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Title Page

Contents

Abstract 13

1. INTRODUCTION 15

1-1. Nonlinear optics 15

1-2. Two-photon excited fluorescence 15

1-3. Second-harmonic generation 16

1-4. Coherent anti-Stokes Raman scattering 17

1-5. Multimodal nonlinear optical imaging system 18

1-6. Aims 19

Application 1. Spatiotemporal impairment of perivascular adipose tissue in atherosclerosis development using multimodal nonlinear optical microscopy 20

2-1. INTRODUCTION 21

2-1-1. Atherosclerosis 21

2-1-2. Perivascular adipose tissue 22

2-1-3. PVAT functions on atherosclerosis 23

2-1-4. Transforming growth factor-beta signaling pathway 24

2-1-5. Application of MNLO microscopy in PVAT 24

2-2. MATERIALS AND METHODS 25

2-3. RESULTS 32

2-3-1. Application of MNLO imaging to characterize of tPVAT 32

2-3-2. Characterization of tPVAT based on the CARS image-based LD analysis 34

2-3-3. Multilocular BAT features of LDs in atherosclerotic tPVAT 37

2-3-4. BAT transition of tPVAT morphology during atherosclerosis progression 40

2-3-5. The expression of BAT markers in atherosclerotic tPVAT 43

2-3-6. Gene expression analysis of atherosclerotic tPVAT by mRNA microarray 45

2-3-7. The association of the BAT transition and fibrosis of atherosclerotic tPVAT with the activation of TGF-β signaling pathway 47

2-3-8. Site-specific features of abnormal LDs, fibrosis, and TGF-β signaling in tPVAT dependent on the atherosclerotic plaques 52

2-3-9. Inflammation of tPVAT in the late stage of atherogenesis 57

2-4. DISCUSSION 58

Application 2. Deposition and penetration of airborne particulate matter on human skin using multimodal nonlinear optical imaging 63

3-1. INTRODUCTION 64

3-1-1. Particulate matter 64

3-1-2. Skin anatomy and function 64

3-1-3. Particulate matter and skin diseases 66

3-1-4. Technology to visualize particulate matter on the skin 66

3-1-5. Application of MNLO microscopy for visualizing particulate matter on human skin 67

3-2. MATERIALS AND METHODS 68

3-3. RESULTS 71

3-3-1. Characterization of airborne collected PM2.5 by Raman microscopy[이미지참조] 71

3-3-2. Visualization of PM2.5 by MNLO imaging[이미지참조] 73

3-3-3. En face MNLO 3D imaging of PM2.5 on human skin[이미지참조] 75

3-3-4. PM2.5 deposition dependent on skin morphology[이미지참조] 77

3-3-5. PM2.5 deposition on skin surface by SC barrier disruption[이미지참조] 79

3-3-6. Percutaneous penetration of PM2.5 deepens with severe skin damage[이미지참조] 81

3-3-7. Inflammatory response by PM2.5 exposure with SC barrier disruption[이미지참조] 84

3-4. DISCUSSION 86

References 90

논문요약 104

List of Tables

Table 2-1. Oligonucleotide primers for Real-time PCR 31

Table 2-2. The analysis of AD-specific group in ApoE-/- by KEGG pathway database[이미지참조] 46

Table 3-1. Raman peaks identified for PM2.5 (AP) and SRM 2786.[이미지참조] 72

List of Figures

Figure 1-1. Energy level diagrams of TPEF, SHG and CARS. 17

Figure 1-2. Schematic of multimodal nonlinear optical (MNLO) microscopy. 19

Figure 2-1. The model of atherosclerosis development. 21

Figure 2-2. Overview of the characteristics of different adipose tissues. 22

Figure 2-3. PVAT roles in vascular remodeling. 23

Figure 2-4. En face CARS imaging of tPVAT using MNLO microscopy. 33

Figure 2-5. Characterization of tPVAT, intBAT, and epiWAT by the CARS image-... 35

