Landscape of transcription and expression regulated by DNA methylation related to age of donor and cell passage in adipose-derived mesenchymal stem cells.

Adipose-derived mesenchymal stem cells (ADSCs) are pluripotent stromal cells that can differentiate into a variety of cell types, including skin cells. High-throughput sequencing was performed on cells of different ages and cell passage, obtaining their methylation, mRNA expression, and protein profile data. The stemness of each sample was then calculated using the TCGAbiolinks package in R. Co-expression modules were identified using WGCNA, and a crosstalk analysis was performed on the corresponding modules. The ClusterProfile package was used for the functional annotation of module genes. Finally, the regulatory network diagram was visualized using the Cytoscape software. First, a total of 16 modules were identified, where 3 modules were screened that were most relevant to the phenotype. 29 genes were screened in combination of the RNA seq, DNA methylation seq and protein iTRAQ. Finally, a comprehensive landscape comprised of RNA expression, DNA methylation and protein profiles of age relevant ADSCs was constructed. Overall, the different omics of ADSCs were comprehensively analyzed in order to reveal mechanisms pertaining to their growth and development. The effects of age, cell passage, and stemness on the therapeutic effect of ADSCs were explored. Additionally, a theoretical basis for selecting appropriate ADSC donors for regenerative medicine was provided.

Multiomics global landscape of stemness-related gene clusters in adipose-derived mesenchymal stem cells.

BACKGROUND: Adipose-derived mesenchymal stem cells (AD-MSCs) are a type of stem cell that is abundant and widely used. The molecular characteristics of AD-MSCs from different passages from donors of different ages have not been well elucidated. METHODS: Six kinds of AD-MSCs ((E1, E2, E3, Y1, Y2, and Y3) with E denoting cells derived from an elderly patient, Y denoting cells derived from a young patient, and 1, 2, and 3 representing passages 3, 6, and 10) were obtained from human abdominal adipose tissue. We obtained the protein expression profile, the mRNA expression profile, the lncRNA expression profile, and the methylation profile of each kind of AD-MSC by sequencing. After calculating the stemness indices, genes related to stemness were extracted. The multiomics correlation analysis was performed in the stemness-related genes. In addition, short time-series expression miner (STEM) analysis was performed for all cell passages and donor ages. To further explore the biological functions of the stemness-related genes, we performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. Finally, the lncRNA-KEGG network and transcription factor (TF)-KEGG network were constructed based on the RNAInter database and TRRUST v2 database. RESULTS: The stemness of the Y1, E1, and Y2 cells was higher than that of the E2, Y3, and E3 cells. The stemness was the highest for Y1 cells and the lowest for E3 cells. STEM analysis showed that five stemness-related gene clusters were associated with the cell passages, and only one gene cluster was associated with age. The enrichment analysis results showed that the biological processes (BPs) and KEGG pathways were mainly involved in the proliferation, differentiation, and migration of cells. The global regulatory landscape of AD-MSCs was constructed: 25 TFs and 16 lncRNAs regulated 21 KEGG pathways through 27 mRNAs. Furthermore, we obtained a core stemness-related gene set consisting of ITGAV, MAD2L1, and PCNA. These genes were expressed at higher levels in Y1 cells than in E3 cells. CONCLUSION: The multiomics global landscape of stemness-related gene clusters was determined for AD-MSCs, which may be helpful for selecting AD-MSCs with increased stemness.

FGF2-induced PI3K/Akt signaling evokes greater proliferation and adipogenic differentiation of human adipose stem cells from breast than from abdomen or thigh.

In this study, human adipose stem cells were isolated from subcutaneous fat in the thigh (htASCs), abdomen (haASCs) and breast (hbASCs). Flow cytometry was used to detect cell surface markers, and an enzyme-linked immunosorbent assay was used to detect paracrine activity. Paracrine gene expression in the three cell types was examined using real-time qPCR, and adipogenic ability was assessed using Oil Red O staining. RNA from third-passage haASCs and hbASCs was sequenced. The results showed that the differentiation potential marker markers CD49d and CD54 were similar across hbASCs from 10 subjects. The hbASCs showed higher colony forming ability and expression of fibroblast growth factor-2, tissue inhibitor of metalloproteinase-1 and stromal cell derived factor-1 than htASCs and haASCs. Stimulating hbASCs with FGF2 promoted adipogenic differentiation, while treating the cells with the PI3K inhibitor LY294002 inhibited differentiation. These results suggest that the PI3K/Akt signaling pathway can promote proliferation and adipogenic differentiation of adipose stem cells, and that activation of this pathway by FGF2 may explain why hbASCs show greater proliferation and adipogenic differentiation than haASCs and htASCs.