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Replicative Senescence (replicative + senescence)
Selected AbstractsSenescence-associated ,-galactosidase is lysosomal ,-galactosidaseAGING CELL, Issue 2 2006Bo Yun Lee Summary Replicative senescence limits the proliferation of somatic cells passaged in culture and may reflect cellular aging in vivo. The most widely used biomarker for senescent and aging cells is senescence-associated ,-galactosidase (SA-,-gal), which is defined as ,-galactosidase activity detectable at pH 6.0 in senescent cells, but the origin of SA-,-gal and its cellular roles in senescence are not known. We demonstrate here that SA-,-gal activity is expressed from GLB1, the gene encoding lysosomal ,-D-galactosidase, the activity of which is typically measured at acidic pH 4.5. Fibroblasts from patients with autosomal recessive GM1 -gangliosidosis, which have defective lysosomal ,-galactosidase, did not express SA-,-gal at late passages even though they underwent replicative senescence. In addition, late passage normal fibroblasts expressing small-hairpin interfering RNA that depleted GLB1 mRNA underwent senescence but failed to express SA-,-gal. GLB1 mRNA depletion also prevented expression of SA-,-gal activity in HeLa cervical carcinoma cells induced to enter a senescent state by repression of their endogenous human papillomavirus E7 oncogene. SA-,-gal induction during senescence was due at least in part to increased expression of the lysosomal ,-galactosidase protein. These results also indicate that SA-,-gal is not required for senescence. [source] Accumulation of multiple forms of lamin A with down-regulation of FACE-1 suppresses growth in senescent human cellsGENES TO CELLS, Issue 3 2007Ryo Ukekawa 5-Bromodeoxyuridine (BrdU) clearly induces a senescence-like phenomenon in every cell type. Proteome analysis revealed that lamin A and C were most highly increased in the nuclei of HeLa cells upon addition of BrdU. Immunoblot analysis also revealed marked accumulation of nuclear prelamin A. Consistently, farnesylated-proteins converting enzyme 1 (FACE-1) was markedly down-regulated in the same cells. Similar phenomena were also observed in normal human fibroblasts undergoing replicative senescence. Immunochemical analysis confirmed the above results. Lamin A is a major component of lamina and responsible for several genetic diseases. Thus, we ectopically expressed a wild-type, a mature type and a premature type of lamin in HeLa cells. All of these forms similarly inhibited colony formation and delayed cell cycle progression mainly through G2 phase. These results suggest that a change in the amount of lamin A, rather than appearance of its truncated form, is responsible for growth retardation in affected cells. [source] Cellular senescence and longevity of osteophyte-derived mesenchymal stem cells compared to patient-matched bone marrow stromal cellsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 4 2009Sanjleena Singh Abstract This study aimed to determine the cellular aging of osteophyte-derived mesenchymal cells (oMSCs) in comparison to patient-matched bone marrow stromal cells (bMSCs). Extensive expansion of the cell cultures was performed and early and late passage cells (passages 4 and 9, respectively) were used to study signs of cellular aging, telomere length, telomerase activity, and cell-cycle-related gene expression. Our results showed that cellular aging was more prominent in bMSCs than in oMSCs, and that oMSCs had longer telomere length in late passages compared with bMSCs, although there was no significant difference in telomere lengths in the early passages in either cell type. Telomerase activity was detectable only in early passage oMSCs and not in bMSCs. In osteophyte tissues telomerase-positive cells were found to be located perivascularly and were Stro-1 positive. Fifteen cell-cycle regulator genes were investigated and only three genes (APC, CCND2, and BMP2) were differentially expressed between bMSC and oMSC. Our results indicate that oMSCs retain a level of telomerase activity in vitro, which may account for the relatively greater longevity of these cells, compared with bMSCs, by preventing replicative senescence. J. Cell. Biochem. 