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During intestinal organogenesis, equipotent epithelial progenitors mature into phenotypically distinct stem cells that are responsible for lifelong maintenance of the tissue. While the morphological changes associated with the transition are well characterized, the molecular mechanisms underpinning the maturation process are not fully understood. Here, we leverage intestinal organoid cultures to profile transcriptional, chromatin accessibility, DNA methylation, and three-dimensional (3D) chromatin conformation landscapes in fetal and adult epithelial cells. We observed prominent differences in gene expression and enhancer activity, which are accompanied by local changes in 3D organization, DNA accessibility, and methylation between the two cellular states. Using integrative analyses, we identified sustained Yes-Associated Protein (YAP) transcriptional activity as a major gatekeeper of the immature fetal state. We found the YAP-associated transcriptional network to be regulated at various levels of chromatin organization and likely to be coordinated by changes in extracellular matrix composition. Together, our work highlights the value of unbiased profiling of regulatory landscapes for the identification of key mechanisms underlying tissue maturation.

Tissues with a high turnover rate produce millions of cells daily and have abundant regenerative capacity. At the core of their maintenance are populations of stem cells that balance self-renewal and differentiation to produce the adequate numbers of specialized cells required for carrying out essential tissue functions. Here, we compare and contrast the intricate mechanisms and elements of homeostasis and injury-driven regeneration in the epidermis, hematopoietic system, and intestinal epithelium-the fastest renewing tissues in mammals. We highlight the functional relevance of the main mechanisms and identify open questions in the field of tissue maintenance.

Weefselspecifieke volwassen stamcellen hebben de aandacht getrokken van de biomedische gemeenschap doordat ze in staat zijn om alle celtypes van een orgaan te vormen waardoor ze bruikbaar zouden kunnen zijn voor stamceltherapie. De hoop is dat deze cellen beschadigd weefsel kunnen herstellen en wellicht zelfs voor volledige vervanging van organen kunnen zorgen. Zo zouden bijvoorbeeld volwassen stamcellen die in staat zijn tot vorming van speekselklierweefsel gebruikt kunnen worden om patiënten die lijden aan het droge mond syndroom (Xerostomie) te behandelen. Xerostomie houdt in dat te weinig speeksel wordt geproduceerd en is een veel voorkomende bijwerking als gevolg van radiotherapie bij hoofd- en hals-tumoren, Sjögren-syndroom, diabetes, ouderdom en het gebruik van sommige medicijnen. Het is echter van het grootste belang dat de nog ontbrekende informatie over de identiteit, lokalisatie, en de kritieke moleculaire signalen die nodig zijn voor het handhaven en kweken in een laboratorium van volwassen speekselklier stamcellen tot in detail onderzocht worden. Het onderzoek beschreven in dit proefschrift draagt hieraan bij en verbetert de huidige kennis over de identiteit van de speekselklierstamcel en de moleculaire signaal routes betrokken bij speekselklier regeneratie. Verder wordt de optimalisatie van een kweekmethode voor volwassen speekselklier stamcellen van muis en mens beschreven. Deze kennis wordt toegepast om een beter begrip van de speekselklier stamcelbiologie tijdens weefsel homeostasis, regeneratie en veroudering te krijgen. De in dit proefschrift gepresenteerde gegevens benadrukken het veelbelovende therapeutische potentieel van speekselklier stamcellen in regeneratieve geneeskunde., The biomedical world has drawn attention to tissue specific stem cells or adult stem cells because of their ability to produce all different cell types of an organ and as such may be useful for stem cell therapies. The hope is that these cells may restore and replenish damaged tissue and even form complete replacement of organs. For example, adult stem cells that give rise to salivary gland tissue could be used to treat patients suffering from xerostomia, a common side effect of head and neck radiotherapy, Sjögren’s syndrome, diabetes, old age and numerous medications. However, paramount information regarding the identity, localization, critical molecular signals maintaining adult salivary gland stem cells and methods for culturing these cells in a laboratory setting are largely missing. Therefore, the work presented in this dissertation contributes to and enhances the current knowledge regarding salivary gland stem cell identity and the core molecular pathways necessary for salivary gland regeneration. Moreover, we describe a development of culture methods of mouse and human adult salivary gland stem cells, which we apply to gain better understanding of salivary gland stem cell biology during tissue maintenance, regeneration and ageing. Thus, the data presented here highlight the promising therapeutic potential of salivary gland stem cells in regenerative medicine.

Tissue regeneration requires dynamic cellular adaptation to the wound environment. It is currently unclear how this is orchestrated at the cellular level and how cell fate is affected by severe tissue damage. Here we dissect cell fate transitions during colonic regeneration in a mouse dextran sulfate sodium (DSS) colitis model, and we demonstrate that the epithelium is transiently reprogrammed into a primitive state. This is characterized by de novo expression of fetal markers as well as suppression of markers for adult stem and differentiated cells. The fate change is orchestrated by remodeling the extracellular matrix (ECM), increased FAK/Src signaling, and ultimately YAP/TAZ activation. In a defined cell culture system recapitulating the extracellular matrix remodeling observed in vivo, we show that a collagen 3D matrix supplemented with Wnt ligands is sufficient to sustain endogenous YAP/TAZ and induce conversion of cell fate. This provides a simple model for tissue regeneration, implicating cellular reprogramming as an essential element. The mechanism that governs tissue regeneration following severe damage to the colonic epithelium remains poorly understood. Jensen and colleagues show that the colonic epithelium undergoes a profound reprogramming into a more primitive state with fetal-like properties. Moreover, they demonstrate that YAP and TAZ operate as essential mechano-sensors during tissue reprogramming.