Effect of aerobic exercise and particulate matter exposure duration on the diversity of gut microbiota.

Inhalation of ambient particulate matter (PM) can disrupt the gut microbiome, while exercise independently influences the gut microbiome by promoting beneficial bacteria. In this study, we analyzed changes in gut microbial diversity and composition in response to combined interventions of PM exposure and aerobic exercise, extending up to 12 weeks. This investigation was conducted using mice, categorized into five groups: control group (Con), exercise group (EXE), exercise group followed by 3-day exposure to PM (EXE + 3-day PM), particulate matter exposure (PM), and PM exposure with concurrent treadmill exercise (PME). Notably, the PM group exhibited markedly lower alpha diversity and richness compared to the Con group and our analysis of beta diversity revealed significant variations among the intervention groups. Members of the Lachnospiraceae family showed significant enhancement in the exercise intervention groups (EXE and PME) compared to the Con and PM groups. The biomarker Lactobacillus, Coriobacteraceae, and Anaerofustis were enriched in the EXE group, while Desulfovibrionaceae, Mucispirillum schaedleri, Lactococcus and Anaeroplasma were highly enriched in the PM group. Differential abundance analysis revealed that Paraprevotella, Bacteroides, and Blautia were less abundant in the 12-week PM exposure group than in the 3-day PM exposure group. Moreover, both the 3-day and 12-week PM exposure groups exhibited a reduced relative abundance of Bacteroides uniformis, SMB53, and Staphylococcus compared to non-PM exposure groups. These findings will help delineate the possible roles and associations of altered microbiota resulting from the studied interventions, paving the way for future mechanistic research.

Temporal variations in the gut microbial diversity in response to high-fat diet and exercise.

High-fat diet-induced obesity is a pandemic caused by an inactive lifestyle and increased consumption of Western diets and is a major risk factor for diabetes and cardiovascular diseases. In contrast, exercise can positively influence gut microbial diversity and is linked to a decreased inflammatory state. To understand the gut microbial variations associated with exercise and high-fat diet over time, we conducted a longitudinal study to examine the effect of covariates on gut microbial diversity and composition. Young mice were divided into four groups: Chow-diet (CHD), high-fat diet (HFD), high-fat diet + exercise (HFX), and exercise only (EXE) and underwent experimental intervention for 12 weeks. Fecal samples at week 0 and 12 were collected for DNA extraction, followed by 16S library preparation and sequencing. Data were analyzed using QIIME 2, R and MicrobiomeAnalyst. The Bacteroidetes-to-Firmicutes ratio decreased fivefold in the HFD and HFX groups compared to that in the CHD and EXE groups and increased in the EXE group over time. Alpha diversity was significantly increased in the EXE group longitudinally (p < 0.02), whereas diversity (Shannon, Faith's PD, and Fisher) and richness (ACE) was significantly reduced in the HFD (p < 0.005) and HFX (p < 0.03) groups over time. Beta diversity, based on the Jaccard, Bray-Curtis, and unweighted UniFrac distance metrics, was significant among the groups. Prevotella, Paraprevotella, Candidatus arthromitus, Lactobacillus salivarius, L. reuteri, Roseburia, Bacteroides uniformis, Sutterella, and Corynebacterium were differentially abundant in the chow-diet groups (CHD and EXE). Exercise significantly reduced the proportion of taxa characteristic of a high-fat diet, including Butyricimonas, Ruminococcus gnavus, and Mucispirillum schaedleri. Diet, age, and exercise significantly contributed to explaining the bacterial community structure and diversity in the gut microbiota. Modulating the gut microbiota and maintaining its stability can lead to targeted microbiome therapies to manage chronic and recurrent diseases and infections.

Effects of Particulate Matter Inhalation during Exercise on Oxidative Stress and Mitochondrial Function in Mouse Skeletal Muscle.

