Project description:These files represent single cell RNA-Seq data generated on a 10x Chromium genomics platform from bioprinted iPSC-derived human kidney organoids differentiated according to our published protocol (Takasato et al., Nature Protocols 2016). The data contains >2000 cells that passed our QC. Data was used to confirm that bioprinted organoids contain a similar cellular composition as standard manually produced organoids - See https://www.biorxiv.org/content/10.1101/505396v1.
Project description:We compared kidney organoids generated manually ('Man') to those generated by bioprinting single cell deposition ('R0') and thin bioprinted lines ('R40').
Project description:Trophoblast organoids offer a unique opportunity to study mechanisms orchestrating placental growth and development during pregnancy. However, many organoid cultures rely on extracellular matrix reagents that are highly variable and unable to be tuned to reflect in vivo tissues. Here we describe the first bioprinted placental organoid model, generated using first trimester trophoblast cell line, ACH-3P, and a synthetic polyethylene glycol matrix. Bioprinted organoids were compared comprehensively to classic Matrigel-embedding using functional assays, immunofluorescence microscopy, transcriptomic and proteomic analyses. Organoids differentiated spontaneously from cytotrophoblasts into two major subtypes: extravillous trophoblasts (EVTs) and syncytiotrophoblasts (STBs), with bioprinted organoids driven towards EVT differentiation. Bioprinted organoids were exposed to inflammation and treated with aspirin or metformin to assess their effects on trophoblast organoid formation and viability. Further, we reversed the inside-out architecture of ACH-3P organoids by suspension culture. Organoid suspension caused STBs to form a syndecan-1+ outer layer on the periphery of organoids, reflecting placental tissue. Here, we present an alternative trophoblast organoid model with further tuning potential to reflect the placental microenvironment in physiological and pathological pregnancies.
Project description:Trophoblast organoids offer a unique opportunity to investigate mechanisms orchestrating placental growth and development during pregnancy. However, many organoid cultures rely on extracellular matrix reagents that are highly variable and unable to be tuned to reflect different tissues. Here we describe a new bioprinted placental organoid model using immortalised first trimester trophoblast cell line, ACH-3P, and a synthetic polyethylene glycol matrix. Bioprinted organoids were comprehensively compared to classical Matrigel-embedding using functional assays, immunofluorescence microscopy, transcriptomic and proteomic analysis. Organoids spontaneously differentiated from cytotrophoblasts into the two major subtypes: extravillous trophoblasts (EVTs) and syncytiotrophoblasts (STBs), with bioprinted organoids driven towards EVT differentiation. Bioprinted organoids were exposed to inflammation and treated with aspirin or metformin to assess their effects on trophoblast organoid formation and viability. Further, we reversed the inside-out architecture of ACH-3P organoids by transferring immature organoids to suspension culture. Organoid suspension caused STB to form a syndecan-1+ outer layer on the periphery of organoids, reflecting placental tissue. We present an alternative trophoblast organoid model with potential for further tuning to accurately reflect the placental microenvironment and provide more reliable insights into early placental development.
Project description:We analyzed single cell transcriptomes over 80,000 cells isolated from 65 organoids differentiated from iPSCs and ESCs using two different protocols. We find that both protocols generate kidney organoids that contain a diverse range of kidney cells at differing ratios as well as non-renal cell types. We reconstructed lineage relationships during organoid differentiation through pseudotemporal ordering, and identified transcription factor networks associated with fate decisions. When comparing to adult human kidney, we reveal immaturity of all kidney organoid cell types. These results define impressive kidney organoid cell diversity, identify incomplete differentiation as a major roadblock for current directed differentiation protocols and provide a human adult kidney snRNA-seq dataset against which to benchmark future progress.
Project description:Bulk RNA-seq comparison of kidney organoids bioprinted in 3 different conformations with varying starting cell densities. Density is dictated by the ratio of bioprinter tip movement to the amount of extrusion, where higher ratios spread cells over a larger surface area. We compare organoids printed with no movement ('blob', ratio 0) to those with moderate ('line 3', ratio 20) or high movement ('line 1', ratio 40).
Project description:Absence of WT1 during kidney organoid development from human induced pluripotent stem cells (iPSCs) induces hallmarks of Wilms tumorigenesis. To define underlying transcriptional alterations and similarities to human patients, we performed timecourse RNA-seq of kidney organoid development from control iPSCs (control, not edited) and in the absence of WT1. Two timepoints for knockout (KO) of WT1 were investigated: In iPSCs (KO in iPSCs), and between day 4 and day 7 of organoid formation (KO d4-7).