Speaker
Description
Induced pluripotent stem cell (iPSC) models combined with single-cell transcriptomics enable the study of cellular consequences resulting from human disease alleles, particularly in early-onset developmental disorders. A significant bottleneck lies in the scalability of iPSC-based experiments. Pooled cell culture or ‘cell villages’ has emerged as a key strategy to enable large-scale studies. However, achieving a balanced representation of cell lines within villages remains challenging, likely due to differences in growth dynamics, non-cell autonomous effects and intrinsic variations in developmental signaling pathways during early embryonic lineages.
To better understand the factors that cause donor imbalance in 2D iPSC villages, we systematically evaluated the effects of different experimental and cell-line related factors on the composition of eight villages (5-18 donors each) differentiated to cortical neurons. We also assessed donor imbalance in the endoderm lineage by differentiating two of the villages to a pancreatic fate, and maintained them in pluripotency as a baseline. All villages included wild type, patient-derived and engineered mutant lines, and their donor composition was deconvoluted using natural genetic variation from either single-cell RNA-seq reads or low-coverage whole-genome sequencing.
We observed that specific cell lines consistently become dominant in villages at neuronal precursor (NPC) stage (50.4% representation), regardless of village size, and were more likely to be female-derived lines (p=0.018). Comparing the transcriptomic profiles of lines differentiated individually and in pools, we confirmed donor identity was preserved at NPC stage. We also found significant shifts in donor representation between neuronal and pancreatic fates, indicating pool imbalance was not constant across cellular lineages. This suggests that pool imbalance arises immediately upon induction due to donor-intrinsic variation in developmental cues that affect fate commitment programs differently. Our future aim is to mitigate donor imbalance in cell villages to create more accurate and accessible disease models for neurodevelopment disorders.
| Author(s) | *Pau Puigdevall1, Benjamin Potter1, Emma Ryhänen2, Adithi Sundaresh1, Kavya Kalpana Ganesh1, Robin Forsén1, Riina Lampela1, Timo Otonkoski2,3, Diego Balboa2, Helena Kilpinen1,4 |
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| Affiliation(s) | "1. Helsinki Institute of Life Science (HiLIFE), University of Helsinki, Finland 2. Stem Cells and Metabolism Research Program, University of Helsinki, Finland 3. Department of Pediatrics, Helsinki University Hospital and University of Helsinki, Finland 4. Faculty of Biological and Environmental Sciences, University of Helsinki, Finland" |
