Proteome analysis of serial-cultured human iPSC-derived MSC with lamin A/C mutation
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo Sapiens
SUBMITTER: 
Liangge He
PROVIDER: PXD037639 | iProX | Fri Oct 21 00:00:00 BST 2022
REPOSITORIES: iProX
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Project description:Sirtuin 3 (SIRT3) is an NAD+-dependent deacetylase involved in various physiological and pathological processes. However, the role of SIRT3 in regulating human stem cell senescence remains largely unknown. Here, we observed the downregulated expression of SIRT3 in senescent human mesenchymal stem cells (hMSCs). SIRT3 deficiency accelerated cellular senescence in hMSCs, along with compromised nuclear integrity, loss of heterochromatin and increased DNA damage. These aging-associated nuclear defects were attenuated by the reintroduction of SIRT3. Mechanistic studies demonstrated the interaction of SIRT3 with nuclear envelope proteins and heterochromatin-associated proteins. Further findings revealed that SIRT3 deficiency led to the loss of lamina-associated domains (LADs) from the nuclear lamina, increased chromatin accessibility and aberrant transcription of repetitive sequences. Meanwhile, the overexpression of nuclear-localized SIRT3 rescued the senescence phenotypes. Taken together, our study reveals a novel role of nuclear SIRT3 in stabilizing heterochromatin and counteracting hMSC senescence, which may provide new clinical therapeutic targets to ameliorate aging-related diseases.
2021-09-09 | PXD023479 | Pride
Project description:To determine miRNA expression changes during in vitro senescence of mesenchymal stem cells (MSC) we have analyzed differential expression of the corresponding early passage (P2) and senescent passage (PX). Keywords: miRNA, senescence, mesenchymal stromal cells, mesenchymal stem cells, MSC
2008-07-29 | GSE9664 | 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:Hutchinson Gilford Progeria Syndrome (HGPS) is a rare, sporadic genetic disease caused by mutations in the nuclear lamin A gene. In most cases the mutation creates an efficient donor-splice site that generates an altered transcript encoding a truncated lamin A protein, progerin. In vitro studies have indicated that progerin can disrupt nuclear function. HGPS affects mainly mesenchymal lineages but the shortage of patient material has precluded a tissue-wide molecular survey of progerin’s cellular impact. We report here a new, induced pluripotent stem cell (iPSC)-based model for studying HGPS. HGPS dermal fibroblasts were reprogrammed into iPSC lines using a cocktail of the transcription factor genes, OCT4, SOX2, KLF4 and C-MYC. The iPSC cells were differentiated into neural progenitors (NPs), endothelial cells (ECs), fibroblast-like cells and mesenchymal stem cells (MSCs). Progerin levels in the different cell types followed the pattern MSC≥ fibroblast>EC>>NP. Functionally, we detected a major impact of progerin on MSC function. We show that HGPS-MSCs are vulnerable to the ischemic conditions found in a murine hind limb recovery model and an in vitro hypoxia assay, as well as showing enhanced sensitivity in a serum starvation assay. Since there is widespread consensus that MSCs reside in low oxygen niches in vivo, we propose that these conditions lead to an accelerated depletion of the MSC pool in HGPS patients with consequent accretion of mesenchymal tissue. Analysis of iPSCs, hESCs and parental fibroblasts at the gene expression level. The comparison analysis in the present study was expected to show the similarity between iPSCs, hESCs and parental fibroblasts. Results provide important information of the differences between iPSCs, hESCs and parental fibroblasts. Total RNA was obtained from different samples (iPSCs, hESCs, and fibroblasts) separately.
