Whole blood transcriptomics to define adaptive biochemical pathways of exercise during aging
ABSTRACT: The aim of the current study was to characterize the genetic adaptive pathways altered by exercise in veteran athletes and age-matched untrained individuals. Two groups of 50-60 year old males: competitive cyclists and untrained, minimally active individuals were examined. All participants completed an acut bout of submaximal endurance exercise and blood samples pre- and post-exercise were analyzed for gene expression changes utilizing genome-wide DNA microarray analysis. Our results indicate distinct differences in gene expression involving energy metabolism, lipids, insuling signaling and cardiovascular function between the two groups. These findings may lead to new insights into beneficial signaling pathways of healthy aging and help identify surrogate markers for monitoring exercise and training load. Blood samples from the control and athlete groups were analyzed at three time-points: T1 (before exercise); T2 (immediately after exercise) and T3 (24 hours after exercise). There were n = 4 samples in each of control and athlete group at T1 and T3; and n = 7 for control group and n = 8 for athlete group at T2. One athlete sample (Sample # 010201) at time - point T2 had a technical replicate.
Project description:Purpose: Next-generation sequencing (NGS) was used to select genes potentially associated with exercise adaptation in Arabian horses. Methods: Whole transcriptome profiling of blood was performed for untrained horses and horses from which samples were collected during at 3 different periods of training procedure (T1-during intense training period - March, T2- before starts - May and T3 -after flat racing season - October). The muscle transcriptome sequencing was performed for 37 blood samples using Illumina HiScan SQ in 75 single-end cycles. The quantifying transcript abundances was made using the RSEM supported by STAR aligner. The raw reads were aligned to the Equus caballus reference genome. Differentially expressed genes in blood tissue were detected by DESeq2. The RNA-seq results were validated using by qPCR. Results: The increase of the number of DEGs between subsequent training periods has been observed and the highest amount of DEGs was detected between untrained horses (T0) and horses at the end of the racing season (T3) – 440. The comparison of transcriptome of T2 vs T3 and T0 vs T3 showed a significant advantage of up-regulated genes during long-term exercise (up-regulation of 266 and 389 DEGs in T3 period compared T2 and T0; respectively). Our results showed that the largest number of identified genes encoded transcription factors, nucleic acid binding proteins and G-protein modulators, which mainly were transcriptional activated at the last training phase (T3) . Moreover, in the T3 period the identified DEGs represented genes coded for cytoskeletal proteins including actin cytoskeletal proteins and kinases. The most abundant exercise-upregulated genes were involved in pathways important in regulating the cell cycle (PI3K-Akt signaling pathway), cell communication (cAMP-dependent pathway), proliferation, differentiation and apoptosis as well as immunity processes (Jak-STAT signaling pathway). We also observed exercise induced expression of genes related in regulation of actin cytoskeleton, gluconeogenesis (FoxO signaling pathway; Insulin signaling pathway), glycerophospholipid metabolism and calcium signaling. Conclusions: TOur results allow to identify changes in genes expression profile following training schedule in Arabian horses. Based on comparison analysis of blood transcriptomes, several exercise-regulated pathways and genes most affected by exercise were detected. We pinpointed overrepresented molecular pathways and genes essential for exercise adaptive response via maintaining of body homeostasis. The observed transcriptional activation of such gene as LPGAT1, AGPAT5, PIK3CG, GPD2, FOXN2, FOXO3, ACVR1B and ACVR2A can be a base for further research in order to identify genes potentially associated with race performance in Arabian horses. Such markers will be essential to choice the training type, and could result in differences in racing performance specific to various breeds. The blood transcriptome sequencing was performed for 37 samples collected form Arabian horses using Illumina HiScan SQ in75 single-end cycles and in 3-4 technical repetitions.repetitions.
