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Unveiling Aging Heterogeneities in Human Dermal Fibroblasts via Nanosensor Chemical Cytometry

ABSTRACT: To investigate aging heterogeneity in tissue-regeneration cells, we conducted RNA-seq analysis of human dermal fibroblasts (hDFs) at two distinct aging stages: passage 5 (p5) and passage 15 (p15). hDFs were cultured under standard conditions, and total RNA was extracted for sequencing. Transcriptomic profiling was performed to identify differentially expressed genes associated with cellular aging. This RNA-seq dataset was used to validate the phenotypic changes observed via a label-free, nondestructive single-cell analytics platform called nanosensor chemical cytometry (NCC). The integrated analysis revealed that transcriptomic aging signatures correspond with aging phenotypes such as changes in cell size, shape, refractive index, and H2O2 efflux. This dataset provides insight into the molecular mechanisms underlying cell aging and supports the development of predictive models for therapeutic performance in regenerative medicine.

ORGANISM(S): Homo sapiens

PROVIDER: GSE297406 | GEO | 2025/06/19

REPOSITORIES: GEO

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Project description:Background: Human bone marrow mesenchymal stem cells (MSCs) expanded in vitro exhibit not only a tendency to lose their proliferative potential, homing ability and telomere length but also genetic or epigenetic modifications such as DNA methylation and the noncoding RNA-mediated mechanism. This results in the senescence of MSCs. We compared differential methylation patterns of genes and miRNAs between early-passage and late-passage cells and estimated the relationship between senescence and DNA methylation patterns. Genomic DNA of MSCs cultured at passage 5 (P5) and passage 15 (P15) was enriched using methylated DNA immunoprecipitation assays, and DNA methylation changes due to long-term culture of MSCs were analyzed using Human 2.1M Deluxe Promoter Arrays (NimbleGen). Results: When we analyzed the methylation differences between P5 and P15 more than twice, 3,338 genes showed more than two-fold higher methylation at P5 than P15, whereas 4,670 genes showed more than two-fold higher methylation at P15 than P5. When we examined hypermethylated genes (methylation peak ≥ 2) at P5 or P15, 2,739 genes, including those related to fructose and mannose metabolism and calcium signaling pathways, and 2,587 genes, including those related to DNA replication, cell cycle and the PPAR signaling pathway, were hypermethylated at P5 and P15, respectively. There was common hypermethylation of 1,205 genes at both P5 and P15. In addition, genes that were hypermethylated at P5 (CPEB1, GMPPA, CDKN1A, TBX2, SMAD9 and MCM2) showed lower mRNA expression than did those hypermethylated at P15, whereas genes that were hypermethylated at P15 (MAML2, FEN1 and CDK4) showed lower mRNA expression than did those that were hypermethylated at P5, demonstrating that hypermethylation at DNA promoter regions inhibited gene expression and that hypomethylation increased gene expression. In the case of hypermethylation on miRNA, 27 miRNAs were hypermethylated at P5, whereas 44 miRNAs were hypermethylated at P15. Conclusion: These results show that hypermethylation increases at genes related to DNA replication, cell cycle and adipogenic differentiation due to long-term culture, which may in part affect MSC senescence.

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Unveiling Aging Heterogeneities in Human Dermal Fibroblasts via Nanosensor Chemical Cytometry

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