Project description:To study safety and stability of kidney organoids, we studied the presence of residual iPSC in the kidney organoids. We also analyzed their potential for malignant transformation in a teratoma assay and following long-term subcutaneous implantation in an immune deficient mouse model.
Project description:Induced pluripotent stem cell (iPSC)-derived kidney organoids are a potential tool for the regeneration of kidney tissue. They represent an early stage of nephrogenesis and have been shown to successfsully vascularize and mature further in vivo. However, there are concerns regarding the long-term safety and stability of iPSC derivatives. Specifically, the potential for tumorigenesis may impede the road to clinical application. To study safety and stability of kidney organoids, we analyzed their potential for malignant transformation in a teratoma assay and following long-term subcutaneous implantation in an immune-deficient mouse model. We did not detect fully functional residual iPSCs in the kidney organoids as analyzed by gene expression analysis, single-cell sequencing and immunohistochemistry. Accordingly, kidney organoids failed to form teratoma. Upon long-term subcutaneous implantation of whole organoids in immunodeficient IL2Ry-/-RAG2-/- mice, we observed tumor formation in 5 out of 103 implanted kidney organoids. These tumors were composed of WT1+CD56+ immature blastemal cells and showed histological resemblance with Wilms tumor. No genetic changes were identified that contributed to the occurrence of tumorigenic cells within the kidney organoids. However, assessment of epigenetic changes revealed a unique cluster of differentially methylated genes that were also present in undifferentiated iPSCs. We discovered that kidney organoids have the capacity to form tumors upon long-term implantation. The presence of epigenetic modifications combined with the lack of environmental cues may have caused an arrest in terminal differentiation. Our results indicate that the safe implementation of kidney organoids should exclude the presence of pro-tumorigenic methylation in kidney organoids.
Project description:Expression profiling of Wilms tumour samples, which identified molecular signatures of the ureteric bud and collecting duct as well as those of the proximal and distal tubules in triphasic histology tumours. These observations indicate Wilms tumours can arise from a precursor cell capable of generating the entire kidney, such as the cells of the intermediate mesoderm from which both the metanephric mesenchym and ureteric bud are derived.
Project description:Human induced pluripotent stem (iPS) cells are capable of differentiating into derivatives of the three embryonic germ layers both in vitro and in vivo. To date the the molecular differences between teratoma-forming cells and non-teratoma-forming cells has not been analysed. A cell line, B1, bears typical ES cell-like morphology, expression of pluripotency-associated genes, and in vitro pluripotency capacity, but fails to form teratomas after subcutaneously injected into immune-deficient mice based on histological analysis. Besides histological analysis, we characterized the tumors derived from line B1, and teratomas derived from bona fida iPS and ES (line H1) cells respectively, using microarray-based gene expression analysis. The expression levels of pluripotency-associated markers in B1 cells were comparable to that in iPS and ES cells, while the complexity of tissue expression commitment was decreased upon spontaneous differentiation of B1 cells as compared to iPS and ES cells. Total RNA obtained from HFF1 (human foreskin fibroblast) cells, line B1, iPS-A4, iPS-B4 and ES (line H1) cells, and their derived tumors in immune-deficient mice.
Project description:Human induced pluripotent stem (iPS) cells are capable of differentiating into derivatives of the three embryonic germ layers both in vitro and in vivo. To date the the molecular differences between teratoma-forming cells and non-teratoma-forming cells has not been analysed. A cell line, B1, bears typical ES cell-like morphology, expression of pluripotency-associated genes, and in vitro pluripotency capacity, but fails to form teratomas after subcutaneously injected into immune-deficient mice based on histological analysis. Besides histological analysis, we characterized the tumors derived from line B1, and teratomas derived from bona fida iPS and ES (line H1) cells respectively, using microarray-based gene expression analysis. The expression levels of pluripotency-associated markers in B1 cells were comparable to that in iPS and ES cells, while the complexity of tissue expression commitment was decreased upon spontaneous differentiation of B1 cells as compared to iPS and ES cells.
Project description:Skeletal muscle stem cells are essential to muscle homeostasis and regeneration after injury. An attractive approach to obtain these cells is via differentiation of pluripotent stem cells (PSCs). We have recently reported that teratomas derived from mouse PSCs are a rich source of skeletal muscle stem cells. Here, we showed that the teratoma formation method is also capable of producing skeletal myogenic progenitors from human PSCs. Using single-cell transcriptomics, we discovered multiple lineages in human PSC-derived teratomas. Interestingly, we observed several distinct skeletal myogenic subpopulations. Trajectory analysis revealed that these subpopulations represented progressive stages of skeletal myogenic development. We further discovered that ERBB3 and CD82 are effective surface markers for prospective isolation of the skeletal myogenic lineage in human PSC-derived teratomas. Therefore, teratoma formation provides an accessible model for obtaining human skeletal myogenic progenitors from PSCs.