Title Page
Contents
Abstract 12
CHAPTER 1. Antagonistic Effect of Magnesium Hydroxide Particles on Vascular Endothelial Activation Induced by Acidic PLGA Degradation Product 17
I. INTRODUCTION 17
II. MATERIALS AND METHODS 21
1. Preparation of PLGA films and degradation products 21
2. Cells and cytotoxicity assays 21
3. RNA-sequencing analysis 22
4. Measurement of pH in vitro and in vivo 23
5. Measurement of magnesium ion (Mg2+) concentration[이미지참조] 23
6. Determination of intracellular oxidative stress 24
7. Leukocyte binding assay 24
8. Western blot analysis 25
9. Cytokine array and ELISA 25
10. RNA extraction, and real-time quantitative PCR 26
11. RNA interference 27
12. In vivo experiments and histological analysis 27
13. Characterization of PLGA/MH composites 28
14. Statistical analysis 29
III. RESULTS 30
1. Protective effect of MH particles on the acidic environment caused by PLGA in vivo and in vitro 30
2. Transcriptional regulation of PLGA degradation products 34
3. Defensive effect of MH particles on PLGA-induced inflammation 36
4. Antagonistic effect of MH particles on influx of PLGA by-products 46
5. Molecular mechanism of PLGA-induced endothelial activation 50
6. A role of magnesium ion in maintaining endothelial function 55
IV. DISCUSSION 58
V. CONCLUSION 62
REFERENCES 63
CHAPTER 2. An Integrated Bioactive Scaffold with PDRN and Inflammatory-primed MSC-EVs for Kidney Regeneration 72
I. INTRODUCTION 72
II. MATERIALS AND METHODS 76
1. Cell culture 76
2. EV isolation and characterization 76
3. EV internalization assay 77
4. Cytokine array 78
5. Wound healing assay 78
6. Tube formation assay 78
7. Macrophage polarization 79
8. Immunocytochemistry 79
9. Western blot analysis 80
10. Real-time quantitative PCR 80
11. Scaffold fabrication 81
12. Characterization of scaffold 81
13. In vitro and in vivo biocompatibility tests 82
14. In Vivo Model Design 83
15. Histological and Functional Analysis 83
16. Statistical analysis 84
III. RESULTS 85
1. Isolation and characterization of EVs 85
2. Synergistic effect of PDRN and TI-EVs on pro-regenerative processes 90
3. Fabrication and characterization of multi-functional composite scaffolds 96
4. Biocompatibility and bioactivity of composite scaffolds 98
5. In vivo evaluation of kidney tissue regeneration capacities of scaffolds 101
6. In vivo evaluation of kidney function restoration 105
IV. DISCUSSION 108
V. CONCLUSION 113
REFERENCES 116
ABSTRACT IN KOREAN 125
Table 1. Water contact angle for fabricated scaffolds 114
Table 2. List of primers sequences used for quantitative real-time PCR analysis 115
CHAPTER 1 10
Figure 1. Effect of MH particles on cytotoxicity of PLGA by-products. (A)... 32
Figure 2. Characteristics of DEGs. (A) Hierarchical clustering of 364 DEGs in the... 35
Figure 3. Effect of MH particles on inflammation induced by PLGA in vivo. PLGA... 37
Figure 4. Residual PLGA or PLGA/MH composites were characterized at 2 weeks... 39
Figure 5. Effect of MH particles on expression of inflammation-related adhesion... 41
Figure 6. Effect of MH particles on expression of inflammation-related factors.... 42
Figure 7. Effect of MH particles on macrophage activation. (A) THP-1 derived... 45
Figure 8. Effect of MH particles on intracellular uptake of monomers of PLGA.... 48
Figure 9. Effect of MH particles on mitochondrial dysfunction and ROS generation... 51
Figure 10. Effect of ROS scavenger and respiratory inhibitor on PLGA-induced... 52
Figure 11. Effect of MH particles on PLGA degradation products-induced MAPK... 54
Figure 12. Effect of magnesium ion on endothelial function. HCAECs were... 57
CHAPTER 2 11
Figure 1. Schematic illustration of an integrated bioactive scaffold fabrication for... 87
Figure 2. Characterization of EVs. (a) Evaluation of morphology by transmission... 88
Figure 3. Effect of PDRN and TI-EV on wound repair and regeneration in vitro.... 92
Figure 4. Effect of PDRN and TI-EVs on EMT. (A) Representative... 94
Figure 5. Effect of PDRN and TI-EVs on macrophage polarization.... 95
Figure 6. Characterization of composite scaffolds. (A) Representative SEM images... 97
Figure 7. Biocompatibility of composite scaffolds in vitro. (a) Evaluation of... 99
Figure 8. Biocompatibility of composite scaffolds in vivo. (A) Representative... 100
Figure 9. In vivo evaluation of kidney tissue regeneration capacity of scaffolds. 102
Figure 10. In vivo evaluation of inflammatory responses. (A) Representative... 104
Figure 11. In vivo evaluation of restoration of kidney function. (a)... 107