108: 839,850, 2009. © 2009 Wiley-Liss, Inc. [source] Defining the transcriptome of accelerated and replicatively senescent keratinocytes reveals links to differentiation, interferon signaling, and Notch related pathways,JOURNAL OF CELLULAR BIOCHEMISTRY, Issue 2 2006Ranjan J. Perera Abstract Epidermal keratinocytes (KCs) undergo highly orchestrated morphological and molecular changes during transition from proliferative compartment into growth arrested early and late differentiation layers, prior to dying in outermost cornified layers of normal skin. Creation of stratum corneum is vital to barrier function protecting against infection. Transcriptional events in KCs regulating complex processes of differentiation and host defense required to maintain constant epidermal thickness and resistance to infection in either young or aged skin are largely unknown. Furthermore, as terminal differentiation is characterized by irreversible loss of replicative potential culminating in dead layers at the skin surface, this process may be viewed as a form of senescence. However, a complete transcriptional profile of senescent (SN) human KCs has not been previously defined to permit delineation of molecular boundaries involving differentiation and senescence. To fill this void, we utilized global transcriptional analysis of KCs maintained in vitro as either cultures of proliferating (PR) cells, early and late confluent (LC) (accelerated senescence) cultures, or KCs undergoing replicative senescence. Global gene expression profiling revealed early confluent (EC) KCs were somewhat similar to PR KCs, while prominent differences were evident when compared to LC KCs; which were also distinct from replicatively SN KCs. While confluent KCs have in common several genes regulating differentiation with replicatively SN KCs, the latter cells expressed elevated levels of genes involved in interferon signaling and inflammatory pathways. These results provide new insights into cell autonomous transcriptional-based programs operative within KCs contributing to replicative senescence, with partial sharing of genes involved in differentiation. In addition, regulation of KC senescence may involve participation of interferon signaling pathways derived from the important role of KCs in protecting skin from infection. Integrating all of the transcriptional data revealed a key role for Notch receptor mediated signaling in the confluency induced differentiation phenotype using this model system. J. Cell. Biochem. 98: 394,408, 2006. © 2006 Wiley-Liss, Inc. [source] Gene positional changes relative to the nuclear substructure during carbon tetrachloride-induced hepatic fibrosis in ratsJOURNAL OF CELLULAR BIOCHEMISTRY, Issue 6 2004Apolinar Maya-Mendoza Abstract In the interphase nucleus the DNA of higher eukaryotes is organized in loops anchored to a substructure known as the nuclear matrix (NM). The topological relationship between gene sequences located in the DNA loops and the NM appears to be very important for nuclear physiology because processes such as replication, transcription, and processing of primary transcripts occur at macromolecular complexes located at discrete sites upon the NM. Mammalian hepatocytes rarely divide but preserve a proliferating capacity that is displayed in vivo after specific stimulus. We have previously shown that transient changes in the relative position of specific genes to the NM occur during the process of liver regeneration after partial ablation of the liver, but also that such changes correlate with the replicating status of the cells. Moreover, since chronic exposure to carbon tetrachloride (CCl4) leads to bouts of hepatocyte damage and regeneration, and eventually to non-reversible liver fibrosis in the rat, we used this animal model in order to explore if genes that show differential activity in the liver change or modify their relative position to the NM during the process of liver fibrosis induction. We found that changes in the relative position of specific genes to the NM occur during the chronic administration of CCl4, but also that such changes correlate with the proliferating status of the hepatocytes that goes from quiescence to regeneration to replicative senescence along the course of CCl4 -induced liver fibrosis, indicating that specific configurations in the higher-order DNA structure underlie the stages of progression towards liver fibrosis. © 2004 Wiley-Liss, Inc. [source] BTG2 antagonizes Pin1 in response to mitogens and telomere disruption during replicative senescenceAGING CELL, Issue 5 2010Keith Wheaton Summary Cellular senescence limits the replicative capacity of normal cells and acts as an intrinsic barrier that protects against the development of cancer. Telomere shortening,induced replicative senescence is dependent on the ATM-p53-p21 pathway but additional genes likely contribute to senescence. Here, we show that the p53-responsive gene BTG2 plays an essential role in replicative senescence. Similar to p53 and p21 depletion, BTG2 depletion in human fibroblasts leads to an extension of cellular lifespan, and ectopic BTG2 induces senescence independently of p53. The anti-proliferative function of BTG2 during senescence involves its stabilization in response to telomere dysfunction followed by serum-dependent binding and relocalization of the cell cycle regulator prolyl isomerase Pin1. Pin1 inhibition leads to senescence in late-passage