Project description:Impact of antibiotics (T2) or antibiotics in combination with stress (T3) in early life on intestinal functioning in pigs on 8, 55, 176 days in jejunum and ileum (blood only day 8) and control pigs (T1) 4 pools consisting of 16 animals were generated per time-point (day 8, 55, 176 after birth) per treatment (T1;control, T2; antibiotics, T3; antibiotics+stress)
Project description:Unconditioned thoroughbred geldings were exercised to maximal heart rate or fatigue on an equine high-speed treadmill. Skeletal muscle biopsies were taken from the middle gluteal muscle before, immediately after and four hours after exercise. Three-condition experiment, Pre exercise (T0), Immediately post exercise (T1), 4 hours post exercise (T2). Hybridisations: T0 vs T1, T0 vs T2 Biological replicates: 8 Technical replication Dye swap
Project description:MiRNAs are essential mediators of many biological processes. The aim of this study was to investigate the dynamics of miRNA-mRNA regulatory networks during exercise and subsequent recovery period. Here we monitored the transcriptome changes by microarray analysis of the whole blood of eight highly trained athletes after 30 min of moderate exercise followed by 30 min and 60 min of recovery period. We combined expression profiling and bioinformatics and analysed metabolic pathways enriched with differentially expressed mRNAs and then mRNAs which are known to be validated targets of differentially expressed miRNAs. Finally we revealed four dynamically regulated networks comprising differentially expressed miRNAs and their known target mRNAs with anti-correlated expression profiles over time. The data suggest that hsa-miR-21-5p regulated TGFBR3, PDGFD and PPM1L mRNAs. Hsa-miR-24-2-5p was likely to be responsible for MYC and KCNJ2 genes and hsa-miR-27a-5p for ST3GAL6. The targets of hsa-miR-181a-5p included ROPN1L and SLC37A3. All these mRNAs are involved in processes highly relevant to exercise response, including immune function, apoptosis, membrane traffic of proteins and transcription regulation. We have identified four miRNA-mRNA networks dynamically regulated following exercise. This work is the first study to monitor miRNAs and mRNAs in parallel into recovery period. The results provide a novel insight into the regulatory role of miRNAs in stress adaptation. Each subject performed a treadmill test with an incremental step protocol until exhaustion. Blood samples were taken from the antecubital vein using indwelling catheter before (T1), immediately after ramp test to exaustion (T2) and after 30 min of recovery (T3). Two weeks later, athletes participated in the main exercise, consisting of running at 80% VO2 peak for 30 min on a treadmill. Blood samples were taken from the antecubital vein using indwelling catheter before (T4), immediately after running at 80% VO2 peak for 30 min on a treadmill (T5), after 30 min (T6) and 60 min (T7) of recovery.
Project description:Unconditioned thoroughbred geldings were exercised to maximal heart rate or fatigue on an equine high-speed treadmill. Skeletal muscle biopsies were taken from the middle gluteal muscle before, immediately after and four hours after exercise. Overall design: Three-condition experiment, Pre exercise (T0), Immediately post exercise (T1), 4 hours post exercise (T2). Hybridisations: T0 vs T1, T0 vs T2 Biological replicates: 8 Technical replication Dye swap
Project description:Purpose: RNA-seq method was used to select genes expressed in muscle tissue and are potentially associated with exercise adaptation in Arabian horses. Methods: Whole transcriptomes between three time points of muscle tissue collection were compared: T0 (untrained horses), T1 (horses after intense gallop phase) and T2 (at the end of the racing season), in total 23 samples. The biopsy of gluteus medius muscle was performed by using minimally invasive ProMag™ Ultra Automatic Biopsy Instrument with a 2 mm diameter biopsy needle. The total RNA was isolated using by TriReagent and 300ng was used to cDNA libraries preparation. The NGS sequencing was performed on HiScan SQ (Illumina). The quantifying transcript abundances was made using the RSEM supported by STAR aligner. The raw reads were aligned to the Equus caballus reference genome. Differentially expressed genes were detected by DESeq2. The RNA-seq results were validated using by qPCR. Results: To detected differentially expressed genes during training preparing to the flat racing, whole transcriptomes between three time points of muscle tissue collection were compared: T0 (untrained horses), T1 (horses after intense gallop phase) and T2 (at the end of the racing season). We identified 1168 DEGs between T0 vs T1; 1593 between T1 vs T2 and 763 between T2 vs T0. The analysis for all DEGs allow to detect 11 pathways which ale significant over represented between at last two training periods. The numerous group of exercise-regulated DEGs was related with muscle cell structure and signaling (‘focal adhesion’, ‘adherens juntion’ and ‘PI3-ATK signaling’) and included insulin-like growth factor 1 receptor (IGF1R); insulin receptor (INSR); transforming growth factor beta receptors 1 and 2 (TGFBR1; TGFBR2); vascular endothelial growth factor B (VEGFB); epidermal growth factor (EGF); hepatocyte growth factor (HGF) and vascular endothelial growth factor D (FIGF). Our results showed that in Arabian horses exercise modified the expression of genes belonging to the ‘PPAR signaling pathway’ (e.g. PPARA; PPARD; PLIN2); ‘calcium signaling pathway’ (e.g. PLN; PLCD1; TNNC1; TNNC2) as well as pathways associated with metabolism processes - ‘oxidative phosphorylation’; ‘fatty acid metabolism’; ‘glycolysis/gluconeogenesis’ and ‘citrate cycle’. Conclusions: Our research allowed to identify the group of exercise-regulated genes which was related with muscle cell structure as well as signaling and pinpointed the significant metabolic processes critical for adaptive response during training. We confirmed that in Arabians, the exercise switch energy generation towards fatty acid utilization, enhance glycogen transport and calcium signaling. The sequencing of skeletal muscle transcriptome allowed to propose the panel of new candidate genes (such as SLC16A1; ME3; ACTN3; PPARα; SH3RF2; TPM3; TNNC1; TNNI3; TGFBR1; TGFBR2; FABP3) potentially related with body homeostasis maintenance and race performance in Arabian horse. Overall design: The muscle (gluteus medius) transcriptome sequencing was performed for 23 samples collected form Arabian horses , using Illumina HiScan SQ in50 single-end cycles and in 6 technical repetitions repetitions.
