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Studies on the effect of oxidative stress on muscle atrophy in cultured C2C12 myotubes
| DC Field | Value | Language |
|---|---|---|
| dc.contributor.advisor | 김혜선 | - |
| dc.contributor.author | 김다정 | - |
| dc.date.accessioned | 2019-04-01T16:41:23Z | - |
| dc.date.available | 2019-04-01T16:41:23Z | - |
| dc.date.issued | 2019-02 | - |
| dc.identifier.other | 28535 | - |
| dc.identifier.uri | https://dspace.ajou.ac.kr/handle/2018.oak/15067 | - |
| dc.description | 학위논문(석사)--아주대학교 일반대학원 :생명과학과,2019. 2 | - |
| dc.description.tableofcontents | Ⅰ. Introduction 1 Ⅱ. Materials and methods 6 1. Materials 6 2. Cell culture 7 3. Measurement of myotube diameter 7 4. Detection of intracellular ROS 7 5. Oxidized protein detection 8 6. Oxidative stress 8 7. RNA isolation and cDNA construction 8 8. Real time-PCR and conventional PCR 9 9. Western blot analysis 11 10. Statistical analysis 12 Ⅲ. Results 13 1. The atrophic effect of serum starvation with HBSS in C2C12 myotubes. 13 1.1 Serum starvation with HBSS reduced myotube diameter in a time-dependent manner. 13 1.2 Serum starvation with HBSS induced muscle atrophy via Akt/Foxo/Atrogin-1 signaling pathway. 13 2. The atrophic effect of Dexamethasone (Dex) treatment in C2C12 myotubes. 16 2.1 Dex treatment decreased diameter of myotubes in a dose-dependent manner. 16 2.2 Dex treatment increased muscle specific E3 ligases via Akt/Foxo signaling pathway. 16 3. An increase in oxidative stress during starvation- and Dex-induced muscle atrophy in C2C12 myotubes. 19 3.1 Both serum starvation and Dex treatment increased intracellular ROS. 19 3.2 Both serum starvation and Dex treatment slightly increased in protein oxidation. 19 4. The atrophic effect of menadione-induced oxidative stress in C2C12 myotubes. 22 4.1 Menadione treatment reduced myotube diameter in a time-dependent manner. 22 4.2Menadione treatment evokes muscle atrophy by increasing Atrogin-1 via Foxo signaling pathway. 22 4.3 Ubiquitinated proteins were increased by menadione-induced oxidative stress in C2C12 myotubes. 22 5. The involvement of Calpain-2 system in relationship between oxidative stress and muscle atrophy in C2C12 myotubes. 27 6. The impairment of partial antioxidant systems in atrophy-induced C2C12 myotubes. 29 7. The involvement of Nrf2 signaling pathway in down-regulation of antioxidant systems during muscle atrophy in C2C12 myotubes. 31 7.1 Keap1 expression was increased in atrophy-induced myoutbes. 31 7.2 Phosphorylation of nuclear Nrf2 was reduced in atrophy-induced myotubes. 31 7.3 Protein kinase C delta (PKC-δ) was reduced in Dex-induced muscle atrophy in a dose-dependent manner. 31 7.4 Pre-treatment of sulforaphane prevented serum starvation-induced muscle atrophy by reducing oxidative stress. 32 Ⅳ. Discussion 37 Ⅴ. References 40 Ⅵ. 국문요약 44 | - |
| dc.language.iso | eng | - |
| dc.publisher | The Graduate School, Ajou University | - |
| dc.rights | 아주대학교 논문은 저작권에 의해 보호받습니다. | - |
| dc.title | Studies on the effect of oxidative stress on muscle atrophy in cultured C2C12 myotubes | - |
| dc.title.alternative | Kim Da Jeong | - |
| dc.type | Thesis | - |
| dc.contributor.affiliation | 아주대학교 일반대학원 | - |
| dc.contributor.alternativeName | Kim Da Jeong | - |
| dc.contributor.department | 일반대학원 생명과학과 | - |
| dc.date.awarded | 2019. 2 | - |
| dc.description.degree | Master | - |
| dc.identifier.localId | 905307 | - |
| dc.identifier.uci | I804:41038-000000028535 | - |
| dc.identifier.url | http://dcoll.ajou.ac.kr:9080/dcollection/common/orgView/000000028535 | - |
| dc.description.alternativeAbstract | Skeletal muscle atrophy is defined as shrinkage of muscle fiber and decrease in muscle strength. It is accompanied by many causes such as lack of physical activity, injury, malnutrition, long-term exposure of corticosteroid, heart failure, cancer and aging. At the cellular aspect, muscle atrophy is occurred owing to increase in protein degradation or decrease in protein synthesis. One of the process of muscle atrophy is mediated by the activation of the ubiquitin-proteasome proteolysis pathway. Especially, muscle specific E3 ubiquitin ligases, atrogin-1/MAFbx and MuRF1, are up-regulated during muscle atrophy. Forkhead box O (Foxo) family of transcription factor is the well-known mediator of atrophy, which induces expression of atrogin-1/MAFbx and MuRF1. Akt is an upstream molecule of Foxo. During muscle atrophy, Akt is inactivated and therefore activated Foxo translocates to the nucleus for induction of atrophy-related genes. To evoke muscle atrophy, C2C12 myotubes were starved by incubating in Hank’s balanced salt solution or were treated with synthetic glucocorticoid dexamethasone. Both serum starvation and treatment of dexamethasone reduced diameter of myotube via Akt/Foxo/Atrogin-1 and MuRF1 pathway. Also, an increase in reactive oxygen species was observed in atrophy-induced myotubes. To study the effect of oxidative stress on muscle atrophy in vitro, we induced oxidative stress by treating menadione on C2C12 myotubes. Treatment of menadione reduced diameter of myotube and increased expression of Atrogin-1 via Foxo pathway. Although it is known that oxidative stress can induce muscle atrophy, it is needed to more explain about the mechanism of oxidative stress-induced muscle atrophy. The nuclear factor erythroid 2-related factor 2 (Nrf2) is the major regulator of antioxidant responses. It induces expression of many antioxidant enzymes such as NAD(P)H quinone oxidoreductase 1, Heme oxygenas-1 and Glutamate-cysteine ligase and so on. Kelch-like ECH-associated protein 1 (Keap1) represses Nrf2 by binding to Nrf2 in cytoplasm. We identified that protein expression of Keap1 was increased in atrophy-induced C2C12. As a result, Nrf2 activity was decreased and several antioxidant enzymes including Nqo1 and Hmox1 were also down-regulated. These results suggest that menadione-induced oxidative stress can be a trigger of muscle atrophy and vice versa. Atrophy-induced myotubes lost oxidant tolerance by decreasing Nrf2 activity. It is indicated that muscle atrophy can be improved by up-regulating impaired antioxidant systems via Nrf2 signaling pathway. | - |
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- Graduate School of Ajou University > Department of Bioscience > 3. Theses(Master)
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