cultured for up to 30 days before analysis. Following incubation, organoids were assessed by immunofluorescence. For comparison, immunohistochemistry and LSFM were performed on native embryonic or foetal pancreata. Results: Immunohistochemistry reveals the spatial and temporal expression profiles of key pancreatic TFs in embryonic and foetal pancreata. Distinct 3D ductal and islet structures were observed in foetal samples using LSFM. The expression of these TFs characterises our human foetal pancreatic organoids. Ductal, PPC and endocrine markers are detected in the organoid system, as represented by CK19, SOX9 and NKX2.2 expression, respectively. Discussion and conclusions: Pancreas forms when foregut endoderm is specified to pancreatic endoderm, followed by PPCs and then acinar or bi-potent ductal and endocrine populations, including beta-cells. 3D organoid culture of foetal pancreas permits expansion of multipotent PPCs, which can be used as a platform for modelling downstream endocrine differentiation. LSFM enables the rapid imaging of whole foetal pancreata and organoids; the localisation and expression of candidate PPC regulators could be investigated using LSFM. The surrounding niche and growth factor environment could also be manipulated, enabling study of their effects on endocrine (and beta-cell) differentiation in both foetal pancreas organoids and PSC-derived pancreatic organoids. In summary, the establishment of this pancreas organoid system is a valuable asset for furthering our knowledge of pancreas development and endocrine commitment.
Date of Award31 Dec 2022
Original languageEnglish
Awarding Institution
  • The University of Manchester
SupervisorAlberto Saiani (Supervisor), Rachel Jennings (Supervisor), Neil Hanley (Supervisor) & Karen Piper Hanley (Supervisor)

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