Particulate matter (PM) has deleterious consequences not only on the respiratory system but also on essential human organs, such as the heart, blood vessels, kidneys, and liver. However, the effects of PM inhalation on skeletal muscles have yet to be sufficiently elucidated. Female C57BL/6 or mt-Keima transgenic mice were randomly assigned to one of the following four groups: control (CON), PM exposure alone (PM), treadmill exercise (EX), or PM exposure and exercise (PME). Mice in the three-treatment group were subjected to treadmill running (20 m/min, 90 min/day for 1 week) and/or exposure to PM (100 µg/m3). The PM was found to exacerbate oxidative stress and inflammation, both at rest and during exercise, as assessed by the levels of proinflammatory cytokines, manganese-superoxide dismutase activity, and the glutathione/oxidized glutathione ratio. Furthermore, we detected significant increases in the levels of in vivo mitophagy, particularly in the PM group. Compared with the EX group, a significant reduction in the level of mitochondrial DNA was recorded in the PME group. Moreover, PM resulted in a reduction in cytochrome c oxidase activity and an increase in hydrogen peroxide generation. However, exposure to PM had no significant effect on mitochondrial respiration. Collectively, our findings in this study indicate that PM has adverse effects concerning both oxidative stress and inflammatory responses in skeletal muscle and mitochondria, both at rest and during exercise.

Proteomic analysis of the effect of high-fat-diet and voluntary physical activity on mouse liver.

Nonalcoholic fatty liver disease (NALFD), characterized by an abnormal accumulation of triglycerides in hepatocytes, is closely linked to insulin resistance, metabolic syndrome, and changes in lipogenesis in the liver. The accumulation of hepatic lipids can lead to a range of pathologies from mild steatosis to severe cirrhosis. Endurance exercise is known to ameliorate the adverse health effects of NAFLD. Therefore, we aimed to investigate the effect of voluntary wheel running (VWR) on the metabolic changes in the livers of high-fat diet (HFD)-induced NAFLD mice and used LC-MS/MS (Liquid chromatography-mass spectrometry) to determine whether the tested intervention affected the protein expression profiles of the mouse livers. Male C57BL/6 mice were randomly divided into three groups: control (CON), high-fat diet sedentary group (HFD), high-fat diet VWR group (HFX). HFX group performed voluntary wheel running into individually cages, given a high-fat diet for 12 weeks. Food consumption, body weight, and running distance were measured every week. Using 2D (2-dimensional)-gel electrophoresis, we detected and quantitatively analyzed the protein expression with >2.0-fold change in the livers of HFD-fed mice, HFD-fed exercise (HFX) mice, and chow-fed mice. Body weight was significantly increased in HFD compared to CON (P < 0.05). The 2D-gel electrophoresis analysis indicated that there was a difference between CON and HFD groups, showing 31 increased and 27 decreased spots in the total 302 paired spots in the HFD group compared to CON. The analysis showed 43 increased and 17 decreased spots in the total 258 spots in the HFX group compared to CON. Moreover, 12 weeks of VWR showed an increase of 35 and a decrease of 8 spots in a total of 264 paired spots between HFD and HFX. LC-MS/MS of HFD group revealed that proteins involved in ketogenesis, lipid metabolism, and the metabolism of drugs and xenobiotics were upregulated, whereas detoxifying proteins, mitochondrial precursors, transport proteins, proteasomes, and proteins involved in amino acid metabolism were downregulated. On the other hand, VWR counteracted the protein expression profile of HFD-fed mice by upregulating molecular chaperones, gluconeogenesis-, detoxification-, proteasome-, and energy metabolism-related proteins. This study provided a molecular understanding of the HFD- and exercise-induced protein marker expression and presented the beneficial effects of exercise during pathophysiological conditions.

Diet Is a Stronger Covariate than Exercise in Determining Gut Microbial Richness and Diversity.