Figure 2-6. En face CARS images of label-free lipids in tPVAT via horizontal tilling. 36

Figure 2-7. En face label-free MNLO imaging of tPVAT in the atherogenic model. 38

Figure 2-8. Cholesterol levels from ND- and AD-C57BL/6 and ApoE-/-.[이미지참조] 39

Figure 2-9. En face label-free CARS imaging of tPVAT in the atherosclerosis... 41

Figure 2-10. En face label-free MNLO imaging of thoracic aorta in the atherosclerosis... 42

Figure 2-11. The analysis of brown fat features in atherosclerotic tPVAT. 44

Figure 2-12. The analysis of brown fat marker expression in atherosclerotic tPVAT. 44

Figure 2-13. Gene profiling analysis of TGF-β signaling pathway in atherosclerotic... 46

Figure 2-14. Oil red O staining for ex vivo differentiated tPVAT. 47

Figure 2-15. Label-free MNLO images of ex vivo differentiated tPVAT. 48

Figure 2-16. The quantitative analysis of MNLO images and gene expressions in ex... 48

Figure 2-17. Oil red O staining for ex vivo differentiated tPVAT from ApoE-/- via...[이미지참조] 49

Figure 2-18. Label-free MNLO images of ex vivo differentiated tPVAT from ApoE-...[이미지참조] 50

Figure 2-19. The quantitative analysis of MNLO images and gene expressions in ex... 50

Figure 2-20. Label-free MNLO images of ex vivo differentiated tPVAT from LDLR... 51

Figure 2-21. The atherosclerotic plaque formation of thoracic aorta with tPVAT. 53

Figure 2-22. Confocal imaging and MNLO imaging of tPVAT in ROIs. 53

Figure 2-23. The quantitative analysis of confocal and MNLO images. 54

Figure 2-24. Confocal images of UCP-1 expression in a longitudinally sectioned thoracic aorta. 54

Figure 2-25. Site-specific features of abnormal LDs and fibrosis in tPVAT dependent... 56

Figure 2-26. Site-specific impairment of tPVAT in advanced atherosclerotic vessel. 56

Figure 2-27. Inflammation of tPVAT in the late stage of atherogenesis. 57

Figure 2-28. A model of the tPVAT changes depend on the state of atherosclerosis development. 58

Figure 2-29. The analysis of tPVAT changes by β3-AR agonist CL316243. 60

Figure 3-1. Human skin anatomy. 65

Figure 3-2. Raman spectra of PM2.5 (AP).[이미지참조] 72

Figure 3-3. Schematic illustration of fine tunable... 74

Figure 3-4. Label-free PM2.5 imaging by MNLO microscopy.[이미지참조] 74

Figure 3-5. En face MNLO 3D imaging for PM2.5 in human skin biopsy.[이미지참조] 76

Figure 3-6. Topology analysis of human skin from en face MNLO 3D images. 78

Figure 3-7. Visualization of PM2.5 deposition on skin surface by SC barrier disruption.[이미지참조] 79

Figure 3-8. The analysis of PM2.5 deposition on skin surface by SC barrier disruption.[이미지참조] 80

Figure 3-9. Visualization of PM2.5 penetration into skin by SC barrier disruption.[이미지참조] 81

Figure 3-10. The analysis of PM2.5 penetration into skin by SC barrier disruption[이미지참조] 83

Figure 3-11. The measurement of the distance between the skin surface and... 83

Figure 3-12. The measurement of secreted cytokines by PM2.5 exposure and SC barrier disruption.[이미지참조] 84

Figure 3-13. The expression of inflammatory genes by PM2.5 exposure and SC barrier disruption.[이미지참조] 85

Figure 3-14. A model of the PM deposition and penetration on human skin. 86

Figure 3-15. En face 3D MNLO imaging of human skin treated with PAH. 87

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