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Systematic biological analysis of metabolic profile and gene expression changes for assessment of nanotoxicity
| DC Field | Value | Language |
|---|---|---|
| dc.contributor.advisor | 유태현, 이광 | - |
| dc.contributor.author | 신태환 | - |
| dc.date.accessioned | 2022-11-29T03:01:16Z | - |
| dc.date.available | 2022-11-29T03:01:16Z | - |
| dc.date.issued | 2017-02 | - |
| dc.identifier.other | 24095 | - |
| dc.identifier.uri | https://dspace.ajou.ac.kr/handle/2018.oak/20952 | - |
| dc.description | 학위논문(박사)--아주대학교 일반대학원 :분자과학기술학과,2017. 2 | - |
| dc.description.tableofcontents | Chapter 1. General introduction 1 1.1. Nanoparticle exposure 2 1.2. Nanotoxicity 2 1.3. MNPs@SiO2(RITC) 2 Chapter 2. Alterations in polyamine profile and related gene expression level in neuronal cells treated with MNPs@SiO2(RITC) 5 2.1. Abstract 6 2.2. Introduction 7 2.3. Materials and Methods 9 2.3.1. Cell culture 9 2.3.2. Gas chromatography-mass spectrometry 9 2.3.3. RNA purification 9 2.3.4. Reverse transcription PCR (RT-PCR) 10 2.3.5. Statistical analysis 10 2.4. Results 12 2.4.1. Polyamine profile 12 2.4.2 Polyamine metabolism related gene expression 16 2.5 Discussion 18 Chapter 3. Overdosed silica-coated magnetic nanoparticles reduce cell deformability and migratory activity via decrement of membrane fluidity 19 3.1. Abstract 20 3.2. Introduction 21 3.3. Materials and Methods 23 3.3.1. Cell culture 23 3.3.2. Microfluidics measurement of cell deformation index 23 3.3.3. Evaluation of intracellular ROS levels 23 3.3.4. Evaluation of lipid peroxidation 23 3.3.5. Measurement of membrane fluidity 24 3.3.6. Immunocytochemistry 25 3.3.7. Microarray data analysis 25 3.3.8. MTS assay 26 3.3.9. Wound healing assay 26 3.3.10. Transwell invasion assay 26 3.3.11. Atomic force microscopy analysis 27 3.3.12. Gas chromatography-mass spectrometry 27 3.3.13. Statistical analysis 27 3.4. Results 28 3.4.1. Deformability decrement in cells treated with MNPs@SiO2(RITC) 28 3.4.2. Generation of intracellular ROS and lipid peroxidation after treatment with MNPs@SiO2(RITC) 30 3.4.3. Membrane fluidity decreases after treatment with MNPs@SiO2(RITC) in HEK293 and hBM-MSC 32 3.4.4. Cytoskeletal abnormality in MNPs@SiO2(RITC)-treated HEK293 and hBM-MSC 34 3.4.5. Altered expression of lipid peroxidation and cytoskeletal abnormality related genes in cells treated with MNPs@SiO2(RITC) 36 3.4.6. Migratory activity of hBM-MSC were decreased with MNPs@SiO2(RITC) treatment 38 3.5 Discussion 40 Chapter 4. Silica-coated magnetic nanoparticle induces microglia activation and excitotoxic D-serine secretion 45 4.1. Abstract 46 4.2. Introduction 47 4.3. Materials and Methods 48 4.3.1. Primary cell isolation and cell culture 48 4.3.2. Evaluation of morphological activation of microglia 48 4.3.3. Flow cytometry 48 4.3.4. Total RNA isolation and cDNA library preparation for transcriptome sequencing (RNA-seq) 49 4.3.5. Differentially expressed gene analysis using RNA-seq data 49 4.3.6. Gene Ontology analysis and pathway analysis 50 4.3.7. Gas chromatography-mass spectrometry 50 4.3.8. Determination of D-serine secreted from microglia 50 4.3.9. Evaluation of cell viability in microglia co-culture system 50 4.3.10. Measurement of the Adenosine Triphosphate (ATP) concentration 51 4.3.11. Evaluation of proteasome activity 51 4.3.12. Statistical analysis 52 4.4. Results 54 4.4.1. Evaluation of microglia activation in MNPs@SiO2(RITC) treatment 54 4.4.2. Transcriptome analysis of microglia in MNPs@SiO2(RITC) treatment 56 4.4.3. Amino acid profiles change and gene co-expression network and amino acid profiles of microglia in MNPs@SiO2(RITC) treatment 58 4.4.4. Cytotoxic effect of activated microglia on neuronal cells in co-culture system 62 4.5 Discussion 64 CONCLUSION 66 REFERENCES 67 ABSTRACT IN KOREAN 74 | - |
| dc.language.iso | eng | - |
| dc.publisher | The Graduate School, Ajou University | - |
| dc.rights | 아주대학교 논문은 저작권에 의해 보호받습니다. | - |
| dc.title | Systematic biological analysis of metabolic profile and gene expression changes for assessment of nanotoxicity | - |
| dc.title.alternative | Tae Hwan Shin | - |
| dc.type | Thesis | - |
| dc.contributor.affiliation | 아주대학교 일반대학원 | - |
| dc.contributor.alternativeName | Tae Hwan Shin | - |
| dc.contributor.department | 일반대학원 분자과학기술학과 | - |
| dc.date.awarded | 2017. 2 | - |
| dc.description.degree | Doctoral | - |
| dc.identifier.localId | 1199318 | - |
| dc.identifier.url | http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000024095 | - |
| dc.description.alternativeAbstract | The emerging field of nanobiotechnology offers the potential for the development of exquisitely sensitive diagnostics and organ/tumor-targeted therapies [1]. Over the past few decades, there has been considerable interest in developing nanoparticles as effective drug delivery devices [1]. MNPs@SiO2(RITC) are expected to have a wide range of applications in the future because they are easy to synthesize and have good biocompatibility by coated with SiO2 [2]. However, there are limits in assessment of potential toxicity on the conventional method. Here, I report the analysis for more detailed assessment of nanotoxicity with multidisciplinary method which is combined with conventional method. In chapter 1, general introduction was described about nanoparticle exposure, toxicity of nanoparticle, and nanoparticle used in my study in assessment of toxicity. In chapter 2, I assessed the changes in polyamine profile by silica-coated magnetic nanoparticles containing rhodamine B isothiocyanate dye (MNPs@SiO2(RITC)) in human neuroblastoma SH-SY5Y cells. In chapter 3, I investigated the characteristics and in vitro cytotoxicity of human bone marrow-derived mesenchymal stem cells (hBM-MSC) labeled with MNPs@SiO2(RITC). I addressed this issue in the present study by investigating deformation, membrane fluidity, cytoskeletal change, the expression of genes related to lipid peroxidation and abnormal cytoskeleton, and migratory activity of hBM-MSC. In chapter 4, I assessed the microglial MNPs@SiO2(RITC)-induced activation by morphological analysis, surface marker protein analysis, transcriptome analysis, intracellular amino acid profiles, and secreted serine family amino acid and neuronal toxicity in co-culture system with neuronal cells, SH-SY5Y, primary rat cortical and dopaminergic neurons. In conclusion, I studied nanotoxicity in neuronal cell line, stem cells, and primary neuronal cells with combinatorial analysis. That is, it should be taken into consideration in potential chronic toxicity of nanoparticle in usage for human. | - |
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