Obesity is a common metabolic disorder caused by a sedentary lifestyle, and a high-fat and a high-glucose diet in the form of fast foods. High-fat diet-induced obesity is a major cause of diabetes and cardiovascular diseases, whereas exercise and physical activity can ameliorate these disorders. Moreover, exercise and the gut microbiota are known to be interconnected, since exercise can increase the gut microbial diversity and contribute to the beneficial health effects. In this context, we analyzed the effect of diet and exercise on the gut microbiota of mice, by next-generation sequencing of the bacterial V4 region of 16S rRNA. Briefly, mice were divided into four groups: chow-diet (CD), high-fat diet (HFD), high-fat diet + exercise (HFX), and exercise-only (EX). The mice underwent treadmill exercise and diet intervention for 8 weeks, followed by the collection of their feces and DNA extraction for sequencing. The data were analyzed using the QIIME 2 bioinformatics platform and R software to assess their gut microbial composition, richness, and diversity. The Bacteroidetes to Firmicutes ratio was found to be decreased manifold in the HFD and HFX groups compared to the CD and EX groups. The gut microbial richness was comparatively lower in the HFD and HFX groups and higher in the CD and EX groups (ACE, Chao1, and observed OTUs). However, the Shannon alpha diversity index was higher in the HFD and HFX groups than in the CD and EX groups. The beta diversity based on Jaccard, Bray-Curtis, and weighted UniFrac distance metrics was significant among the groups, as measured by PERMANOVA. Paraprevotella, Desulfovibrio, and Lactococcus were the differentially abundant/present genera based on the intervention groups and in addition to these three bacteria, Butyricimonas and Desulfovibrio C21c20 were differentially abundant/present based on diet. Hence, diet significantly contributed to the majority of the changes in the gut microbiota.

Intertwined Relationship of Mitochondrial Metabolism, Gut Microbiome and Exercise Potential.

The microbiome has emerged as a key player contributing significantly to the human physiology over the past decades. The potential microbial niche is largely unexplored in the context of exercise enhancing capacity and the related mitochondrial functions. Physical exercise can influence the gut microbiota composition and diversity, whereas a sedentary lifestyle in association with dysbiosis can lead to reduced well-being and diseases. Here, we have elucidated the importance of diverse microbiota, which is associated with an individual's fitness, and moreover, its connection with the organelle, the mitochondria, which is the hub of energy production, signaling, and cellular homeostasis. Microbial by-products, such as short-chain fatty acids, are produced during regular exercise that can enhance the mitochondrial capacity. Therefore, exercise can be employed as a therapeutic intervention to circumvent or subside various metabolic and mitochondria-related diseases. Alternatively, the microbiome-mitochondria axis can be targeted to enhance exercise performance. This review furthers our understanding about the influence of microbiome on the functional capacity of the mitochondria and exercise performance, and the interplay between them.

Protective effects of extra virgin olive oil and exercise training on rat skeletal muscle against high-fat diet feeding.

A diet high in saturated fat leads to skeletal muscle deteriorations including insulin resistance, mitochondrial dysfunction and muscle fiber atrophy. Consumption of long-chain polyunsaturated fatty acids and exercise have shown promise in ameliorating high-fat diet (HFD)-induced oxidative stress and inflammation. However, the impact of extra virgin olive oil (EVOO) on mitochondrial homeostasis in muscle is largely unknown. This study aimed to investigate whether 12 wks of EVOO feeding alone and in conjunction with endurance training could protect against metabolic and mitochondrial dysfunction rat muscle with HFD. Female Sprague-Dawley rats were divided into 4 groups fed a control diet (C), HFD, EVOO diet, and EVOO diet with training (EVOO+T). Mitochondrial enzyme activity and protein content decreased with HFD compared to C, but were restored with EVOO and EVOO+T. EVOO+T elevated muscle cytochrome c and PGC-1α levels. HFD increased muscle proteolytic markers and protein ubiquitination, whereas these effects were not seen in EVOO and EVOO+T. HFD suppressed mitochondrial fusion protein level while increasing fission protein levels, but were restored with EVOO and EVOO+T. Mitophagy marker PINK1 content decreased with HFD, but was unchanged in EVOO and EVOO+T. EVOO+T upregulated autophagy markers, along with decreased phosphorylated/dephosphorylated FoxO3 ratio. Antioxidants enzyme levels were upregulated by EVOO and EVOO+T, and EVOO+T reduced HFD-induced lipid peroxidation. In conclusion, HFD impaired muscle oxidative capacity, promoted protein ubiquitination and mitochondrial fission, and upregulated autophagy markers. Replacement of HFD with EVOO corrected the observed adverse effects, while exercise training in conjunction with EVOO provided additional protection to the muscle.

