RESULTS: Dot blot analyses of exosome-specific proteins (CD9, CD63, and CD81) and TRPS confirmed the presence of isolated exosomes in all samples following SEC elution. Western Blot confirmed the absence of contaminating proteins. Although no differences in exosome concentration were seen following any of the exercise interventions, exosome profiling showed a decrease in exosomes associated with cancer metastasis (MCSP) between pre-(0.029 ± 0.01) and post-intervention (0.017 ± 0.019, p=0.03). The exosome expression of the monocyte and macrophage marker was also reduced from 0.023 ± 0.029 to 0.007 ± 0.019 (p=0.02) post-intervention in all groups. CONCLUSION: This evaluation and validation of methods have shown that exosome isolation, characterization, and profiling can be achieved with minimal volume. Funded by NIH R21AG058181.
Sarcopenic obesity (SO) differentiates itself from sarcopenia and obesity by exhibiting a more detrimental and complex condition that increases the risk of mortality in aging individuals. Sarcopenic obese individuals suffer from greater physical disability and muscle weakness than sarcopenic and obese individuals alone. In addition, altered cellular signaling that regulates increased muscle loss in SO is largely unknown. Identifying key differentially expressed genes in SO muscle are important for future investigation and the creation of therapeutics and strategies aimed at attenuating or preventing muscle loss. PURPOSE: To examine global gene expression in muscle of SO mice to determine differentially expressed (DE) genes. METHODS: After weaning, twenty-four mice were randomly assigned to young (3-4 months old) or aged (22-24 months old) groups and then randomly assigned to either a high-fat (HFD, 60% fat) or normal chow (NC, 14% fat) diet. RNA was isolated from the plantaris muscle and Next Generation RNA sequencing and bioinformatics was conducted to determine DE genes among experimental groups. Log 2 FoldChange (FC) ≥ 0.6, Log 2 FC ≤ -0.6 and p ≤ 0.05 was the criteria used to determine significant DE genes. Real-Time Polymerase Chain Reaction was used to confirm mRNA expression of DE genes. RESULTS: Aged-HFD mice had an ̴ 39-49% (p ≤ 0.05) reduction in plantaris to body mass ratio compared to all other groups. CYP1A1, RAB15, and CDH22 showed significant main effects for diet. Age and diet interacted to alter GDF5 mRNA abundance. There was a ̴ 26-fold increase (p ≤ 0.05) in GDF-5 mRNA abundance in the aged-HFD mice compared to all other groups. CONCLUSION: GDF-5 is associated with TGF-B and SMAD signaling. The increased GDF-5 mRNA abundance seen in SO mice could be promoting increased collagen deposition within skeletal muscle affecting muscle function. These findings show that GDF5 may play a key role in altered cellular signaling and muscle atrophy in SO.
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