Unknown,Transcriptomics,Genomics,Proteomics

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Immunomodulatory Effects of Massage on Non-Perturbed Skeletal Muscle in Rats

ABSTRACT: Microarray was used as a screening tool to determine changes in gene transcription with cyclic compressive loading (CCL) (massage mimetic) applied at varying magnitudes in non-perturbed skeletal muscle. 24 male Wistar Rats were randomly assigned to one of three groups based on magnitude of load: Control: 0N; Low Load: 1.4N, and Moderate Load 4.5N. Animals were anesthetized using isofluorane, postioned lateral-recumbant with one limb secured to a platform with the Tibialis Anterior (TA) muscle facing superiorly. Using a custom CCL device, load was applied along the length of the TA cycled at 15mm proximally and distally at a frequency of 0.5Hz. Each animal received a 30-min bout of CCL once a day for four consecutive days. Rats were euthanized 24 hour following the fourth treatment. TA muscles were harvested and preserved cryogenically. Muscle tissue was homogenized, and RNA was isolated using ToTALLY RNA kit. Microarray was performed using Affymetrix GeneChip system.

ORGANISM(S): Rattus norvegicus

SUBMITTER: Christine Waters-Banker

PROVIDER: E-MEXP-3957 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Belt electrode-skeletal muscle electrical stimulation (B-SES) involves the use of belt-shaped electrodes to simultaneously contract multiple muscle groups. Twitch contractions have been demonstrated to protect against denervation-induced muscle atrophy in rats, possibly via mitochondrial biosynthesis. In this study, we examined whether inducing tetanus contractions with B-SES suppresses muscle atrophy and identified the underlying molecular mechanisms. We evaluated the effects of acute (60 Hz, 5 min) and chronic (60 Hz, 5 min, every alternate day for 1 week) B-SES on the tibialis anterior (TA) and gastrocnemius (GAS) muscles in Sprague Dawley rats using belt electrodes attached to both ankle joints. In acute stimulation, a significant decrease in glycogen content in the left and right TA and GAS was observed, suggesting that B-SES causes simultaneous contractions in multiple muscle groups. B-SES also enhanced p70S6K phosphorylation, an indicator of the mechanistic target of rapamycin complex 1 activity. During chronic stimulations, rats were divided into control (CONT), denervation-induced atrophy (DEN), and DEN+electrically stimulated with B-SES (DEN＋ES) groups. After 7 days of treatment, muscle wet weight (n = 8-11 for each group) and muscle fiber cross-sectional area (CSA, n = 6 for each group) of the TA and GAS muscles were reduced in the DEN and DEN+ES groups compared to those in the CON group. The DEN+ES group showed significantly higher muscle weight and CSA than the DEN group. Although RNA-seq and pathway analysis suggested that mitochondrial and ribosome biogenesis are key events in this phenomenon, mitochondrial content showed no difference. In contrast, ribosomal RNA 28S and 45S (n = 6) levels in the DEN+ES group were higher than those in the DEN group. The mRNA levels of the muscle proteolytic molecules Atrogin-1 and MuRF1 were significantly higher in DEN than in CONT but were suppressed in DEN+ES. In conclusion, tetanic electrical stimulation of both ankles using belt electrodes was effective in preventing denervation-induced atrophy in multiple muscle groups. Unlike twitch contractions, ribosomal biosynthesis plays a key role in tetanic contractions to prevent muscle atrophy.

Project description:Higher-order chromosomal organization for transcription regulation is poorly understood in eukaryotes. Using genome-wide Chromatin Interaction Analysis with Paired-End-Tag sequencing (ChIA-PET), we mapped long-range chromatin interactions associated with RNA polymerase II in human cells and uncovered widespread promoter-centered intragenic, extragenic, and intergenic interactions. These interactions further aggregated into higher-order clusters, wherein proximal and distal genes were engaged through promoter-promoter interactions. Most genes with promoter-promoter interactions were active and transcribed cooperatively, and some interacting promoters could influence each other implying combinatorial complexity of transcriptional controls. Comparative analyses of different cell lines showed that cell-specific chromatin interactions could provide structural frameworks for cell-specific transcription, and suggested significant enrichment of enhancer-promoter interactions for cell-specific functions. Furthermore, genetically-identified disease-associated noncoding elements were found to be spatially engaged with corresponding genes through long-range interactions. Overall, our study provides insights into transcription regulation by three-dimensional chromatin interactions for both housekeeping and cell-specific genes in human cells. RNA polymerase II (RNAPII) bound chromatin interactions were extracted with Chromatin Interaction Analysis with Paired-End Tag (ChIA-PET) sequencing, in order to study the transcription regulations with RNAPII-associated long-range chromatin interactions. Five cell lines, namely MCF7 (ATCC# HTB-22), K562 (ATCC# CCL-243), HCT116 (ATCC# CCL-247), HeLa (ATCC# CCL-2.2), and NB4 (Roussel and Lanotte, 2001) (provided by Dr. Sherman Weissman, Yale University), were grown under standard culture conditions and harvested at log phase. Harvested cells were cross-linked using 1% formaldehyde followed by neutralization with 0.2M glycine. Chromatin was isolated and subjected to ChIA-PET protocol as described in Fullwood et al (Fullwood et al: An oestrogen-receptor-alpha-bound human chromatin interactome. Nature 2009, 462(7269):58-64). The ChIA-PET sequence reads were processed and analyzed using ChIA-PET Tool (Li et al: ChIA-PET tool for comprehensive chromatin interaction analysis with paired-end tag sequencing. Genome Biol 2010, 11(2):R22).

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Immunomodulatory Effects of Massage on Non-Perturbed Skeletal Muscle in Rats

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