Control of Mitochondrial Homeostasis In Disused and Aging Skeletal Muscle: Role of Pgc-1Alpha and Sirtuins

University of Minnesota Ph.D. dissertation.June 2020. Major: Kinesiology. Advisors: Li Li Ji, Dawn Lowe. 1 computer file (PDF); viii, 85 pages.Prolonged period of muscle disuse circumstances such as limb immobilization and bedrest are associated with loss of skeletal muscle mass and function (Kandarian et al., 2006). This disuse-induced skeletal muscle atrophy has an important clinical implication to the elderly population because it is directly related to increased mortality and a decreased quality of life (Christensen et al., 1982). In the elderly population, age-dependent muscle loss is a serious health threat known as sarcopenia (Rosenberg, 1997). Thus, the elderly population is much vulnerable to the muscle disuse atrophy. Furthermore, age-related muscle loss leads to increased incidence of falls and fractures (Mithal et al., 2012). Epidemiological evidences report that hip fractures can lead to serious mobility impairment and thus have a strong correlation with mortality (Kim et al., 2012; Cornwall et al., 2004). Indeed, 34% of elderly hip fracture patients dying within 1 year of surgery and this possibility increases with age (Holt et al., 2012). Therefore, enhancing our understanding of the underlying mechanisms for skeletal muscle atrophy in elderly population is vital in order to develop prevention and therapeutic treatments to these problems. Mitochondrial dysfunction with advanced age plays an important role in sarcopenia due to its critical roles including ATP production, redox homeostasis as well as apoptosis (Handchin & Spiegelman, 2008). In the past decade, the peroxisome proliferator-activated receptor-ɤ coactivator-1α (PGC-1α) and sirtuins (SIRTs) emerged as key regulators for transcriptional and/or post-translational modifications, which are involved in mitochondrial biogenesis, dynamics, antioxidant defense system as well as mitophagy (Kim et al., 2017). PGC-1α activates mitochondrial biogenesis via coactivating nuclear-encoded mitochondrial protein transcription factors, and stimulates replication of mitochondrial DNA (mtDNA) (Lin et al., 2005). PGC-1α also has a regulatory mechanism for the expression of antioxidant enzymes such as SOD2, Catalase, and GPx1 (Wenz et al., 2009). In addition, recent studies have shown that PGC-1α is highly involved in mitochondrial fusion/fission and mitophagy machinery to maintain mitochondrial homeostasis, through interacting with mitochondrial fusion proteins (Mfn1/2 and Opa-1) and FoxO transcription factors (Sakellariou et al., 2016). SIRTs are NAD+-dependent protein deacetylases that, along with GCN5, plays a major role in controlling acetylation/deacetylation status of important intracellular proteins (Blander and Guarente, 2002). For example, deacetylation activity of SIRT1 is required for PGC-1α mediated mitochondrial biogenesis in skeletal muscles (Lagouge et al., 2006). Activity of NFkB and FoxO transcription factors, the key controllers for pro-inflammatory cytokine expression and mitophagy, are also regulated by SIRTs (Yeung et al 2004; Kawahara et al., 2009). SIRTs can also activates certain mitochondrial antioxidant enzymes (such as SOD2), enzymes in the TCA cycle and ETC via protein deacetylation resulting in enhancing mitochondrial oxidative phosphorylation (Lombard et al., 2007). Several experimental procedures have been utilized to boost PGC-1α and/or SIRTs activities to improve mitochondrial function and antioxidant capacity in various age-related diseases, such as neurodegenerative diseases, chronic inflammation, type 2 diabetes, and cardio vascular diseases (Rodgers et al., 2008; Wenz, 2011). Overexpression of PGC-1α and/or SIRTs is intended to ameliorate mitochondrial dysregulation and protein breakdown in aged skeletal muscles. Currently, however, direct evidences in the role of PGC-1α and SIRTs on aged muscles are limited. Therefore, this dissertation aims to investigate the impact of PGC-1α and SIRTs on skeletal muscle under two pathophysiological conditions, immobilization and aging. The work will focus on muscle mitochondrial quality control by investigating mitochondrial biogenesis, fusion and fission dynamics, mitophagy and oxidative capacity. The dissertation consists of three studies. The first study investigated the effect of muscle immobilization on mitochondrial fission/fusion and mitophagy in mice. The second study investigated the effect of PGC-1α overexpression on mitochondrial homeostasis in aged skeletal muscle in mice. The third study elucidated the impact of protein acetylation during aging in skeletal and cardiac muscles and the role of SIRTs and NAD+, the limiting factor of protein deacetylation

Maintenance of NAD+ Homeostasis in Skeletal Muscle during Aging and Exercise

Nicotinamide adenine dinucleotide (NAD) is a versatile chemical compound serving as a coenzyme in metabolic pathways and as a substrate to support the enzymatic functions of sirtuins (SIRTs), poly (ADP-ribose) polymerase-1 (PARP-1), and cyclic ADP ribose hydrolase (CD38). Under normal physiological conditions, NAD+ consumption is matched by its synthesis primarily via the salvage pathway catalyzed by nicotinamide phosphoribosyltransferase (NAMPT). However, aging and muscular contraction enhance NAD+ utilization, whereas NAD+ replenishment is limited by cellular sources of NAD+ precursors and/or enzyme expression. This paper will briefly review NAD+ metabolic functions, its roles in regulating cell signaling, mechanisms of its degradation and biosynthesis, and major challenges to maintaining its cellular level in skeletal muscle. The effects of aging, physical exercise, and dietary supplementation on NAD+ homeostasis will be highlighted based on recent literature

