Evaluation of variability in human kidney organoids

Abstract

The utility of human pluripotent stem cell–derived kidney organoids relies implicitly on the robustness and transferability of the protocol. Here we analyze the sources of transcriptional variation in a specific kidney organoid protocol. Although individual organoids within a differentiation batch showed strong transcriptional correlation, we noted significant variation between experimental batches, particularly in genes associated with temporal maturation. Single-cell profiling revealed shifts in nephron patterning and proportions of component cells. Distinct induced pluripotent stem cell clones showed congruent transcriptional programs, with interexperimental and interclonal variation also strongly associated with nephron patterning. Epithelial cells isolated from organoids aligned with total organoids at the same day of differentiation, again implicating relative maturation as a confounder. This understanding of experimental variation facilitated an optimized analysis of organoid-based disease modeling, thereby increasing the utility of kidney organoids for personalized medicine and functional genomics.

Kidney organoid reproducibility across multiple human iPSC lines and diminished off target cells after transplantation revealed by single cell transcriptomics

“…1C, Supplementary Fig. 2), were found in D29 organoids, including melanoma-like cells (PMEL), SOX2-positive(+) neuronal precursors, STMN2+ neuron-like cells, and MYOG+ muscle-like cells, as reported previously (Phipson et al, 2019;Wu et al, 2018). A rare population of endothelial cells was also observed ( Fig.…”

“…To harness the full potential of iPSC derived kidney organoid technology, we must address critical unsolved questions about their reproducibility, faithfulness, and quality. First, we must establish organoid reproducibility: the comparability and range of variability in cellular composition and state between different iPSC lines from normal individuals across replicates and protocols (building on previous efforts to draw comparisons between iPSC and embryonic stem cell (ESC) derived organoids (Wu et al, 2018) or bulk RNASeq data comparing iPSC derived organoids (Phipson et al, 2019)). This is critically important, because individual patient iPSCs offer significant advantages over ESCs for precision medicine and drug development projects (Boreström et al, 2018;Burrows et al, 2016;Takasato et al, 2016b).…”

Reporter‐based fate mapping in human kidney organoids confirms nephron lineage relationships and reveals synchronous nephron formation

“…Multiple stromal populations were also identified, which expressed collagens COL1A1 and COL3A1, as well as kidney stromal markers DCN and CXCL12. We note the apparent absence of an endothelial cluster in this dataset, despite clear evidence for this cell type in previous studies [19][20][21], including in bioprinted organoids [preprint: 15].…”

Mouse embryonic stem cells self-organize into trunk-like structures with neural tube and somites

“…Our TLS already display remarkable reproducibility both at 166 the morphological and molecular level. Nevertheless, we realize the importance of further 167 integrated comparative analysis of single TLS at single-cell resolution, such as recently 168 published for brain and kidney organoids 37,38 . Finally, in vivo somites are formed in a rhythmic 169 process involving an oscillator -the segmentation clock 39 .…”