cells, and ectopic Pin1 expression rescues cells from BTG2-induced senescence. The neutralization of Pin1 by BTG2 provides a critical mechanism to maintain senescent arrest in the presence of mitogenic signals in normal primary fibroblasts. [source] Protein modification and replicative senescence of WI-38 human embryonic fibroblastsAGING CELL, Issue 2 2010Emad K. Ahmed Summary Oxidized proteins as well as proteins modified by the lipid peroxidation product 4-hydroxy-2-nonenal (HNE) and by glycation (AGE) have been shown to accumulate with aging in vivo and during replicative senescence in vitro. To better understand the mechanisms by which these damaged proteins build up and potentially affect cellular function during replicative senescence of WI-38 fibroblasts, proteins targeted by these modifications have been identified using a bidimensional gel electrophoresis-based proteomic approach coupled with immunodetection of HNE-, AGE-modified and carbonylated proteins. Thirty-seven proteins targeted for either one of these modifications were identified by mass spectrometry and are involved in different cellular functions such as protein quality control, energy metabolism and cytoskeleton. Almost half of the identified proteins were found to be mitochondrial, which reflects a preferential accumulation of damaged proteins within the mitochondria during cellular senescence. Accumulation of AGE-modified proteins could be explained by the senescence-associated decreased activity of glyoxalase-I, the major enzyme involved in the detoxification of the glycating agents methylglyoxal and glyoxal, in both cytosol and mitochondria. This finding suggests a role of detoxification systems in the age-related build-up of damaged proteins. Moreover, the oxidized protein repair system methionine sulfoxide reductase was more affected in the mitochondria than in the cytosol during cellular senescence. Finally, in contrast to the proteasome, the activity of which is decreased in senescent fibroblasts, the mitochondrial matrix ATP-stimulated Lon-like proteolytic activity is increased in senescent cells but does not seem to be sufficient to cope with the increased load of modified mitochondrial proteins. [source] DNA methylation pattern changes upon long-term culture and aging of human mesenchymal stromal cellsAGING CELL, Issue 1 2010Simone Bork Summary Within 2,3 months of in vitro culture-expansion, mesenchymal stromal cells (MSC) undergo replicative senescence characterized by cell enlargement, loss of differentiation potential and ultimate growth arrest. In this study, we have analyzed DNA methylation changes upon long-term culture of MSC by using the HumanMethylation27 BeadChip microarray assessing 27 578 unique CpG sites. Furthermore, we have compared MSC from young and elderly donors. Overall, methylation patterns were maintained throughout both long-term culture and aging but highly significant differences were observed at specific CpG sites. Many of these differences were observed in homeobox genes and genes involved in cell differentiation. Methylation changes were verified by pyrosequencing after bisulfite conversion and compared to gene expression data. Notably, methylation changes in MSC were overlapping in long-term culture and aging in vivo. This supports the notion that replicative senescence and aging represent developmental processes that are regulated by specific epigenetic modifications. [source] Senescence-associated ,-galactosidase is lysosomal ,-galactosidaseAGING CELL, Issue 2 2006Bo Yun Lee Summary Replicative senescence limits the proliferation of somatic cells passaged in culture and may reflect cellular aging in vivo. The most widely used biomarker for senescent and aging cells is senescence-associated ,-galactosidase (SA-,-gal), which is defined as ,-galactosidase activity detectable at pH 6.0 in senescent cells, but the origin of SA-,-gal and its cellular roles in senescence are not known. We demonstrate here that SA-,-gal activity is expressed from GLB1, the gene encoding lysosomal ,-D-galactosidase, the activity of which is typically measured at acidic pH 4.5. Fibroblasts from patients with autosomal recessive GM1 -gangliosidosis, which have defective lysosomal ,-galactosidase, did not express SA-,-gal at late passages even though they underwent replicative senescence. In addition, late passage normal fibroblasts expressing small-hairpin interfering RNA that depleted GLB1 mRNA underwent senescence but failed to express SA-,-gal. GLB1 mRNA depletion also prevented expression of SA-,-gal activity in HeLa cervical carcinoma cells induced to enter a senescent state by repression of their endogenous human papillomavirus E7 oncogene. SA-,-gal induction during senescence was due at least in part to increased expression of the lysosomal ,-galactosidase protein. These