Effect of Aerobic Exercise on Oxidative Stress and Inflammatory Response During Particulate Matter Exposure in Mouse Lungs.

Regular exercise provides several health benefits that can improve the cardiovascular and musculoskeletal systems, but clear evidence on the effect of exercise-induced hyperventilation in particulate matter (PM) exposure is still lacking. This study aimed to investigate the effects of exercise in PM exposure on reactive oxygen species (ROS) generation, inflammatory response, and mitochondrial integrity in human lung epithelial cells (A549), as well as in mouse lung tissue. In in vitro experiments, PM treatment was shown to significantly increased ROS production, and reduced cell viability and mitochondrial function in A549 cells. The mice were divided into four groups for an in vivo exercise experiment: control (CON), PM inhalation (PI), PM inhalation during exercise (PIE), and exercise (EX) groups. The PI and PIE groups were exposed to 100 µg/m3 of PM for 1 h per day for a week. The PIE and EX groups performed treadmill exercises every day for 1 h at 20 m/min for a week. The levels of pro-inflammatory markers (IL-6 and TNF-α) were significantly higher in the PI group than in the CON group (P < 0.001 and P < 0.01, respectively). The carbonyl protein level was decreased in EX vs. PI (P < 0.001). Mitochondrial fission (Drp1) content was significantly decreased in the EX vs. CON group (P < 0.01), but anti-mitochondrial fission (P-Drp1 Ser637) was increased in the EX vs. PI group (P < 0.05). Mitochondrial autophagy (mitophagy), which is an assessment of mitochondrial integrity, was markedly increased in PI vs. CON (P < 0.001), but the level was reversed in PIE (P < 0.05). Lung fibrosis was increased in PI vs. CON group (P < 0.001), however, the cells were rescued in the PIE (P < 0.001). The number of apoptotic cells was remarkably increased in the PI vs. CON group (P < 0.001), whereas the level was decreased in the PIE (P < 0.001). Taken together, these results showed that short-term exposure to PM triggers oxidative stress, pro-inflammatory responses, and apoptosis in the lungs, but the PM-induced adverse effects on the lung tissue are not exacerbated by exercise-induced PM hyperventilation but rather has a protective effect.

Voluntary exercise training improves body weight of leptin-deficient ob/ob mice by altering hepatic stearoyl-CoA desaturase 1 and deleted in breast cancer 1 protein levels.

PURPOSE: Deleted in breast cancer 1 (DBC1) ablation causes obesity, and stearoyl-CoA desaturase 1 (SCD1) induces the biosynthesis of monounsaturated fatty acids. This study examined whether voluntary wheel running (VWR) alters SCD-1 and DBC1 protein levels in the liver of leptin-deficient ob/ob mice. METHODS: Twenty-five Ob/Ob mice were divided into two groups (ob/ob-Sed and ob/ob-Ex). The expression of DBC1 and SCD1 in the mouse liver was determined using western blotting. RESULTS: After 10 weeks, VWR significantly reduced body weight without affecting the fatty acid synthase and CD36 protein levels. The average daily running distance was 4.0±1.0 km/day. This improvement was associated with changes in the hepatic SCD1 and DBC1 levels. Hepatic SCD-1 protein levels increased significantly, and DBC1 protein levels decreased in ob/ob-Sed animals. On the other hand, VWR inhibited the obesity-induced increase in SCD1 expression and impaired the obesity-induced decrease in DBC1 expression in the liver of leptin-deficient ob/ob mice. CONCLUSION: This is the first study showing that VWR has strong effects on hepatic SCD1 and DBC1 in ob/ob mice, and provides key insights into the effects of exercise on obesity.

