ABSTRACT: Transcription profiling of skeletal muscles from young, old, and old calorie restricted C57BL/6NHsd mice to investigate age-related changes in the transcriptional profile
Project description:We investigated age-related changes in the transcriptional profile of skeletal muscle in 5 month old (young) and 25 month old (old) C57BL/6NHsd mice using high density oligonucleotide arrays (22,690 transcripts probed). We identified 712 transcripts that are differentially expressed in young (5 month old) and old (25-month old) mouse skeletal muscle. Caloric restriction (CR) completely or partially reversed 87% of the changes in expression. Examination of individual genes revealed a transcriptional profile indicative of increased p53 activity in the older muscle. To determine whether the increase in p53 activity is associated with transcriptional activation of apoptotic targets, we performed RT-PCR on four well known mediators of p53-induced apoptosis: puma, noxa, tnfrsf10b and bok. Expression levels for these proapoptotic genes increased significantly with age (P<0.05), while CR significantly lowered expression levels for these genes as compared to control fed old mice (P<0.05). Age-related induction of p53-related genes was observed in multiple tissues, but was not observed in SOD2+/- and GPX4+/- mice, suggesting that oxidative stress does not mediate the observed age-related increase in expression. Western blot analysis confirmed that protein levels for both p21 and GADD45a, two established transcriptional targets of p53, were higher in the older muscle tissue. These observations support a role for p53-mediated apoptotic activity in mammalian aging. Keywords: aging, calorie restriction, muscle, p53
Project description:During aging, the number and functionality of muscle stem cells (MuSCs) decreases leading to impaired regeneration of aged skeletal muscle. In addition to intrinsic changes in aged MuSCs, extracellular matrix (ECM) proteins deriving from other cell types, e.g., fibrogenic-adipogenic progenitor cells (FAPs), contribute to the aging phenotype of MuSCs and impaired regeneration in the elderly. So far, no comprehensive analysis on how age-dependent changes in the whole skeletal muscle proteome affect MuSC function have been conducted. Here, we investigated age-dependent changes in the proteome of different skeletal muscle types by applying deep quantitative mass spectrometry. We identified 183 extracellular matrix proteins that show different abundances in skeletal muscles of old mice. By integrating single cell sequencing data, we reveal that transcripts of those ECM proteins are mainly expressed in FAPs, suggesting that FAPs are the main contributors to ECM remodelling during aging. We functionally investigated one of those ECM molecules, namely Smoc2, which is aberrantly expressed during aging. We show that Smoc2 levels are elevated during regeneration and that its accumulation in the aged MuSC niche causes impairment of MuSCs function through constant activation of integrin/MAPK signaling. In vivo, supplementation of exogenous Smoc2 hampers the regeneration of young muscles following serial injuries, leading to a phenotype reminiscent of regenerating aged skeletal muscle. Taken together, we provide a comprehensive resource of changes in the composition of the ECM of aged skeletal muscles, we pinpoint the cell types driving these changes, and we identify a new niche protein causing functional impairment of MuSCs thereby hampering the regeneration capacity of skeletal muscles.
Project description:During aging, the number and functionality of muscle stem cells (MuSCs) decreases leading to impaired regeneration of aged skeletal muscle. In addition to intrinsic changes in aged MuSCs, extracellular matrix (ECM) proteins deriving from other cell types, e.g., fibrogenic-adipogenic progenitor cells (FAPs), contribute to the aging phenotype of MuSCs and impaired regeneration in the elderly. So far, no comprehensive analysis on how age-dependent changes in the whole skeletal muscle proteome affect MuSC function have been conducted. Here, we investigated age-dependent changes in the proteome of different skeletal muscle types by applying deep quantitative mass spectrometry. We identified 183 extracellular matrix proteins that show different abundances in skeletal muscles of old mice. By integrating single cell sequencing data, we reveal that transcripts of those ECM proteins are mainly expressed in FAPs, suggesting that FAPs are the main contributors to ECM remodelling during aging. We functionally investigated one of those ECM molecules, namely Smoc2, which is aberrantly expressed during aging. We show that Smoc2 levels are elevated during regeneration and that its accumulation in the aged MuSC niche causes impairment of MuSCs function through constant activation of integrin/MAPK signaling. In vivo, supplementation of exogenous Smoc2 hampers the regeneration of young muscles following serial injuries, leading to a phenotype reminiscent of regenerating aged skeletal muscle. Taken together, we provide a comprehensive resource of changes in the composition of the ECM of aged skeletal muscles, we pinpoint the cell types driving these changes, and we identify a new niche protein causing functional impairment of MuSCs thereby hampering the regeneration capacity of skeletal muscles.
