Senescence‐associated β‐galactosidase reveals the abundance of senescent CD8+ T cells in aging humans
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ABSTRACT:
PROVIDER: PRJNA714897 | ENA |
REPOSITORIES: ENA
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Project description:Aging leads to a progressive functional decline of the immune system, rendering the elderly increasingly susceptible to disease and infection. The degree to which immune cell senescence contributes to this decline remains unclear, however, since markers that label immune cells with classical features of cellular senescence accurately and comprehensively have not been identified. Using a second‐generation fluorogenic substrate for β‐galactosidase and multi‐parameter flow cytometry, we demonstrate here that peripheral blood mononuclear cells (PBMCs) isolated from healthy humans increasingly display cells with high senescence‐associated β‐galactosidase (SA‐βGal) activity with advancing donor age. The greatest age‐associated increases were observed in CD8+ T‐cell populations, in which the fraction of cells with high SA‐βGal activity reached average levels of 64% in donors in their 60 s. CD8+ T cells with high SA‐βGal activity, but not those with low SA‐βGal activity, were found to exhibit features of telomere dysfunction‐induced senescence and p16‐mediated senescence, were impaired in their ability to proliferate, developed in various T‐cell differentiation states, and had a gene expression signature consistent with the senescence state previously observed in human fibroblasts. Based on these results, we propose that senescent CD8+ T cells with classical features of cellular senescence accumulate to levels that are significantly higher than previously reported and additionally provide a simple yet robust method for the isolation and characterization of senescent CD8+ T cells with predictive potential for biological age.
2021-04-14 | GSE169009 | GEO
Project description: Not available
| S-EPMC8135084 | biostudies-literature
Project description:Noncoding RNAs include small transcripts, such as microRNAs and piwi-interacting RNAs, and a wide range of long noncoding RNAs (lncRNAs). Although many lncRNAs have been identified, only a small number of lncRNAs have been characterized functionally. Here, we sought to identify lncRNAs differentially expressed during replicative senescence. We compared lncRNAs expressed in proliferating, early-passage, 'young' human diploid WI-38 fibroblasts [population doubling (PDL) 20] with those expressed in senescent, late-passage, 'old' fibroblasts (PDL 52) by RNA sequencing (RNA-Seq). Numerous transcripts in all lncRNA groups (antisense lncRNAs, pseudogene-encoded lncRNAs, previously described lncRNAs and novel lncRNAs) were validated using reverse transcription (RT) and real-time, quantitative (q)PCR. Among the novel senescence-associated lncRNAs (SAL-RNAs) showing lower abundance in senescent cells, SAL-RNA1 (XLOC_023166) was found to delay senescence, because reducing SAL-RNA1 levels enhanced the appearance of phenotypic traits of senescence, including an enlarged morphology, positive β-galactosidase activity, and heightened p53 levels. Our results reveal that the expression of known and novel lncRNAs changes with senescence and suggests that SAL-RNAs play direct regulatory roles in this important cellular process. RNA was extracted from both young and senescent WI-38 cells and used for total RNA-Seq.
2016-02-08 | E-GEOD-77675 | biostudies-arrayexpress
Project description:The presence of senescent cells in the aging/degenerating human disc is now well-recognized. Senescent cells are viable, cannot divide, remain metabolically active and accumulate within the disc over time. Molecular analysis of senescent cells in tissue, however, offers a special challenge since there are no cell surface markers for senescence which would let one use fluorescence-activated cell sorting as a method for separating out senescent cells. Here we use a novel experimental design using laser capture microdissection to selectively separately harvest senescent and non-senescent annulus cells in paraffin-embedded tissue, and then compare their gene expression with microarray analysis. An initial in vitro study using cultured human annulus cells was first performed to test whether there was any difference in identification of senescent cells using the accepted histochemical methodology vs. the immunofluoresent identification of cells positive for senescence-associated-ß-galactosidase in control cells and cells induced into stress-induced premature senescence via hydrogen peroxide exposure. No statistically significant difference was found between the 2 methods. Laser capture microdissection was used to separately harvest senescent and non-senescent cells from 11 human annulus specimens, and microarray analysis was used to determine gene expression levels. Genes with established relationships to senescence were found to be significantly upregulated in senescent cells vs. non-senescent cells. Additional genes related to cytokines, cell proliferation, and other cell processes were also identified. Disc Tissue samples were obtained from surgical disc procedures performed on patients with herniated discs and degenerative disc disease. Tissue was fixed and paraffin embedded. Standard laser capture microdissection (LCM) techniques were used to collect senescent cells. Remaining non-senescent cells were scraped from the histology slide. Total RNA was isolated and analyzed via mircoarray. Gene expression from senescent cells was compared to non-senescent cells. Eight histological samples were used to obtain both senescent and non-senescent cells. From an additional 3 samples, only senescent cells were harvested.