The role of mitochondria in redox signaling of muscle homeostasis.

In the past, contraction-induced production of reactive oxygen species (ROS) has been implicated in oxidative stress to skeletal muscle. As research advances, clear evidence has revealed a more complete role of ROS under both physiologic and pathologic conditions. Central to the role of ROS is the redox signaling pathways that control exercise-induced major physiologic and cellular responses and adaptations, such as mitochondrial biogenesis, mitophagy, mitochondrial morphologic dynamics, antioxidant defense, and inflammation. The current review focuses on how muscle contraction and immobilization may activate or inhibit redox signalings and their impact on muscle mitochondrial homeostasis and physiologic implications.

Avenanthramide C suppresses hypoxia-induced cyclooxygenase-2 expression through sirtuin1 activation in non-small-cell lung cancer cells.

Avenanthramide C (AVC), found mainly in oats, mediates anti-inflammatory activities by reducing the anti-inflammatory cytokine levels. This study investigated the effects of AVC on hypoxia-induced cyclooxygenase-2 (COX-2) expression in A549 cells. AVC suppressed the hypoxia-induced increase in COX-2 protein levels and promoter activity. We also observed that the effects of AVC were reversed by a SIRT1 inhibitor, indicating that the inhibitory effects of AVC on hypoxia-induced COX-2 expression are mediated by SIRT1. Therefore, AVC inhibits the hypoxic induction of COX-2 expression via SIRT1 activation. Our results suggest that AVC could be beneficial for preventing lung inflammation under hypoxia.

Aging alters acetylation status in skeletal and cardiac muscles.

During aging, organs such as skeletal muscle and heart require sufficient NAD+ both as a coenzyme for oxidative-reductive electron transfer and as a substrate for multiple signaling pathways. Sirtuins (SIRTs), a family of NAD+-dependent deacetylase, play an important role in regulating mitochondrial homeostasis and antioxidant defense by deacetylating transcription factors and enzymes such as PGC-1α, p65, GCN5, and SOD2. However, age-related DNA damage and increased SASP activate PARP-1 and CD38, the enzymes competing with SIRTs for NAD+. Thus, it is important to know how aging alters intracellular NAD+ status and NAD+-depending enzyme expression in muscles. In this study, we report that the acetylation level of muscle protein pool, as well as major SIRTs target proteins (PGC-1α, GCN5, p65, and SOD2), was significantly increased in hindlimb and cardiac muscles of 24-month old mice compared with their 6-month old counterparts, despite the fact that most members of the SIRT family were upregulated with aging. Aging increased the protein content of PARP-1 and CD38, whereas decreased NAD+ levels in both skeletal and heart muscles. Aged muscles demonstrated clear signs of mitochondrial dysfunction, oxidative stress, and inflammation. Taken together, our data suggest that despite the upregulation of SIRTs, aged muscles suffered from NAD+ deficit partly due to the competition of elevated CD38 and PARP-1. The enhanced acetylation of several key proteins involved in broad cellular functions may contribute to the age-related muscle deterioration.

Muscle Disuse Atrophy Caused by Discord of Intracellular Signaling.