Project description:High and moderate intensity endurance exercise alters gene expression in human white blood cells (WBCs), but the understanding of how this effect occurs is limited. To increase our knowledge of the nature of this process, we investigated the effects of passing the anaerobic threshold (AnT) on the gene expression profile in WBCs of athletes. Nineteen highly trained skiers participated in a treadmill test with an incremental step protocol until exhaustion (ramp test to exhaustion; RTE). The average total time to exhaustion was 14:40 min and time after AnT was 4:50 min. Two weeks later, seven of these skiers participated in a moderate treadmill test (MT) at 80% peak O2 uptake for 30 min, which was slightly below their AnTs. Blood samples were obtained before and immediately after both tests. RTE was associated with substantially greater leukocytosis and acidosis than MT. Gene expression in WBCs was measured using whole genome microarray expression analysis before and immediately after each test. A total of 310 upregulated genes were found after RTE, and 69 genes after MT, of which 64 were identical to RTE. Both tests influenced a variety of known gene pathways related to inflammation, stress response, signal transduction and apoptosis. A large group of differentially expressed, and previously unknown, small nucleolar RNA and small Cajal body RNA was found. In conclusion, a 15 min test to exhaustion was associated with substantially greater changes of gene expression than a 30 min test just below the AnT. After a general medical checkup, the subjects performed a ramp-type progressive exercise test on a treadmill. Blood samples were taken from the antecubital vein using indwelling catheter before (T0) and immediately after ramp test to exaustion (T1), and before (T2) and immediately after moderate treadmill test (T3).
Project description:Haploid (1N) and diploid (2N) Emiliania huxley (RCC 1217 and RCC1216) were grown in dilute batch culture and achieved stationary phase. We sampled two time points, at early (T2) and late (T3) stationary phase. Controls are haploid or diploid cells derived from dilute cultures (T1) growing in replete control medium. All samples were hybridized against a common, pooled baseline RNA. All experiments and hybridization were done in independent biological triplicates (A, B and C).
Project description:GalNAc-transferase (GalNAc-T) isoforms modify distinct subsets of the O-glycoproteome and GalNAc-type O-glycosylation is found on most proteins trafficking through the secretory pathway in metazoan cells. The O-glycoproteome is regulated by up to 20 polypeptide GalNAc-Ts and the contributions and biological functions of individual GalNAc-Ts are poorly understood. Here, we used a zinc-finger nuclease (ZFN)-directed knockout strategy to probe the contributions of the major GalNAc-Ts (GalNAc-T1 and T2) in liver cells, and explore how the GalNAc-T repertoire quantitatively affects the O-glycoproteome. We demonstrate that the majority of the O-glycoproteome is covered by redundancy, whereas distinct subsets of substrates are modified by non-redundant functions of GalNAc-T1 and T2. Differential transcriptomic analysis indicates that loss of function of a GalNAc-T induces specific transcriptional response. The non-redundant O-glycoproteome subsets for and the transcriptional responses for each isoform appeared to be related to different cellular processes, and for the GalNAc-T2 isoform supporting a role in lipid metabolism. The results demonstrate that GalNAc-Ts have non-redundant glycosylation functions, and that these may affect distinct cellular processes. The data provides a comprehensive resource for unique substrates for individual GalNAc-Ts. Our study provides a new view on the regulation of the O-glycoproteome, suggesting that the plurality of GalNAc-Ts arose to regulate distinct protein functions and cellular processes.
Project description:Monocytes play a central role in the inflammatory response that follows acute myocardial infarction (MI). In order to study phenotypic adaptation of this cell type, we investigated patterns of monocyte gene expression in circulating monocytes at various stages of MI. Circulating monocytes were isolated from venous blood of MI patients at three time points: t1: within 6 hours after onset of chest pain (acute phase), t2: 3 days after MI (subacute phase), t3: 90 days after MI (chronic phase). For comparison, we studied a control group (n=21, data to be submitted later) with stable coronary artery disease. Using this transcriptomic analysis, we aimed to provide a more comprehensive reference of monocyte biology following acute MI and to aid in the identification of novel pathways and genes influencing the course of MI. Monocytes play a central role in the inflammatory response that follows acute myocardial infarction (MI). In order to study phenotypic adaptation of this cell type, we investigated patterns of monocyte gene expression in circulating monocytes at various stages of MI. Circulating monocytes were isolated from venous blood of MI patients at three time points: t1: within 6 hours after onset of chest pain (acute phase), t2: 3 days after MI (subacute phase), t3: 90 days after MI (chronic phase). For comparison, we studied a control group (n=21, data to be submitted later) with stable coronary artery disease. Illumina Ref-8 v3.0 microarray arrays were used for whole-genome transcriptional profiling.