results also indicate that SA-,-gal is not required for senescence. [source] Nuclear accumulation of glycogen synthase kinase-3 during replicative senescence of human fibroblastsAGING CELL, Issue 5 2004Jaroslaw W. Zmijewski Summary Activation of the tumor suppressor protein p53 contributes to cellular senescence. As glycogen synthase kinase-3 (GSK3) was recently found to interact with p53 and contribute to the actions of p53, this study examined whether GSK3 accumulated in the nucleus and associated with p53 in senescent cells. Compared with young and middle-aged human WI-38 fibroblasts, senescent cells were found to contain increased nuclear levels of GSK3,, and also tended to accumulate in the nucleus the other isoform of GSK3, GSK3,. Co-immunoprecipitation experiments demonstrated that GSK3, and p53 formed a complex in the nucleus. Further experiments tested whether inhibition of GSK3 altered the development of senescence using long-term treatment with the selective GSK3 inhibitor lithium. Lithium treatment reduced the senescence-associated accumulation of p53 and caused cells to enter a reversible quiescent state. These results indicate that a portion of the p53 that is activated in senescent cells is modulated by its association with GSK3, in the nucleus, an association that is known to facilitate the actions of p53 and that may contribute to senescence. [source] Bcl-2 overexpression in hepatic stellate cell line CFSC-2G, induces a pro-fibrotic stateJOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY, Issue 7 2010Viridiana Y González-Puertos Abstract Background and Aim:, Development of hepatic fibrosis is a complex process that involves oxidative stress (OS) and an altered balance between pro- and anti-apoptotic molecules. Since Bcl-2 overexpression preserves viability against OS, our objective was to address the effect of Bcl-2 overexpression in the hepatic stellate cells (HSC) cell-line CFSC-2G under acetaldehyde and H2O2 challenge, and explore if it protects these cells against OS, induces replicative senescence and/or modify extracellular matrix (ECM) remodeling potential. Methods:, To induce Bcl-2 overexpression, HSC cell line CFSC-2G was transfected by lipofection technique. Green fluorescent protein-only CFSC-2G cells were used as a control. Cell survival after H2O2 treatment and total protein oxidation were assessed. To determine cell cycle arrest, proliferation-rate, DNA synthesis and senescence were assessed. Matrix metalloproteinases (MMP), tissue-inhibitor of MMP (TIMP), transglutaminases (TG) and smooth muscle a-actin (,-SMA) were evaluated by western blot in response to acetaldehyde treatment as markers of ECM remodeling capacity in addition to transforming growth factor-, (TGF-,) mRNA. Results:, Cells overexpressing Bcl-2 survived , 20% more than control cells when exposed to H2O2 and , 35% proteins were protected from oxidation, but Bcl-2 did not slow proliferation or induced senescence. Bcl-2 overexpression did not change ,-SMA levels, but it increased TIMP-1 (55%), tissue transglutaminases (tTG) (25%) and TGF-, mRNA (49%), when exposed to acetaldehyde, while MMP-13 content decreased (47%). Conclusions:, Bcl-2 overexpression protected HSC against oxidative stress but it did not induce replicative senescence. It increased TIMP-1, tTG and TGF-, mRNA levels and decreased MMP-13 content, suggesting that Bcl-2 overexpression may play a key role in the progression of fibrosis in chronic liver diseases. [source] Proteomic profiling of tumor cells after induction of telomere dysfunctionPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 3 2009Stefan Zimmermann Dr. Abstract Cell division in the absence of telomerase causes progressive telomere shortening which ultimately leads to telomere dysfunction and initiation of genome instability. In order to identify factors related to loss of telomere function, the effects of telomerase inhibition on the proteome of five tumor cell lines were followed by SELDI-TOF-MS. Five differentially expressed protein peaks (p<0.01) were found in a total of 60 clones of five cell lines representing four tissues (lung, breast, prostate, and colon) in which telomerase was inhibited by retroviral overexpression of a dominant negative (DN) mutant of human telomerase reverse transcriptase (hTERT). Among these, a 11.3,kDa peak diminished in DN-hTERT clones was identified as histone H4 by nanoflow-HPLC-MS/MS. Immunoblot analysis not only confirmed the decline of histone H4, but also of other core histone proteins including histone H3. Furthermore, upregulation of several cytokeratins was found to be associated with telomere attrition. In conclusion, loss of telomere function is associated with alterations in the proteome which may represent novel biomarkers for the detection of replicative senescence. [source] | ||||||||||