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Characterizing drivers of intestinal tissue maturation in vitro and in vivo

Periodic Reporting for period 1 - fetISC (Characterizing drivers of intestinal tissue maturation in vitro and in vivo)

Reporting period: 2016-02-01 to 2018-01-31

Inflammatory bowel disease (IBD) affects millions of European citizens. Currently patients are treated with immunosuppressive palliative therapies and in some cases large segments of patients intestines are surgically resected. There is, however no medical cure. We proposed that IBD patients could benefit from a from epithelial cell transplantations if a suitable source can be identified. Human induced pluripotent stem cells (hiPSC) constitute a promising source of cells for transplantation, but current protocols for directed in vitro differentiation in a defined media produce cells with foetal characteristics. Moreover our group and others (Klein OD lab) recently publish that either during colitis regeneration or during parasitic helminths infections, intestinal cells are reprogrammed into a fetal-like state. In order to develop intestinal cell transplantations as well as in order to understand the cellular and molecular characteristics of IBD it becomes instrumental to understand: how intestinal stem cells are originated during foetal development, characterise their unique molecular properties, and elucidate the mechanisms driving foetal intestinal maturation towards an adult intestinal epithelium.
During the development of this project we have worked in several main aspects related to: basic research, results dissemination, communication and public engagement.

BASIC RESEARCH:

During the development of this project we have identified the place and the time where intestinal stem cells are specified during development as well as the mechanism driving this process. Our results demonstrate that large scale tissue remodelling and cell fate specification are intertwined processes. These findings provide a direct link between the observed plasticity and cellular reprogramming of differentiating cells in adult tissue following damage, revealing that stem cell identity is an induced rather than a hardwired property.Moreover we have identified several cellular populations in the fetal intestinal epithelium with engraftment and regenerative potential. Importantly we have identified the unique molecular and epigenetic properties of foetal and adult intestinal cells that we have validated with human foetal samples from abortions. This has been crucial in order to validate the conservation of mechanism identified.

RESULTS DISSEMINATION:

The results of this project have been shown in several scientific conferences including:

-The Stem cell Niche Conference 2018. Hillerød. Contribution: Oral presentation. Title: Tracing the origin of adult intestinal stem cells.
-International Society for Stem Cell Research-ISSCR 2017 (Innovation Showcase, Stem Cell Technologies). Boston. Contribution: Oral presentation. Title: Origin and Ancestry of Intestinal Stem Cells Precursors.
-Keystone Symposia: Stem Cells and Regeneration in the Digestive Organs 2016. Keystone. Contribution: Poster presentation. Title: Modelling human intestinal development in vitro. Authors: Jordi Guiu, Marianne T. Pedersen, Ludovic Vallier, Kim Jensen.

Moreover we expect to published the results in Peer reviewed journals. The first part of the work regarding origin and specification of intestinal stem cells was submitted and we expect to submit a second manuscript this year regarding molecular and epigenetic properties of foetal and adult intestinal cells. Additionally to this we have published a review emphasising the relevance of understanding intestinal development.

COMMUNICATION AND PUBLIC ENGAGEMENT:

Through the development of the project we have shown and explained some of our results as well as technical expertise to the general public in order to rise the general awareness and communicate the importance of our research. Moreover I have provided coursed for young and future researchers.

-Workshops addressed to High school teachers, where we showed and provided a hands-on training where they learned how to generated 3D intestinal stem cell cultures.
-Hands-on course for high school talented students where we showed the resistance of tumor and normal cells to radiotherapy.
-Supervision of a master student project.
-Provide state of the art knowledge in imaging and 3D analysis to PhD training course.
During the development of this project we have used novel and state of the art techniques. This has been crucial in order to generate a deep and high resolution knowledge of intestinal maturation process. In order to find the place of origin of intestinal stem cells we introduce fluorescent marks (lineage tracing) to specific cells. Then we followed them during time and we analysed the progeny of the labeled cells by 3-dimensional reconstructions of the foetal and adult intestine using confocal microscopy. In order to identify the cellular mechanism we have done live imaging of the fetal intestine. In order to identify the molecular mechanisms driving intestinal maturation we have use sequencing techniques in order to identify the transcriptome of the cells (Single cell sequencing, CAGE sequencing), the epigenetic profile (ATAC sequencing, Methyl-sequencing) and the 3-D configuration of the cellular nuclei (Hi-C sequencing). Finally we have challenged different foetal cell populations to describe their engraftment and regenerative potential. This project has paved the way in order to develop transplantations in humans, and the lab will pursue this way moving to the study of bigger animals and eventual clinical trials.
Figure illustrating the project relevance and the main technologies used