Project description:Utilizing glycerol intramuscular injections in M. musculus provide a models of skeletal muscle damage followed by skeletal muscle regeneration. In particular, glycerol-induced muscle injury triggers accute activation of skeletal muscle stem cells, called satellite cells. However, aging dramatically impairs the regenerative capacity of satellite cells. We characterized genome-wide expression profiles of young and old satellite cells in the non-proliferative and activated state, freshly isolated to non-injured or damaged muscles, respectively. Our goal was to uncover new regulatory signaling specific to satellite cells entry into the activation and myogenic program that are affected with age. Satellite cells were isolated in either quiescent / non-proliferative or activated state from uninjured or 3 days after glycerol-induced injury of tibialis anterior, gastrocnemius and quadriceps, respectively. Young (2-4 months old) and old (20-24 months old) wildtype C57BL/6J male were used, with five to six biological replicates per group.
Project description:Aging animals undergo a variety of changes in molecular processes. Among these, the cellular circadian clock has been shown to change as animals age. Moreover, there is evidence that also core circadian clock proteins could influence the ageing behavior of vertebrates. To investigate the interplay between aging and the circadian clock, we studied circadian mRNA expression in skeletal muscles from young (8 weeks) and aged (80 weeks) mice. In order to detect differences in circadian patterns, we used microarray-based transcriptome-wide time series of mRNA expression, containing 16 independent measurements for both young and aged animals. Each individual time point consists of total RNA from hind limb skeletal muscles from 3 different animals. Young and aged mice where entrained to 12 hr/12 hr light-dark conditions. From these mice, hind limb skeletal muscles were extracted at different times of day, in order to measure circadian mRNA expression patterns.
Project description:Background: Age-related physiological, biochemical and functional changes in mammalian skeletal muscle have been shown to begin at the mid-point of the lifespan. However, the underlying changes in DNA methylation that occur during this turning point of the muscle aging process have not been clarified. To explore age-related genomic methylation changes in skeletal muscle, we employed young (0.5 years old) and middle-aged (7 years old) pigs as models to survey genome-wide DNA methylation in the longissimus dorsi muscle using a methylated DNA immunoprecipitation sequencing approach. Results: We observed a tendency toward a global loss of DNA methylation in the gene-body region of the skeletal muscle of the middle-aged pigs compared with the young group. We determined the genome-wide gene expression pattern in the longissimus dorsi muscle using microarray analysis and performed a correlation analysis using DMR (differentially methylated region)-mRNA pairs, and we found a significant negative correlation between the changes in methylation levels within gene bodies and gene expression. Furthermore, we identified numerous genes that show age-related methylation changes that are potentially involved in the aging process. The methylation status of these genes was confirmed using bisulfite sequencing PCR. The genes that exhibited a hypomethylated gene body in middle-aged pigs were over-represented in various proteolysis and protein catabolic processes, suggesting an important role for these genes in age-related muscle atrophy. In addition, genes associated with tumorigenesis exhibited aged-related differences in methylation and expression levels, suggesting an increased risk of disease associated with increased age. Conclusions: This study provides a comprehensive analysis of genome-wide DNA methylation patterns in aging pig skeletal muscle. Our findings will serve as a valuable resource in aging studies, promoting the pig as a model organism for human aging research and accelerating the development of comparative animal models in aging research. We collected the longissimus dorsi muscles tissue from Jinhua pigs which aged 0.5 year and seven years and study the genome-wide DNA methylation difference between the two age periods.
Project description:For additional details see Ebert et al, Identification and Small Molecule Inhibition of an ATF4-dependent Pathway to Age-related Skeletal Muscle Weakness and Atrophy. Weight-matched cohorts of 22-month-old male C57BL/6 mice were provided ad libitum access to standard chow (control) or standard chow supplemented with 0.27% ursolic acid (UA) or 0.05% tomatidine (TM) for 2 months. After the 2 month treatment period, quadriceps femoris muscles were harvested. mRNA levels in muscles harvested from ursolic acid or tomatidine fed mice were normalized to levels in muscles fed control diet.
Project description:We carried out a global survey of age-related changes in mRNA levels in the C57BL/6NIA mouse hippocampus and found a difference in the hippocampal gene expression profile between 2-month-old young mice and 15-month-old middle-aged mice correlated with an age-related cognitive deficit in hippocampal-based explicit memory formation. Middle-aged mice displayed a mild but specific deficit in spatial memory in the Morris water maze. Keywords: age comparison
Project description:Aging animals undergo a variety of changes in molecular processes. Among these, the cellular circadian clock has been shown to change as animals age. Moreover, there is evidence that also core circadian clock proteins could influence the ageing behavior of vertebrates. To investigate the interplay between aging and the circadian clock, we studied circadian mRNA expression in skeletal muscles from young (8 weeks) and aged (80 weeks) mice. In order to detect differences in circadian patterns, we used microarray-based transcriptome-wide time series of mRNA expression, containing 16 independent measurements for both young and aged animals. Each individual time point consists of total RNA from hind limb skeletal muscles from 3 different animals.
Project description:Comprehensive analyses of mRNA expression were performed using three different cell populations isolated from skeletal muscle of young and aged mice to investigate age-related changes of each cell population.