Epigenetic biomarkers to track differentiation of pluripotent stem cells

Quality control of induced pluripotent stem cells remains a challenge. For validation of the pluripotent state, it is crucial to determine trilineage differentiation potential toward endoderm, mesoderm, and ectoderm. Here, we report GermLayerTracker, a combination of site-specific DNA methylation (DNAm) assays that serve as biomarker for early germ layer specification. CG dinucleotides (CpGs) were identified with characteristic DNAm at pluripotent state and after differentiation into endoderm, mesoderm, and ectoderm. Based on this, a pluripotency score was derived that tracks reprogramming and may indicate differentiation capacity, as well as lineage-specific scores to monitor either directed differentiation or self-organized multilineage differentiation in embryoid bodies. Furthermore, we established pyrosequencing assays for fast and cost-effective analysis. In the future, the GermLayerTracker could be used for quality control of pluripotent cells and to estimate lineage-specific commitment during initial differentiation events.

Automated summary

Quality control of induced pluripotent stem cells is challenging, and determining trilineage differentiation potential is crucial for validating their pluripotent state. In this study, GermLayerTracker, a combination of site-specific DNA methylation assays, was developed as a biomarker for early germ layer specification. Specific CG dinucleotides (CpGs) with characteristic DNA methylation patterns were identified in the pluripotent state and during differentiation into endoderm, mesoderm, and ectoderm. The GermLayerTracker can be used for quality control of pluripotent cells and to estimate lineage-specific commitment during initial differentiation events.