Gazdhar, Amiq

Geiser, Thomas

Ozan, Vedat Burak

Tschirren, Anna-Barbara

English

Bern Open Repository and Information System (BORIS)

source: https://doi.org/10.48350/169202 | downloaded: 2.8.2022iPSCs were grown and differentiated intoalveolar lung organoids using different cocktailsof growth factors and cytokines; iPSCs weredifferentiated to endothelial cells by chemicalinduction. Each stage of differentiation wasvalidated via flow cytometry, microscopy andrelative gene expression analysis.Induced Pluripotent Stem Cell Derived Alveolar Lung OrganoidsVedat Burak Ozan1,2, Anna-Barbara Tschirren1,2, Amiq Gazdhar1,2, Thomas Geiser1,21Department of Pulmonary Medicine, University Hospital Bern, 2Department for BioMedical Research, University of BernPoster Number: P06Idiopathic Pulmonary Fibrosis• Chronic and progressive lung disease• Causes fibrosis• High morbidity and mortality• Main symptoms• Shortness of breath• Dry cough• Can lead to further complications• Lung cancer• Pulmonary emboli• Pulmonary hypertensionThere is no cure, only limited treatment options.Introduction Methods ResultsiPSCs cultured in defined media were firstdifferentiated into Definitive Endoderm;media was changed directing the cellstowards Anterior Foregut Endoderm(AFE).AFE were differentiated into Alveolar LungOrganoids in two stages with branchingstructures emerging in matrigel after d35,and maturing after d40. Mature ALOs hadAECI and AECII markers with multiple celltypes.iPSC were separately differentiated viachemical induction into endothelial cells asvalidated by CD31 staining on the endothelialcells.ConclusioniPSC differentiate into self organizing alveolarorganoids that have both alveolar epithelialtype I and alveolar epithelial type II cellswhen grown in matrigel in defined media.Further investigations are ongoing to combine alveolar epithelial cells and endothelial cells into a complex organoid.ReferencesTamò, L., et al., Generation of an alveolar epithelial type II cell line from induced pluripotent stem cells. American Journal of Physiology-Lung Cellular and Molecular Physiology, 2018. 315(6): p. L921-L932.Geiser, T., Idiopathic pulmonary fibrosis--a disorder of alveolar wound repair? Swiss Med Wkly, 2003. 133(29-30): p. 405-11.Nishihara, H., et al., Advancing human induced pluripotent stem cell-derived blood-brain barrier models for studying immune cell interactions. FASEB J. 2020 Dec;34(12):16693-16715. doi: 10.1096/fj.202001507RR.Dutta, D., I. Heo, and H. Clevers, Disease modeling in stem cell-derived 3D organoid systems. Trends in molecular medicine, 2017. 23(5): p. 393-410..Illustrations created with BioRender (Fig 2, 3 & 7)Figure 1.IPF Lung(Dr. K Watts)Our goal is to create a complex in vitro organoid model to address the lack of representative models.Figure 2. Alveolar Lung OrganoidCulture SchemeFigure 5. Endothelial Tube Formation Assay on Matrigel on iPSC derived Endothelial Cells after 6 hoursBrightfield image of Definitive Endoderm on Day 4CXCR4 Expression of DEBrightfield image of Anterior Foregut Endoderm on Day 8Sox2 Expression of AFEBrightfield image of Lung Organoids Day 25Brightfield image of Lung Organoids Day 42OCT4,SOX2, DAPI Staining on iPSCs(orange, green and blue respectively)Figure 4. Validation of iPSC Derived Endothelial Cells. a) CD31 expression ofEndothelial Cells after sorting Endothelial Progenitor Cells, b) iPSC DerivedEndothelial Cells after one round of selective passaging, c) Immunofluorescenceimage of Endothelial Cells with von Willebrand factor (Red) and DAPI (Blue)Figure 3. Alveolar Lung Organoid Differentiation. a) iPSC, b) Definitive Endoderm, c)Anterior Foregut Endoderm, d) Alveolar Lung OrganoidsFigure 6. Immunofluorescence image of Alveolar Lung Organoid. SP-C (Green), SP-B (Red), DAPI (Blue)a) b) c) d)a) b) c)iPSC Definitive EndodermAnterior Foregut Endoderm Alveolar Lung Organoids4 days 4 days ~40 daysFigure 7. qPCR results showing mRNA expression of Alveolar Epithelial Type I and Type II marker expression in mature Alveolar Lung Organoids,

Induced pluripotent stem cell derived alveolar lung organoids

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Induced pluripotent stem cell derived alveolar lung organoids

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