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      Human pluripotent stem cell-derived acinar/ductal organoids generate human pancreas upon orthotopic transplantation and allow disease modelling

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          Abstract

          Objective

          The generation of acinar and ductal cells from human pluripotent stem cells (PSCs) is a poorly studied process, although various diseases arise from this compartment.

          Design

          We designed a straightforward approach to direct human PSCs towards pancreatic organoids resembling acinar and ductal progeny.

          Results

          Extensive phenotyping of the organoids not only shows the appropriate marker profile but also ultrastructural, global gene expression and functional hallmarks of the human pancreas in the dish. Upon orthotopic transplantation into immunodeficient mice, these organoids form normal pancreatic ducts and acinar tissue resembling fetal human pancreas without evidence of tumour formation or transformation. Finally, we implemented this unique phenotyping tool as a model to study the pancreatic facets of cystic fibrosis (CF). For the first time, we provide evidence that in vitro , but also in our xenograft transplantation assay, pancreatic commitment occurs generally unhindered in CF. Importantly, cystic fibrosis transmembrane conductance regulator (CFTR) activation in mutated pancreatic organoids not only mirrors the CF phenotype in functional assays but also at a global expression level. We also conducted a scalable proof-of-concept screen in CF pancreatic organoids using a set of CFTR correctors and activators, and established an mRNA-mediated gene therapy approach in CF organoids.

          Conclusions

          Taken together, our platform provides novel opportunities to model pancreatic disease and development, screen for disease-rescuing agents and to test therapeutic procedures.

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          Most cited references 43

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          Pancreatic endoderm derived from human embryonic stem cells generates glucose-responsive insulin-secreting cells in vivo.

          Development of a cell therapy for diabetes would be greatly aided by a renewable supply of human beta-cells. Here we show that pancreatic endoderm derived from human embryonic stem (hES) cells efficiently generates glucose-responsive endocrine cells after implantation into mice. Upon glucose stimulation of the implanted mice, human insulin and C-peptide are detected in sera at levels similar to those of mice transplanted with approximately 3,000 human islets. Moreover, the insulin-expressing cells generated after engraftment exhibit many properties of functional beta-cells, including expression of critical beta-cell transcription factors, appropriate processing of proinsulin and the presence of mature endocrine secretory granules. Finally, in a test of therapeutic potential, we demonstrate that implantation of hES cell-derived pancreatic endoderm protects against streptozotocin-induced hyperglycemia. Together, these data provide definitive evidence that hES cells are competent to generate glucose-responsive, insulin-secreting cells.
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            Production of pancreatic hormone-expressing endocrine cells from human embryonic stem cells.

            Of paramount importance for the development of cell therapies to treat diabetes is the production of sufficient numbers of pancreatic endocrine cells that function similarly to primary islets. We have developed a differentiation process that converts human embryonic stem (hES) cells to endocrine cells capable of synthesizing the pancreatic hormones insulin, glucagon, somatostatin, pancreatic polypeptide and ghrelin. This process mimics in vivo pancreatic organogenesis by directing cells through stages resembling definitive endoderm, gut-tube endoderm, pancreatic endoderm and endocrine precursor--en route to cells that express endocrine hormones. The hES cell-derived insulin-expressing cells have an insulin content approaching that of adult islets. Similar to fetal beta-cells, they release C-peptide in response to multiple secretory stimuli, but only minimally to glucose. Production of these hES cell-derived endocrine cells may represent a critical step in the development of a renewable source of cells for diabetes cell therapy.
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              Reversal of diabetes with insulin-producing cells derived in vitro from human pluripotent stem cells.

              Transplantation of pancreatic progenitors or insulin-secreting cells derived from human embryonic stem cells (hESCs) has been proposed as a therapy for diabetes. We describe a seven-stage protocol that efficiently converts hESCs into insulin-producing cells. Stage (S) 7 cells expressed key markers of mature pancreatic beta cells, including MAFA, and displayed glucose-stimulated insulin secretion similar to that of human islets during static incubations in vitro. Additional characterization using single-cell imaging and dynamic glucose stimulation assays revealed similarities but also notable differences between S7 insulin-secreting cells and primary human beta cells. Nevertheless, S7 cells rapidly reversed diabetes in mice within 40 days, roughly four times faster than pancreatic progenitors. Therefore, although S7 cells are not fully equivalent to mature beta cells, their capacity for glucose-responsive insulin secretion and rapid reversal of diabetes in vivo makes them a promising alternative to pancreatic progenitor cells or cadaveric islets for the treatment of diabetes.
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                Author and article information

                Journal
                Gut
                Gut
                gutjnl
                gut
                Gut
                BMJ Publishing Group (BMA House, Tavistock Square, London, WC1H 9JR )
                0017-5749
                1468-3288
                March 2017
                15 September 2016
                : 66
                : 3
                : 473-486
                Affiliations
                [1 ]Department of Internal Medicine I, University Medical Center Ulm , Ulm, Germany
                [2 ]Department of Pediatrics I, Pediatric Infectiology and Immunology, Translational Genomics and Gene Therapy in Pediatrics, University of Tuebingen , Tuebingen, Germany
                [3 ]Medical Faculty, IZKF Computational Biology Research Group, RWTH Aachen University , Aachen, Germany
                [4 ]Medical Faculty, Department of Cell Biology, Institute for Biomedical Engineering, RWTH Aachen University , Aachen, Germany
                [5 ]Department of Medicine A, University Medicine, Ernst-Moritz-Arndt-University , Greifswald, Germany
                [6 ]Institute of Human Genetics, University Hospital , Magdeburg, Germany
                [7 ]Department of Pediatrics and Adolescent Medicine, University Medical Center Ulm , Ulm, Germany
                [8 ]Department of Internal Medicine I, Division of Medicine, Martin-Luther-University Halle-Wittenberg , Halle (Saale), Germany
                [9 ]Department of Biochemistry, Universidad Autónoma de Madrid, Instituto de Investigaciones Biomédicas “Alberto Sols” CSIC-UAM , Madrid, Spain
                [10 ]Institute of Neuroanatomy, Eberhard Karls University Tuebingen , Tuebingen, Germany
                Author notes
                [Correspondence to ] Prof Dr Alexander Kleger, Department of Internal Medicine I, University Medical Center Ulm, Albert-Einstein-Allee 23, Ulm 89081, Germany; alexander.kleger@ 123456uni-ulm.de

                SL and AK jointly supervised this work and contributed equally.

                Article
                gutjnl-2016-312423
                10.1136/gutjnl-2016-312423
                5534761
                27633923
                Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/

                This is an Open Access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See: http://creativecommons.org/licenses/by-nc/4.0/

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                1506
                1507
                Pancreas
                Original article
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                Gastroenterology & Hepatology

                cystic fibrosis, pancreas, stem cells

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