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Natural Compounds, Genetic Variation, and the Gut-Muscle Axis as Novel Therapeutic Targets in DexamethasoneInduced Muscle Atrophy : 덱사메타손 유발 근육 위축에서 천연 화합물, 유전적 변이 및 장근육 축을 새로운 치료 표적으로 탐색

초록/요약 도움말

Skeletal muscle, essential for locomotion, posture, and metabolic regulation, constitutes approximately 40% of total body mass and holds nearly half of the body's protein content. Muscle atrophy—a condition characterized by decreased muscle mass, strength, and fiber cross-sectional area—can result from various pathological conditions such as chronic diseases, cancer, prolonged glucocorticoid exposure, and aging. Sarcopenia, a progressive and generalized skeletal muscle disorder particularly prevalent in the elderly, poses a growing public health burden. Despite its significance, no pharmacological treatments are currently approved, and its pathogenesis remains incompletely understood, warranting the search for novel therapeutic approaches. This dissertation investigates natural product-based therapeutic strategies to counteract muscle atrophy, using a dexamethasone (DEX)-induced model to mimic catabolic muscle wasting. Part 2 evaluates how genetic background modulates muscle and bone responses to Rg3, a ginsenoside compound, across six genetically diverse Collaborative Cross founder mouse strains. The findings highlight strain-specific differences in susceptibility to DEX and responsiveness to Rg3, underscoring the importance of host genetics in treatment outcomes. Part 3 explores the efficacy of Centaurea cyanus extract and its active constituent Graveobioside A in alleviating DEX-induced muscle atrophy. The results demonstrate that CC modulates cannabinoid receptor pathways and beneficially reshapes the gut microbiota to protect muscle integrity. Part 4 investigates Polyporus umbellatus extract and its gut-muscle axis-mediated protective effects, revealing Lactobacillus gasseri as a critical microbial mediator enhancing PU’s anti-atrophic activity. Collectively, this dissertation identifies three promising natural therapeutics—Rg3, Centaurea cyanus extract, and Polyporus umbellatus—for mitigating glucocorticoid-induced muscle atrophy. Beyond simply identifying these compounds, this work provides a comprehensive understanding of the multifaceted nature of muscle wasting. The studies collectively demonstrate the critical interplay among diverse biological systems—from host genetics that dictate an individual’s susceptibility and responsiveness to a compound, to intricate molecular signaling pathways and the profound influence of the gut microbiota. This integrated perspective represents a significant advance over traditional, single-pathway approaches to muscle atrophy research. The findings of this research lay the groundwork for developing more sophisticated, mechanism-based therapeutic strategies. By considering an individual's unique genetic makeup and microbial profile, our work suggests a potential paradigm shift towards personalized interventions for sarcopenia and other related muscle disorders. This research not only contributes to the search for effective treatments but also opens up new avenues for exploring the complex biology of muscle health.