2010-12-23 | E-GEOD-26093 | biostudies-arrayexpress
Project description:Human mesenchymal stem cells (hMSCs) promote endogenous tissue regeneration and have become a promising candidate for cell therapy. However, in vitro culture expansion of hMSCs induces a rapid decline of stem cell properties through replicative senescence. Here we characterize metabolic profiles of hMSCs during expansion. We show that alterations of cellular nicotinamide adenine dinucleotide (NAD+ /NADH) redox balance and activity of the Sirtuin (Sirt) family enzymes regulate cellular senescence of hMSCs. Treatment with NAD+ precursor nicotinamide increases the intracellular NAD+ level and re-balances the NAD+ /NADH ratio, with enhanced Sirt-1 activity in hMSCs at high passage, partially restores mitochondrial fitness and rejuvenates senescent hMSCs. By contrast, human fibroblasts exhibit limited senescence as their cellular NAD+ /NADH balance is comparatively stable during expansion. These results indicate a potential metabolic and redox connection to replicative senescence in adult stem cells and identify NAD+ as a metabolic regulator that distinguishes stem cells from mature cells. This study also suggests potential strategies to maintain cellular homeostasis of hMSCs in clinical applications.
2021-09-09 | PXD022395 | Pride
Project description:Cells in culture undergo replicative senescence and unequivocally stop proliferation after a limited number of cell divisions. In this study, we have expanded mesenchymal stem cells (MSC) from human adipose tissue and analyzed genetic and epigenetic sequels. The subpopulation of highly proliferative cells and the in vitro differentiation potential decayed already within early passages. Relevant chromosomal aberrations were not detected by karyotyping and SNP-microarrays. Comparison of early and late passage of five samples of mesenchymal stem cells from human adipose tissue.
2013-02-15 | E-GEOD-27135 | biostudies-arrayexpress
Project description:DallePazze2014 - Cellular senescene-induced
mitochondrial dysfunction
This model is described in the article:
Dynamic modelling of
pathways to cellular senescence reveals strategies for targeted
interventions.
Dalle Pezze P, Nelson G, Otten EG,
Korolchuk VI, Kirkwood TB, von Zglinicki T, Shanley DP.
PLoS Comput. Biol. 2014 Aug; 10(8):
e1003728
Abstract:
Cellular senescence, a state of irreversible cell cycle
arrest, is thought to help protect an organism from cancer, yet
also contributes to ageing. The changes which occur in
senescence are controlled by networks of multiple signalling
and feedback pathways at the cellular level, and the interplay
between these is difficult to predict and understand. To
unravel the intrinsic challenges of understanding such a highly
networked system, we have taken a systems biology approach to
cellular senescence. We report a detailed analysis of
senescence signalling via DNA damage, insulin-TOR, FoxO3a
transcription factors, oxidative stress response, mitochondrial
regulation and mitophagy. We show in silico and in vitro that
inhibition of reactive oxygen species can prevent loss of
mitochondrial membrane potential, whilst inhibition of mTOR
shows a partial rescue of mitochondrial mass changes during
establishment of senescence. Dual inhibition of ROS and mTOR in
vitro confirmed computational model predictions that it was
possible to further reduce senescence-induced mitochondrial
dysfunction and DNA double-strand breaks. However, these
interventions were unable to abrogate the senescence-induced
mitochondrial dysfunction completely, and we identified
decreased mitochondrial fission as the potential driving force
for increased mitochondrial mass via prevention of mitophagy.
Dynamic sensitivity analysis of the model showed the network
stabilised at a new late state of cellular senescence. This was
characterised by poor network sensitivity, high signalling
noise, low cellular energy, high inflammation and permanent
cell cycle arrest suggesting an unsatisfactory outcome for
treatments aiming to delay or reverse cellular senescence at
late time points. Combinatorial targeted interventions are
therefore possible for intervening in the cellular pathway to
senescence, but in the cases identified here, are only capable
of delaying senescence onset.
