title: Senescence-associated DNA methylation is stochastically acquired in subpopulations of mesenchymal stem cells.
abstract: Replicative senescence has a major impact on function and integrity of cell preparations. This process is reflected by continuous DNA methylation (DNAm) changes at specific CpG dinucleotides in the course of in vitro culture, and such modifications can be used to estimate the state of cellular senescence for quality control of cell preparations. Still, it is unclear how senescence-associated DNAm changes are regulated and whether they occur simultaneously across a cell population. In this study, we analyzed global DNAm profiles of human mesenchymal stem cells (MSCs) and human umbilical vein endothelial cells (HUVECs) to demonstrate that senescence-associated DNAm changes are overall similar in these different cell types. Subsequently, an Epigenetic-Senescence-Signature, based on six CpGs, was either analyzed by pyrosequencing or by bar-coded bisulfite amplicon sequencing. There was a good correlation between predicted and real passage numbers in bulk populations of MSCs (R2= 0.67) and HUVECs (R2= 0.97). However, when we analyzed the Epigenetic-Senescence-Signature in subclones of MSCs, the predictions revealed high variation and they were not related to the adipogenic or osteogenic differentiation potential of the subclones. Notably, in clonally derived subpopulations, the DNAm levels of neighboring CpGs differed extensively, indicating that these genomic regions are not synchronously modified during senescence. Taken together, senescence-associated DNAm changes occur in a highly reproducible manner, but they are not synchronously co-regulated. They rather appear to be acquired stochastically-potentially evoked by other epigenetic modifications.
authors: Helmholtz-Institute for Biomedical Engineering, RWTH Aachen University Medical School, 52074, Aachen, Germany.
authors: Julia
authors: J
authors: Franzen
authors: Center for Molecular Medicine (CMMC), University of Cologne, 50931, Cologne, Germany.
authors: Anne
authors: A
authors: Zirkel
authors: Genomics Core Facility, European Molecular Biology Laboratory (EMBL), 69117, Heidelberg, Germany.
authors: Jonathon
authors: J
authors: Blake
authors: Department for Orthopedics, RWTH Aachen University Medical School, 52074, Aachen, Germany.
authors: Björn
authors: B
authors: Rath
authors: Genomics Core Facility, European Molecular Biology Laboratory (EMBL), 69117, Heidelberg, Germany.
authors: Vladimir
authors: V
authors: Benes
authors: Center for Molecular Medicine (CMMC), University of Cologne, 50931, Cologne, Germany.
authors: Argyris
authors: A
authors: Papantonis
authors: Helmholtz-Institute for Biomedical Engineering, RWTH Aachen University Medical School, 52074, Aachen, Germany.
authors: Wolfgang
authors: W
authors: Wagner
mesh_terms: D001483:Base Sequence; D002454:Cell Differentiation / Q000235:genetics; D016922:Cellular Senescence / Q000235:genetics*; D002999:Clone Cells; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation / Q000235:genetics*; D044127:Epigenesis, Genetic; D061307:Human Umbilical Vein Endothelial Cells / Q000378:metabolism; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D013269:Stochastic Processes; D013447:Sulfites / Q000378:metabolism
chemical_list: D013447:Sulfites; C042345:hydrogen sulfite
keywords: DNA methylation; bisulfite sequencing; epigenetics; human umbilical vein endothelial cell; mesenchymal stem cell; senescence