Significance: Regular contractile activity plays a critical role in maintaining skeletal muscle morphological integrity and physiological function. If the muscle is forced to stop contraction, such as during limb immobilization (IM), the IGF/Akt/mTOR signaling pathway that normally stimulates protein synthesis and inhibits proteolysis will be suppressed, whereas the FoxO-controlled catabolic pathways such as ubiquitin-proteolysis and autophagy/mitophagy will be activated and dominate, resulting in muscle fiber atrophy. Recent Advances: Mitochondria occupy a central position in the regulation of both protein synthesis and degradation through several redox-sensitive pathways, including peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), mitochondrial fusion and fission proteins, mitophagy, and sirtuins. Prolonged IM downregulates PGC-1α due to AMPK (5'-AMP-activated protein kinase) and FoxO activation, thus decreasing mitochondrial biogenesis and causing oxidative damage. Decrease of mitochondrial inner membrane potential and increase of mitochondrial fission can trigger cascades of mitophagy leading to loss of mitochondrial homeostasis (mitostasis), inflammation, and apoptosis. The phenotypic outcomes of these disorders are compromised muscle function and fiber atrophy. Critical Issues: Given the molecular mechanism of the pathogenesis, it is imperative that the integrity of intracellular signaling be restored to prevent the deterioration. So far, overexpression of PGC-1α via transgene and in vivo DNA transfection has been found to be effective in ameliorating mitostasis and reduces IM-induced muscle atrophy. Nutritional supplementation of select amino acids and phytochemicals also provides mechanistic and practical insights into the prevention of muscle disuse atrophy. Future Directions: In light of the importance of mitochondria in regulating the various critical signaling pathways, future work should focus on exploring new epigenetic strategies to restore mitostasis and redox balance.

Foam rolling is an effective recovery tool in trained distance runners.

Purpose: Many endurance athletes use foam rolling (FR) to decrease muscle soreness, but it is unclear whether FR effectively treats soreness in this population. Moreover, the effects of FR in highly trained runners are unknown. The aim of this study was to use downhill running (DHR) to induce muscle soreness in runners and to determine the influence of FR on soreness and running performance when compared to sham compression tights. Methods: Participants performed a running economy (RE) test at 75% of 5-km race speed and a 3-km time trial (TT). In a crossover design, subjects then completed DHR followed by either a FR protocol or wearing sham compression tights. Two days post-DHR, subjects repeated the RE and TT tests. Crossover visits occurred 2-4 weeks later. During RE tests, VO2 and rating of perceived exertion (RPE) were recorded. Passive and active soreness were measured on a scale of 0 (no soreness) to 10 (extreme soreness). Results: Eight runners (aged 31 ± 7 years; four females; VO2peak 57 ± 7 ml kg-1 min-1) completed the study. Both treatment conditions experienced passive (p = 0.026) and active soreness (p = 0.012) induced by DHR. Active soreness 2 days postDHR was significantly lower after FR than after sham compression tights (p = 0.025). With tights, there was a trend for an increased RPE compared to pre-DHR (p = 0.056). Conclusions: Foam rolling decreases leg soreness in well-trained runners and attenuates soreness-related increases in perceived exertion during sub-maximal running.

PGC-1α Overexpression via Local In Vivo Transfection in Mouse Skeletal Muscle.

The overexpression of a specific protein is a common method for investigating the specific biological function of the substance and the mechanism of action. In vivo electrotransfer has been confirmed to be one of the most reliable, efficient and cost-effective way to overexpress a protein in a select biological tissue. Typically, this technique involves a physical injection of plasmid DNA followed by electric pulses across the injection site. Here, we introduce this method that we used to transfect green fluorescent protein (GFP)-tagged PGC-1α plasmid DNA into mouse tibialis anterior (TA) muscle, which attained high transfection efficiency with no muscle damage. To quantify the transfection efficiency, we also demonstrate the visualization of plasmid DNA transfected fibers via immunohistochemical staining on muscle cross sections.

Acute estradiol treatment reduces skeletal muscle protein breakdown markers in early- but not late-postmenopausal women.