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-1β 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. Keywords: Atrogenes, Interleukin, NF-κB, Skeletal muscle, TNF-

Data on the mode of binding between avenanthramides and IKKβ 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 (IKKβ) 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

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

The impact of pulsed electromagnetic field therapy on blood pressure and circulating nitric oxide levels: a double blind, randomized study in subjects with metabolic syndrome

Purpose: Regulation of blood pressure (BP) is important in reducing the risk for cardiovascular disease. There is growing interest in non-pharmacological methods to treat BP including a novel approach using pulsed electromagnetic field therapy (PEMF). PEMF therapy has been proposed to impact physiological function at the cellular and tissue level and one possible mechanism is through an impact on endothelial function and nitric oxide (NO) related pathways. The focus of the present study was to evaluate the effect of PEMF on BP and NO in subjects with mild to moderate metabolic syndrome. Materials and methods: For 12 weeks, 23 subjects underwent PEMF therapy and 21 subjects underwent sham therapy. BP was measured at rest and near the end of submaximal exercise pre- and 12 week post-therapy. Additionally, plasma NO was measured at similar time points. Results: The PEMF demonstrated an increase in NO after therapy (p = .04) but SHAM did not (p = .37). For resting BP, there were no differences in systolic BP (SBP), diastolic BP (DBP) or mean arterial pressure (MAP) between groups (p > .05). During exercise, PEMF had a reduction in peak SBP (p = .04), but not SHAM (p = .57). PEMF demonstrated significant relationships between baseline SBP and change in SBP following therapy (r = −0.71, p  .05). Conclusion: PEMF may increase plasma NO availability and improve BP at rest and during exercise. However, this beneficial effect appears to be more pronounced in subjects with existing hypertension

Factors Associated with Emergency Department Visits and Consequent Hospitalization and Death in Korea Using a Population-Based National Health Database

We aim to investigate common diagnoses and risk factors for emergency department (ED) visits as well as those for hospitalization and death after ED visits. This study describes the clinical course of ED visits by using the 2014&ndash;2015 population data retrieved from the National Health Insurance Service. Sociodemographic, medical, and behavioral factors were analyzed through multiple logistic regression. Older people were more likely to be hospitalized or to die after an ED visit, but younger people showed a higher risk for ED visits. Females were at a higher risk for ED visits, but males were at a higher risk for ED-associated hospitalization and death. Individuals in the highest quartile of income had a lower risk of ED death relative to lowest income level individuals. Disabilities, comorbidities, and medical issues, including previous ED visits or prior hospitalizations, were risk factors for all ED-related outcomes. Unhealthy behaviors, including current smoking, heavy alcohol consumption, and not engaging in regular exercise, were also significantly associated with ED visits, hospitalization, and death. Common diagnoses and risk factors for ED visits and post-visit hospitalization and death found in this study provide a perspective from which to establish health polices for the emergency medical care system

Additional file 1 of Exercise training attenuates pulmonary inflammation and mitochondrial dysfunction in a mouse model of high-fat high-carbohydrate-induced NAFLD

Additional file 1. The ARRIVE Essential 10: Compliance Questionnaire

Additional file 2 of Exercise training attenuates pulmonary inflammation and mitochondrial dysfunction in a mouse model of high-fat high-carbohydrate-induced NAFLD

Additional file 2

Absorption and Elimination of Oat Avenanthramides in Humans after Acute Consumption of Oat Cookies

Background. Avenanthramides (AVA) are a group of diphenolic acids found only in oats that have anti-inflammatory and antioxidant effects. Absorption of AVAs in humans after oral consumption of natural oat flour is unknown. Objective. To examine the appearance of AVAs in plasma after oral ingestion of oat cookies and estimate key pharmacokinetic parameters. Methods. Male and female nonobese participants (n=16) consumed three cookies made with oat flour containing high (229.6 mg/kg, H-AVA) or low (32.7 mg/kg, L-AVA) amounts of AVAs, including AVA-A, AVA-B, and AVA-C. Blood samples were collected at 0, 0.5, 1, 2, 3, 5, and 10 h after ingestion. Plasma total (conjugated and free) AVA concentrations were quantified using UPLC-MS, and pharmacokinetic parameters for each AVA were estimated. Results. AVAs reached peak concentrations in plasma between 2 and 3 h for the H-AVA group and between 1 and 2 h for the L-AVA group. Maximal plasma concentrations for AVAs were higher in the H-AVA than in the L-AVA group. AVA-B demonstrated a longer half-life and slower elimination rate than AVA-A and AVA-C. Conclusions. AVAs found naturally in oats are absorbed in the plasma after oral administration in humans. AVA-B has the slowest elimination rate and the longest half-life compared to AVA-A and AVA-C, while AVA-C demonstrated the lowest plasma concentrations. This study is registered with ClinicalTrials.gov identifier NCT02415374