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GUIDE OAK 국가리포지터리 부경대학교

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Abstract: Oxidative damage occurred by reactive oxygen species (ROS) give rise to not only oxidation of cell membrane lipid but also that of the genetic material called RNA or DNA in cells. Besides, excessive ROS lead to many kind of disease such as abnormal aging, inflammation, cancer, neurodegenerative disorders in living organism. Some researcher have reported uncontrolled activation of microglia in neuronal degeneration and prevention of this phenomenon is one of the target for therapy.
This study compared protective effect of 4-hydroxybenzyl-COS (HB-COS) with ascorbic acid (vitamin C) from oxidative damage by H2O2. It was shown that HB-COS showed similar effect of reducing the cellular oxidation level and effective scavenge ROS, compared with ascorbic acid in murine microglia (BV-2). In addition, HB-COS and ascorbic acid up-regulated antioxidants protein and gene expression level. Furthermore, they suppressed nuclear factor kappa B (NF-κB) and encouraged expression of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) cell signaling molecule related oxidative stress. The results of this study elucidate that how HB-COS can prevent oxidative stress via to regulate the level of the antioxidants and expression of cell signal molecules.

URI: https://repository.pknu.ac.kr:8443/handle/2021.oak/12880
http://pknu.dcollection.net/jsp/common/DcLoOrgPer.jsp?sItemId=000002235116

Table Of Contents: Abstract i
Table of Contents iii
List of Figures v
List of Tables vii
List of abbreviation viii
1. Introduction 1
1.1. Background 1
1.2. Microglia and relationship with Alzheimer’s disease (AD) 4
1.3. Protection system from Reactive oxygen species (ROS) in cells 5
1.4. Cell response to oxidative stress from a cell signaling perspective 11
1.5. Chitin, chitosan and chitooligomer 19
2. Materials and Methods 20
2.1. Materials and chemicals 20
2.2. Synthesis of a 4-hydroxybenzyl-chitooligomer (HB-COS) 20
2.3. Cell culture 21
2.4. Cytotoxicity determination 21
2.5. Nuclear and cytosol protein extraction 22
2.6. Western blot analysis 23
2.7. Isolation of genomic DNA 24
2.8. DNA oxidation assay 24
2.9. Determination of intracellular formation of reactive oxygen species (ROS) by DCFH-DA 25
2.10. Analysis of intracellular ROS by fluorescence-activated cell sorting (Flow cytometry) and fluorescence microscopy 26
2.11. Statistical analysis 27
3. Results and discussion 28
3.1. Structural analysis of HB-COS 28
3.2. Cell viability assay 33
3.3. Intracellular ROS scavenging effect of HB-COS 35
3.4. HB-COS engage the expression level of Nrf2 41
3.5. Effect of HB-COS on antioxidants expression 44
3.6. Effect on expression of inhibitor κB and NF-κB by HB-COS 47
3.7. DNA protective effect of HB-COS 50
4. Conclusion 52
5. References 53
Acknowledgement 67

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