OBJECTIVES: Menopause and decline in estradiol (E2) may contribute to sarcopenia (i.e., age-related decline in muscle mass and strength) in women. E2 may directly impact skeletal muscle protein breakdown via estrogen receptor (ER) signaling, primarily ERα. It is not yet known whether: 1) E2 regulates pathways of skeletal muscle protein breakdown; 2) E2-mediated changes in protein breakdown markers are associated with ERα activation and insulin sensitivity; and 3) the effects of E2 on protein breakdown markers differ by increasing time since menopause. STUDY DESIGN: We studied 27 women who were ≤6 years past menopause (early postmenopausal, EPM; n = 13) or ≥10 years past menopause (late postmenopausal, LPM; n = 14). Fasted skeletal muscle samples were collected following 1 week of transdermal E2 or placebo treatment in a randomized cross-over design. MAIN OUTCOME MEASURES: We analyzed for cytosolic protein content of the: 1) structural proteins myosin heavy chain (MHC) and tropomyosin; and 2) protein regulatory markers: protein kinase B (Akt), muscle-specific ring finger protein1 (MuRF1), atrogin1, and forkhead box O3 (FOXO3) using Western blot. RESULTS: In response to acute E2, FOXO3 activation (dephosphorylation) and MuRF1 protein expression decreased in EPM but increased in LPM women (p < 0.05). ERα activation was not associated with these protein breakdown markers, but FOXO3 activation tended to be inversely correlated (r = -0.318, p = 0.065) to insulin sensitivity. CONCLUSIONS: These preliminary studies suggest the effects of E2 on skeletal muscle protein breakdown markers were dependent on time since menopause, which is consistent with our previous study on insulin sensitivity.

Avenanthramides attenuate inflammation and atrophy in muscle cells.

BACKGROUND: Chronic inflammation is an important etiologic mechanism for muscle atrophy. Oat-derived phytochemical avenanthramides (AVAs) have been shown to suppress inflammatory responses in human clinical studies and in several cell lines in vitro, but their role in skeletal muscle is unclear. The aim of this study was to investigate whether AVA treatment can prevent tumor necrosis factor (TNF)-α-induced muscle fiber atrophy in C2C12 cells. METHODS: We treated 70% confluent cells for 24 h with AVA. Then, TNF-α was added to cell-cultured medium. Subsequently, cells were harvested at different time points. The cells were examined using various biochemical techniques for measuring protein, messenger RNA levels, nuclear binding activity, and viability. Fluorescence microscope was used for analysis of the myotube morphology. RESULTS: Cells treated with TNF-α significantly increased nuclear factor κB activation, indicated by a marked decrease of IκB (p < 0.05) and a 6.6-fold increase in p65-DNA binding (p < 0.01); however, 30 µmol of AVA-A, -B, and -C treatment reduced the binding by 33%, 18%, and 19% (p < 0.01), respectively, compared with cells treated with TNF-α without AVA. The interleukin-6 level increased by 2.5 fold (p < 0.01) with TNF-α, but decreased by 24%, 32%, and 28% (p < 0.01), respectively, with AVA-A, -B, and -C. The interleukin-1ß level also showed a 47% increase with TNF-α (p < 0.01), whereas this increment was abolished in all AVA-treated cells. Reactive oxygen species production was 1.3-fold higher in the TNF-α-treated group (p < 0.01) but not in the TNF-α + AVAs groups. Messenger RNA levels of muscle-specific E3 ubiquitin ligase atrogin-1 increased 23% in TNF-α vs. control (p < 0.05) but was decreased by 46%, 34%, and 53% (p < 0.01), respectively, with treatment of AVA-A, -B, and -C. Moreover, TNF-α treatment increased the muscle RING finger 1 messenger RNA level by 76% (p < 0.01); this change was abolished by AVAs. Cells treated with TNF-α demonstrated a reduced proliferation compared with control cells (p < 0.01), but this effect was not seen in TNF-α + AVAs cells. The diameter of the C2C12 myotube decreased by 28% (p < 0.01) with TNF-α, whereas it showed no change when AVAs were included in the cell media. CONCLUSION: These results indicated that AVAs can reduce proinflammatory cytokine and reactive oxygen species production and ameliorate TNF-α-induced myotube atrophy in muscle cells.

