Title Page
Contents
PART Ⅰ. CO-induced TTP attenuates combined CTLA-4 and PD-1 blockade immunotherapy toxicity and promotes anti-tumor immunity 12
Ⅰ-Ⅰ. ABSTRACT 13
Ⅰ-Ⅱ. INTRODUCTION 14
Ⅰ-Ⅲ. MATERIALS AND METHODS 16
1. Reagents and Chemicals 16
2. Cell culture 16
3. Mice 16
4. DSS-induced colitis models and tumor models 17
5. Treatments 17
6. Magnetic activated cell sorting (MACS) 17
7. Flow cytometry analysis 18
8. RNA Isolation and Reverse Transcription-Polymerase Chain Reaction 18
9. Western blot 19
10. Statistical Analysis 20
Ⅰ-Ⅳ. RESULTS 21
1. irAEs, exacerbated by ICB, is attenuated by CO 21
2. TTP is required for the protective effects of CO on ICB-induced irAEs 21
3. CO-induced TTP augments ICB efficacy 22
4. CO dampens the immunosuppressive M2-TAM population in TME 23
5. TTP reprogrammed M2 macrophages to an M1-like population 24
Ⅰ-Ⅵ. DISCUSSION 38
Ⅰ-Ⅶ. CONCLUSION 41
Ⅰ-Ⅷ. REFERENCES 42
Ⅰ-Ⅸ. 국문 요약 49
PART Ⅱ. Activation of ROS-PERK-TFEB by filbertone ameliorates neurodegenerative disease via enhancing the autophagy-lysosomal pathway 50
Ⅱ-Ⅰ. ABSTRACT 51
Abbreviations 52
Ⅱ-Ⅱ. INTRODUCTION 53
Ⅱ-Ⅲ. MATERIALS AND METHODS 55
1. Reagents and Chemicals 55
2. Cell culture 55
3. Mice 55
4. Behavioral tests 56
5. Western blot 57
6. Measurement of mitochondrial ROS 58
7. Real-time quantitative RT-PCR 58
8. Transfection 59
9. Fluorescence cell imaging 59
10. Immunofluorescence 59
11. Subcellular fractionation 60
12. Statistical analysis 60
Ⅱ-Ⅳ. RESULTS 61
1. Filbertone treatment activates the PERK-eIF2α-ATF4 pathway in SH-SY5Y human neuroblastoma cells. 61
2. PERK activation by filbertone requires mitochondrial ROS 62
3. Filbertone promotes the autophagy-lysosomal pathway (ALP) in a PERK dependent manner 62
4. PERK is required for TFEB nuclear translocation by filbertone 63
5. Filbertone-induced PERK activation reduces α-syn accumulation by enhancing the TFEB-ALP axis 64
6. Filbertone exerts a neuroprotective effect in the HFD and MPTP murine models of Parkinson's disease 65
Ⅱ-Ⅵ. DISCUSSION 78
Ⅱ-Ⅶ. CONCLUSION 80
Ⅱ-Ⅷ. REFERENCES 81
Ⅱ-Ⅸ. 국문 요약 86
PART Ⅲ. Filbertone attenuates MPP⁺/MPTP-induced ferroptosis in neurological diseases via Nrf2-GPX4 activation 87
Ⅲ-Ⅰ. ABSTRACT 88
Ⅲ-Ⅱ. INTRODUCTION 89
Ⅲ-Ⅲ. MATERIALS AND METHODS 91
1. Reagents 91
2. Cell lines 91
3. Animals 91
4. Western blot 92
5. Measurement of lipid peroxidation 92
6. Real-time quantitative RT-PCR 93
7. Subcellular fractionation 93
8. WST-8 assay 94
9. Generation of knockdown cells 94
10. Statistical analysis 94
Ⅲ-Ⅳ. RESULTS 95
1. Filbertone attenuates MPTP-induced ferroptosis in mouse PD model 95
2. Filbertone inhibits RSL3-induced ferroptosis in SH-SY5Y human neuroblastoma cells 96
3. Filbertone inhibits MPP⁺ -induced ferroptosis in SH-SY5Y cells 96
4. Filbertone protects against ferroptosis by increasing SLC7A11-GPX4 axis 97
5. PERK-Nrf2 activation is required to anti-ferroptosis effect of filbertone 97
Ⅲ-Ⅵ. DISCUSSIONS 107
Ⅲ-Ⅶ. CONCLUSION 109
Ⅲ-Ⅷ. REFERENCES 110
Ⅲ-Ⅸ. 국문 요약 117
PART Ⅰ. CO-induced TTP attenuates combined CTLA-4 and PD-1 blockade immunotherapy toxicity and promotes anti-tumor immunity 6
Figure 1. Protective effects of CO inhalation on irAEs. 26
Figure 2. Effects of TTP on CO-induced protection of irAEs. 29
Figure 3. TTP is required for CO-induced antitumor effects. 31
Figure 4. CO decreases M2-TAM in tumor tissues. 32
Figure 5. TTP reduces macrophage M2 polarization. 34
Figure 6. TTP induces macrophage M1 polarization. 36
Schematic figure 1. CO-induced TTP protects against ICB-induced irAEs. 30
Schematic figure 2. CO-induced TTP enhances immune response by reprogramming macrophages. 37
PART Ⅱ. Activation of ROS-PERK-TFEB by filbertone ameliorates neurodegenerative disease via enhancing the autophagy-lysosomal pathway 9
Fig. 1. Filbertone treatment activates the PERK-eIF2α-ATF4 pathway in SH-SY5Y human neuroblastoma cells. 67
Fig. 2. PERK activation by filbertone requires mitochondrial ROS. 68
Fig. 3. Filbertone promotes the autophagy-lysosomal pathway (ALP) in a PERK dependent manner. 71
Fig. 4. PERK is required for TFEB translocation by filbertone 72
Fig. 5. Filbertone-induced PERK activation reduces α-syn accumulation by enhancing the TFEB-ALP axis. 73
Fig. 6. Filbertone exerts a neuroprotective effect in the HFD and MPTP murine models of Parkinson's disease 75
PART Ⅲ. Filbertone attenuates MPP⁺/MPTP-induced ferroptosis in neurological diseases via Nrf2-GPX4 activation 10
Fig. 1. Filbertone prevents MPTP-induced PD and inhibit ferroptosis in mouse models. 99
Fig. 2. Filbertone inhibits RSL3-induced ferroptosis in SH-SY5Y human neuroblastoma cells. 100
Fig. 3. Filbertone inhibits MPP⁺-induced ferroptosis in SH-SY5Y cells. 101
Fig. 4. Filbertone increases GPX4 and SLC7A11 levels. 102
Fig. 5. Filbertone activates PERK-Nrf2 pathway. 103
Fig. 6. Ablation of Nrf2 diminishes anti-ferroptosis effect of filbertone. 105