2009-07-15 | E-GEOD-17077 | biostudies-arrayexpress
Project description:EGFR inhibitor Erlotinib treatment can induce NHBE into profound cell cycle arrest that include senescence and quiescence. Because senescent cells feature high senescence assocaited-beta-galactosidase(SA-β-gal) activty, which has been used as marker for FACS based collection of senescent and quiescent cells, respectively.
2018-07-27 | GSE100014 | GEO
Project description:Noncoding RNAs include small transcripts, such as microRNAs and piwi-interacting RNAs, and a wide range of long noncoding RNAs (lncRNAs). Although many lncRNAs have been identified, only a small number of lncRNAs have been characterized functionally. Here, we sought to identify lncRNAs differentially expressed during replicative senescence. We compared lncRNAs expressed in proliferating, early-passage, 'young' human diploid WI-38 fibroblasts [population doubling (PDL) 20] with those expressed in senescent, late-passage, 'old' fibroblasts (PDL 52) by RNA sequencing (RNA-Seq). Numerous transcripts in all lncRNA groups (antisense lncRNAs, pseudogene-encoded lncRNAs, previously described lncRNAs and novel lncRNAs) were validated using reverse transcription (RT) and real-time, quantitative (q)PCR. Among the novel senescence-associated lncRNAs (SAL-RNAs) showing lower abundance in senescent cells, SAL-RNA1 (XLOC_023166) was found to delay senescence, because reducing SAL-RNA1 levels enhanced the appearance of phenotypic traits of senescence, including an enlarged morphology, positive β-galactosidase activity, and heightened p53 levels. Our results reveal that the expression of known and novel lncRNAs changes with senescence and suggests that SAL-RNAs play direct regulatory roles in this important cellular process.
2016-02-08 | GSE77675 | GEO
Project description:The presence of senescent cells in the aging/degenerating human disc is now well-recognized. Senescent cells are viable, cannot divide, remain metabolically active and accumulate within the disc over time. Molecular analysis of senescent cells in tissue, however, offers a special challenge since there are no cell surface markers for senescence which would let one use fluorescence-activated cell sorting as a method for separating out senescent cells. Here we use a novel experimental design using laser capture microdissection to selectively separately harvest senescent and non-senescent annulus cells in paraffin-embedded tissue, and then compare their gene expression with microarray analysis. An initial in vitro study using cultured human annulus cells was first performed to test whether there was any difference in identification of senescent cells using the accepted histochemical methodology vs. the immunofluoresent identification of cells positive for senescence-associated-ß-galactosidase in control cells and cells induced into stress-induced premature senescence via hydrogen peroxide exposure. No statistically significant difference was found between the 2 methods. Laser capture microdissection was used to separately harvest senescent and non-senescent cells from 11 human annulus specimens, and microarray analysis was used to determine gene expression levels. Genes with established relationships to senescence were found to be significantly upregulated in senescent cells vs. non-senescent cells. Additional genes related to cytokines, cell proliferation, and other cell processes were also identified.