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목차 도움말

1 Introduction 1
1.1 Skeletal muscle and muscle atrophy 1
1.2 Molecular Mechanisms of Muscle Atrophy 2
1.2.1 Protein Synthesis Pathway 2
1.2.2 Protein Degradation Pathway 3
1.3 Sarcopenia disease: An Age-Related Muscle Wasting Disorder 4
1.4 Experimental Models of Muscle Atrophy 6
1.5 Current Therapeutic Approaches and Unmet Need 9
1.6 Rationale and Specific Aims of the Dissertation 10
2 Dexamethasone-induced muscle atrophy and bone loss in six genetically diverse collaborative cross founder strains demonstrates phenotypic variability by Rg3 treatment 13
2.1 Introduction 14
2.2 Materials and methods 19
2.2.1 Animals 19
2.2.2 Grip strength test and treadmill running exercise 20
2.2.3 Biochemical analysis in plasma 20
2.2.4 Histological analysis of skeletal muscle 20
2.2.5 Data analysis and statistics 21
2.3 Results 21
2.3.1 Effect of genetic backgrounds and Rg3 on grip strength, muscle mass, and myofiber size 21
2.3.2 Effect of genetic backgrounds and Rg3 on exercise performance and biomarkers of mitochondrial biogenesis and muscle injury 24
2.3.3 Effect of genetic backgrounds and Rg3 on bone health-related traits 27
2.3.4 Rg3 strengthened the association between musculoskeletal traits and immune function 29
2.3.5 Effect of genetic backgrounds and Rg3 on the expression of key muscle and bone-related genes and trait-gene correlation 31
2.3.6 The immune function of the spleen was not significantly affected by Rg3 treatment or genetic background 35
2.4 Discussion 37
3 Cornflower Extract and Its Active Components Alleviate Dexamethasone-Induced Muscle Wasting by Targeting Cannabinoid Receptors and Modulating Gut Microbiota 48
3.1 Introduction 49
3.2 Materials and methods 50
3.2.1 Plant Materials and Extraction 50
3.2.2 Isolation and Identification of CC-Derived Compounds 50
3.2.3 High-Performance Liquid Chromatography (HPLC) Profiling 51
3.2.4 Chemicals and Apparatus 52
3.2.5 C2C12 Cell Differentiation and Cytotoxicity Assay 52
3.2.6 Quantification of Myofiber 53
3.2.7 Detection of Reduced GSH Measurement 54
3.2.8 Mitotracker Assay 54
3.2.9 Determination of Cellular ATP Content 54
3.2.10 Measurement of ROS Production 54
3.2.11 Computational Docking 55
3.2.12 Mouse Model of DEX-Induced Muscle Atrophy 55
3.2.13 Grip Strength and Exercise Performance Test 56
3.2.14 Histological Analysis of Skeletal Muscle 57
3.2.15 Determination of Muscle ATP and Aconitase Contents 57
3.2.16 RNA Extraction and Quantitative Reverse Transcription–Polymerase Chain Reaction (qRT-PCR) 57
3.2.17 Western Blot Analysis 59
3.2.18 16S rRNA Gene Sequencing in Cecum Samples 60
3.2.19 Statistical Analyses 60
3.3 Results 61
3.3.1 CC Demonstrates Protective Activity against DEX-Induced Atrophy in C2C12 Myotubes 61
3.3.2 CC Mitigates DEX-Induced Oxidative Stress and Boosts Mitochondrial Content in C2C12 Myotubes 64
3.3.3 Chemical Characterization of CC Reveals a Diverse Profile of Bioactive Phytochemicals 67
3.3.4 Multiple Compounds Isolated from CC Exhibit Anti-Atrophic, Antioxidant, and ATP-Stimulatory Properties in DEX-Treated C2C12 Myotubes 69
3.3.5 Graveobioside A Exerts Muscle-Protective Effects through Modulatory Action on Cannabinoid Receptors CB1 and CB2 71
3.3.6 CC Ameliorates DEX-Induced Muscle Atrophy in a Murine Mode 75
3.3.7 CC Enhances Exercise Capacity and Promotes Mitochondrial Function in Skeletal Muscle 78
3.3.8 CC Modulates Gut Microbial Diversity and Composition, Reversing DEX-Induced Dysbiosis 80
3.3.9 Correlation Analysis Suggests a Potential Role for Clostridium Sensu Stricto 1 in Mediating the Muscle-Protective and Performance-Enhancing Effects of CC 84
3.4 Discussion 86
4 Modulation of muscle anabolism and gut microbiota by Polyporus umbellatus extract attenuates dexamethasone-induced muscle atrophy 93
4.1 Introduction 94
4.2 Materials and methods 96
4.2.1 Plant material and Polyporus umbellatus extract (PU) Preparation and characterization 96
4.2.2 Isolation and identification of PU-derived compounds 96
4.2.3 C2C12 cell culture and differentiation 97
4.2.4 Cytotoxicity assay 97
4.2.5 Intracellular glutathione (GSH) level measurement 97
4.2.6 Mitochondrial activity measurement 98
4.2.7 Cellular ATP quantification 98
4.2.8 Immunofluorescence microscopy 98
4.2.9 Intracellular ROS Measurement 99
4.2.10 RNA extraction and quantitative reverse transcription-polymerase chain reaction (qRT-PCR) 99
4.2.11 Lactobacillus gasseri culture and growth rate assessment 100
4.2.12 Animal studies 101
4.2.13 Grip strength and exercise performance test 102
4.2.14 Skeletal muscle histology 103
4.2.15 Gut Microbiome analysis by 16 S rRNA gene sequencing 103
4.2.16 Statistical analysis 104
4.3 Results 104
4.3.1 PU attenuates DEX-Induced atrophy and modulates myogenic regulatory factors and key signaling pathways in C2C12 myotubes 104
4.3.2 PU treatment mitigates DEX-induced oxidative stress and improves mitochondrial function in C2C12 myotubes 107
4.3.3 PU is effective in mitigating DEX-induced muscle atrophy in mice 112
4.3.4 PU mitigates DEX-induced decline in exercise performance and promotes mitochondrial biogenesis 115
4.3.5 PU administration markedly shifts gut microbial composition and diversity in mice 117
4.3.6 Correlation analysis implicates L. gasseri as a potential mediator in the muscle-protective effects of PU 119
4.3.7 L. gasseri mitigates DEX-induced muscle wasting and restores exercise performance in mice by modulating protein homeostasis and enhancing mitochondrial biogenesis 123
4.3.8 L. gasseri exerts targeted modulation of gut microbiota composition, promoting muscle health 126
4.4 Discussion 128
5 Future perspective and Conclusion 132
5.1 Recapitulation of Dissertation Aims and Key Findings 132
5.2 Synthesis and Significance of the Dissertation Findings 133
5.3 Limitations of the Overall Thesis Research 134
5.4 Future Research Perspectives 136
5.5 Overall Conclusions 137
References 139
ACKNOWLEDGEMENTS 155

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