Intensified mitophagy in skeletal muscle with aging is downregulated by PGC-1alpha overexpression in vivo.

Mitochondrial dysfunction plays an important role in the etiology of age-related muscle atrophy known as sarcopenia. PGC-1α is positioned at the center of crosstalk in regulating mitochondrial quality control, but its role in mitophagy in aged skeletal muscle is currently unclear. The present study investigated the effects of aging and PGC-1α overexpression via in vivo DNA transfection on key mitophagy protein markers, as well as mitochondrial dynamics related proteins, metabolic function and antioxidant capacity in mouse muscle. C57BL/6J mice at the age of 2 mo (young, Y; N = 14) and 24 mo (old, O; N = 14) were transfected in vivo with either PGC-1α DNA (OE, N = 7) or GFP (N = 7) into the tibialis anterior (TA) muscle followed by electroporation. PINK1 and Parkin protein contents were 3.6 and 1.4-fold higher (P < 0.01), whereas mitochondrial ubiquitination (Ub) increased 1.5-fold (P < 0.05), in O vs. Y mice. PGC-1 OE suppressed PINK and Parkin protein levels by 50-60% (P < 0.01), and decreased Ub by 20% (P < 0.05) in old mice. Aging significantly increased the protein content of LC3II (30%, P < 0.05), p62 (42%, P < 0.05), RheB (5.5-fold, P < 0.01), Beclin-1 (3-fold, P < 0.01) and Mfn2 (~4-fold, P < 0.01) in the TA muscle. However, these age-related increases in mitophagy markers were attenuated by PGC-1α OE. Furthermore, aging dramatically increased Fis-1 protein content by 14-fold (P < 0.01), along with a severe reduction of citrate synthase activity (64%, P < 0.01) and cytochrome c oxidase subunit IV (COXIV) protein content (85%, P < 0.01). PGC-1α OE mitigated the age effects on Fis-1 and Drp-1 (P < 0.05). Moreover, PGC-1α OE enhanced mitochondrial oxidative function and antioxidant enzyme activities, and decreased lipid peroxidation and inner membrane damage found in old mice (P < 0.01). In summary, our data demonstrate that mitophagy protein expression in skeletal muscle was enhanced at old age driven possibly by increased mitochondrial dysfunction, damage, and fission. PGC-1α OE was effective in ameliorating mitochondrial deficits but did not restore muscle fiber atrophy.

Data on in vivo PGC-1alpha overexpression model via local transfection in aged mouse muscle.

The data presented in this article are related to the research paper entitled "Intensified mitophagy in skeletal muscle with aging is downregulated by PGC-1alpha overexpression in vivo" (Yeo et al., 2019). The data explained the surgical procedure of in vivo local transfection by electroporation method in aged mouse tibialis anterior muscle, and plasmid DNA preparation and verification protocol. The data also showed the transfection efficiency levels of GFP or GFP-tagged PGC-1alpha through immunohistochemistry method for frozen muscle cross-sections.

Data on the mode of binding between avenanthramides and IKKß domains in a docking model.

The data presented in this article are related to the research paper entitled "Anti-inflammatory effect of avenanthramides via NF-κB pathways in C2C12 skeletal muscle cells." (Kang et al., in press) [1] This article includes experimental procedures used to analyze the mode of binding between and IkB kinase (IKKß) and avenanthramides which are a group of phenolic alkaloids found in oats. The protein-ligand docking and the computer simulation method of molecular dynamics (MD) for studying the physical interactions of molecules were performed.