2009-07-15 | GSE17077 | GEO
Project description:<p><strong>BACKGROUND:</strong> Periodontal ligament mesenchymal stem cells (PDLSCs) are a promising cell resource for cell-based regenerative medicine in dentistry. PDLSCs inevitably acquire a senescent phenotype after prolonged in vitro expansion, and the key regulators of cells during replicative senescence remain unclear.</p><p><strong>METHODS:</strong> We cultured periodontal ligament stem cells to passages 4, 10 and 20 (P4, P10, P20). The senescent phenotypes, proliferation and migration ability of PDLSCs (P4, P10, P20) were detected, and non-targeted metabonomic sequencing was performed. We treated PDLSCs with AICAR and detected the expression of FOXO1, FOXO3a, FOXO6 and AMPK phosphorylation (p-AMPK) levels of replicating senescent PDLSCs to explore the correlation between the metabolic changes and the AMPK pathway.</p><p><strong>RESULTS:</strong> Immunofluorescence staining of γ-H2AX, β-galactosidase staining, cell scratch test and qPCR were performed to confirm the occurrence of replicative senescence in PDLSCs during passaging. Three groups of cells at passage 4 (P4), passage 10 (P10) and passage 20 (P20) were collected for non-targeted metabolomics analysis. Metabonomic sequencing showed that the metabolism of replicative senescence in PDLSCs varied significantly. In particular, the content of fatty acid metabolites decreased with senescence, including capric acid, stearic acid, myristic acid and dodecanoic acid. KEGG pathway analysis showed that the AMPK signaling pathway was closely related to AMP levels. The AMP:ATP ratio increased in senescent PDLSCs; however, the levels of p-AMPK and the profile of FOXO1 and FOXO3a, which are downstream of the AMPK signaling pathway, decreased with senescence. We treated PDLSCs with AICAR, an activator of the AMPK pathway, and the phosphorylated AMPK level at P20 PDLSCs was partially restored. </p><p><strong>CONCLUSION:</strong> In summary, our study suggests that the metabolic process of PDLSCs is active in the early stage of senescence, prefers to consume fatty acids, and is attenuated in the later stages of senescence. AMP accumulates in replicative senescent PDLSCs; however, the sensitivity of AMPK phosphorylation sites is impaired, causing senescent PDLSCs to fail to respond to changes in energy metabolism. Our findings provide a new basis for the clinical application of periodontal ligament stem cells.</p>
2022-11-15 | MTBLS4568 | MetaboLights
Project description:Abstract: Cellular senescence, an integral component of aging and cancer, arises in response to diverse triggers, including telomere attrition, macromolecular damage, and signaling from activated oncogenes. At present, senescent cells are identified by the combined presence of multiple traits, such as senescence-associated protein expression and secretion, DNA damage, and β-galactosidase activity; unfortunately, these traits are neither exclusively nor universally present in senescent cells. To identify robust shared markers of senescence, we have performed RNA-sequencing analysis across 8 diverse models of senescence triggered in human diploid fibroblasts (WI-38, IMR-90) and endothelial cells (HUVEC, HAEC) by replicative exhaustion, exposure to ionizing radiation or doxorubicin, and expression of the oncogene HRASG12V. The intersection of the altered transcriptomes revealed 47 RNAs consistently elevated and 26 RNAs consistently reduced across all senescence models, including many protein-coding mRNAs and some long noncoding RNAs. We propose that these shared transcriptome profiles will enable the identification of senescent cells in vivo, the investigation of their roles in aging and malignancy, and the development of strategies to target senescent cells therapeutically.
2019-06-12 | GSE130727 | GEO
Project description:Tripeptidyl peptidase II (TPPII) degrades N-terminal tripeptides from proteins and peptides. Studies in both human and mice have shown that TPPII deficiency is linked to cellular immune-senescence, lifespan regulation, and the aging process. However, the mechanism of how TPPII participates in these processes is less clear. In this study, we established a chemical probe-based assay and found that while the mRNA and protein levels of TPPII were not altered during senescence, its enzymatic activity was reduced in senescent human fibroblasts. We also showed that elevation of serine protease inhibitor serpinB2 reduced TPPII activity in senescent cells. Moreover, suppression of TPPII led to elevation of lysosomal contents as well as TPPI and -galactosidase activities, suggesting that the lysosome biogenesis is induced to compensate for the reduction of TPPII activity in senescent cells. Together this study discloses a critical role of the serpinB2/TPPII signaling pathway in proteostasis during senescence. Since serpinB2 level can be increased by a variety of cellular stresses, reduction of TPPII activity through activation of serpinB2 might represent a common pathway for cells to respond to different stress conditions.
2022-05-31 | PXD033808 | Pride