Project description:Here, we used single cell RNA sequencing of iPSC17 WT 7B2-derived day 10 lung spheroids, unsorted 3-, 6-, and 10-week lung progenitor organoids (LPOs) and induced bud tip progenitor organoids (iBTOs) sorted from 4- and 10-week LPOs, sequenced 4 weeks post-sort. All cultures were grown in serum-free basal media supplemented with WNT activator CHIR99021, FGF7, and all-trans retinoic acid (‘3F media’).
Project description:Underdeveloped lungs are the primary cause of death in premature infants, however, little is known about stem and progenitor cell maintenance during human lung development. In this study, we have identified that FGF7, Retinoic Acid and CHIR-99021, a small molecule that inhibits GSK3 to activate Wnt signaling, support in vitro maintenance of primary human fetal lung bud tip progenitor cells in a progenitor state. Furthermore, these factors are sufficient to derive a population of human bud tip-like progenitor cells in 3D organoid structures from human pluripotent stem cells (hPSC). Functional studies showed that hPSC-derived bud tip progenitor organoids do not contain any mesenchymal cell types, maintain multilineage potential in vitro and are able to engraft into the airways of injured mice and respond to systemic factors. We performed RNA-sequencing to assess the degree of similarity in global gene expression profiles between the full human fetal lung (59-127 days gestation), isolated human fetal bud tip progenitors, organoids grown from primary fetal bud tip progenitors, and hPSC-derived bud tip organoids. Results showed that hPSC-derived organoids have molecular profiles similar to organoids generated from primary human fetal lung tissue. Gene expression differences between hPSC-derived bud tip organoids and fetal progenitor organoids may be related to the presence of contaminating mesenchymal cells in primary cultures. hPSC-derived bud tip organoids are generated from a well-defined human cell sources, offering a distinct advantage over rare primary tissue as a means to study human specific lung development, homeostasis and disease.<br>Sample Nomenclature - Description<br> -------------------------------------------------------------------------<br> Peripheral fetal lung the distal/peripheral portion of the fetal lung (i.e., distal 0.5 cm) was excised from the rest of the lung using a scalpel. This includes all components of the lung (e.g., epithelial, mesenchymal, vascular). <br>Isolated fetal bud tip the bud peripheral portion of the fetal lung was excised with a scalpel and subjected to enzymatic digestion and microdissection. The epithelium was dissected and separated from the mesenchyme, but a small amount of associated mesenchyme likely remained. <br>Fetal progenitor organoid 3D organoid structures that arose from culturing isolated fetal epithelial bud tips. <br>Foregut spheroid 3D foregut endoderm structure as described in Dye et al. (2015). Gives rise to patterned lung organoid (PLO) when grown in 3F medium. <br> Patterned lung organoid (PLO) lung organoids that were generated by differentiating hPSCs, as described throughout the manuscript. <br> Bud tip organoid organoids derived from PLOs, enriched for SOX2/SOX9 co-expressing cells, and grown/passaged in 3F medium.
Project description:Retinal organoids samples that derived from human embryonic stem cells were analyzed by single-cell RNA sequencing. Two samples at different differentiation stages (day57 and day 171) were included in this study for cell type comparison.
Project description:Here, we used a lentiviral-based barcoding method (CellTagging) to lineage trace progenitor cells in a human lung organoid model. 21-day cultures were transduced for 7 days, fluorescence activated cell sorted for GFP+ expression, and regrown as sparsely plated cells for additional 30 days before sequencing. Cultures were grown in serum-free basal media supplemented with FGF-10, A-8301, NOGGIN, and Y-27632 (‘DSI media’).
Project description:Patient-derived organoids have shown great promise as models of human tumors. However, a detailed single-cell description of the heterogeneity of these models is still lacking and limits their future clinical use. Here we performed a 10X single-cell RNA sequencing experiment to study the single-cell phenotype of breast cancer patient-derived organoids from 6 donors.
Project description:Organoids were generated from H9 cells. Single cells were sorted from 4-month-old brain organoids differentiated using the telencephalon organoids protocol.