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2015-08-12 | BIOMD0000000582 | BioModels
Project description:Hutchinson Gilford Progeria Syndrome (HGPS) is a rare, sporadic genetic disease caused by mutations in the nuclear lamin A gene. In most cases the mutation creates an efficient donor-splice site that generates an altered transcript encoding a truncated lamin A protein, progerin. In vitro studies have indicated that progerin can disrupt nuclear function. HGPS affects mainly mesenchymal lineages but the shortage of patient material has precluded a tissue-wide molecular survey of progerin’s cellular impact. We report here a new, induced pluripotent stem cell (iPSC)-based model for studying HGPS. HGPS dermal fibroblasts were reprogrammed into iPSC lines using a cocktail of the transcription factor genes, OCT4, SOX2, KLF4 and C-MYC. The iPSC cells were differentiated into neural progenitors (NPs), endothelial cells (ECs), fibroblast-like cells and mesenchymal stem cells (MSCs). Progerin levels in the different cell types followed the pattern MSC≥ fibroblast>EC>>NP. Functionally, we detected a major impact of progerin on MSC function. We show that HGPS-MSCs are vulnerable to the ischemic conditions found in a murine hind limb recovery model and an in vitro hypoxia assay, as well as showing enhanced sensitivity in a serum starvation assay. Since there is widespread consensus that MSCs reside in low oxygen niches in vivo, we propose that these conditions lead to an accelerated depletion of the MSC pool in HGPS patients with consequent accretion of mesenchymal tissue.
2010-12-23 | GSE26093 | GEO
Project description:<p><strong>BACKGROUND:</strong> In all living organisms, DNA replication is exquisitely regulated in a wide range of growth conditions to achieve timely and accurate genome duplication prior to cell division. Failures in this regulation cause DNA damage with potentially disastrous consequences for cell viability and human health, including cancer. To cope with these threats, cells tightly control replication initiation using well-known mechanisms. They also couple DNA synthesis to nutrient richness and growth rate through a poorly understood process thought to involve central carbon metabolism. One such process may involve the cross-species conserved pyruvate kinase (PykA) which catalyzes the last reaction of glycolysis. Here we have investigated the role of PykA in regulating DNA replication in the model system <em>Bacillus subtilis.</em></p><p><strong>RESULTS: </strong>On analysing mutants of the catalytic (Cat) and C-terminal (PEPut) domains of <em>B. subtilis</em> PykA we found replication phenotypes in conditions where PykA is dispensable for growth. These phenotypes are independent from the effect of mutations on PykA catalytic activity and are not associated with significant changes in the metabolome. PEPut operates as a nutrient-dependent inhibitor of initiation while Cat acts as a stimulator of replication fork speed. Disruption of either PEPut or Cat replication function dramatically impacted the cell cycle and replication timing even in cells fully proficient in known replication control functions. <em>In vitro</em>, PykA modulates activities of enzymes essential for replication initiation and elongation via functional interactions. Additional experiments showed that PEPut regulates PykA activity and that Cat and PEPut determinants important for PykA catalytic activity regulation are also important for PykA-driven replication functions.</p><p><strong>CONCLUSIONS:</strong> We infer from our findings that PykA typifies a new family of cross-species replication control regulators that drive the metabolic control of replication through a mechanism involving regulatory determinants of PykA catalytic activity. As disruption of PykA replication functions causes dramatic replication defects, we suggest that dysfunctions in this new family of universal replication regulators may pave the path to genetic instability and carcinogenesis.</p>
2022-04-19 | MTBLS4415 | MetaboLights
Project description:Primary cells enter replicative senescence after a limited number of cell divisions. This process is associated with reproducible changes in DNA methylation (DNAm) at specific sites in the genome. The mechanism that drives senescence-associated DNAm changes remains unknown and may arise through drift in DNAm or through regulated, senescence dependent modifications at specific sites in the genome. In this study, we analyzed the reorganization of nuclear architecture and DNA methylation during long-term culture of human fibroblasts and mesenchymal stromal cells (MSCs). [MethylCap-seq] Fibroblasts of two female donors (both 43 years old) were culture expanded and DNA was harvested of 10,000,000 cells at early passage (P3 or P5) and late passage (P30 and P33). DNA methylation changes were subsequently analyzed by MethylCap-Seq.
2015-03-26 | E-GEOD-59960 | biostudies-arrayexpress