Schulz A, Brockmann E, Zentgraf M, Baur A, Uebe S, Ekici AB, Dedden M, Zundler S, Thiel C (2026)
Publication Type: Journal article
Publication year: 2026
Book Volume: 17
Article Number: 276
Journal Issue: 1
DOI: 10.1186/s13287-026-05223-x
Background: Urine-derived stem cells (USCs) represent an accessible and non-invasive cell source with reported chondrogenic differentiation potential. However, the cellular heterogeneity and transcriptional dynamics underlying USC differentiation remain incompletely understood, limiting their translational interpretation. Methods: We combined functional differentiation assays with single-cell RNA sequencing to characterize USC differentiation at both phenotypic and transcriptional levels. Chondrogenic and osteogenic differentiation were assessed using histological staining, quantitative PCR, and three-dimensional spheroid cultures. Single-cell transcriptomic analysis was performed on integrated datasets of undifferentiated and differentiated USCs, followed by pseudotime trajectory inference and mapping to a human cartilage reference atlas. Results: Chondrogenic induction resulted in reproducible acquisition of cartilage-associated features, including glycosaminoglycan-rich extracellular matrix deposition, increased expression of SOX9, and formation of aggrecan-positive spheroids. In this donor, single-cell analysis mapped an inferred differentiation trajectory from proliferative states towards differentiated populations, although the fine-grained pseudotemporal ordering was sensitive to analytical choices and is therefore interpreted qualitatively. Along this inferred trajectory, we identified a candidate transient transcriptional state associated with elevated CDH1 expression and epithelial-like aggregation features. Probabilistic mapping to a human cartilage reference atlas indicated that overall mapping confidence was low (median prediction score 0.34) and that only a minority of cells showed confident transcriptional similarity (prediction score ≥ 0.5) to mature/articular cartilage-associated reference states (7.8% of all cells and 17.6% of chondrogenically induced cells). This confident similarity was concentrated in a few clusters at the differentiated end of the trajectory rather than representing the bulk of the culture, and label-transfer confidence was not equated with chondrocyte identity. Despite this enrichment, differentiated populations exhibited transcriptional heterogeneity, including subsets of cells associated with hypertrophic, fibrocartilage-like, and contractile gene programmes, indicating the presence of multiple differentiation trajectories. Conclusions: This single-donor proof-of-concept study suggests that USC differentiation may involve a candidate transient, aggregation-associated transcriptional state accompanied by CDH1 expression and gives rise to heterogeneous lineage-associated outcomes, with only a minority of cells acquiring confident transcriptional similarity to mature cartilage. Because these observations derive from one donor, they should be interpreted as hypothesis-generating and require validation across independent donors before donor-independent or translational conclusions for cartilage regeneration can be drawn. These findings nonetheless provide a single-cell resolution framework for future multi-donor validation of USC differentiation and its inherent transcriptional heterogeneity.
APA:
Schulz, A., Brockmann, E., Zentgraf, M., Baur, A., Uebe, S., Ekici, A.B.,... Thiel, C. (2026). A single-donor proof-of-concept single-cell analysis maps heterogeneous differentiation trajectories toward cartilage-like states in human urine-derived stem cells. Stem Cell Research and Therapy, 17(1). https://doi.org/10.1186/s13287-026-05223-x
MLA:
Schulz, Alexander, et al. "A single-donor proof-of-concept single-cell analysis maps heterogeneous differentiation trajectories toward cartilage-like states in human urine-derived stem cells." Stem Cell Research and Therapy 17.1 (2026).
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