Title Page
Contents
ABSTRACT 9
Chapter I. PEP-1-GSTA2 inhibits hippocampal neuronal cell death in HT-22 cells and in an animal model of ischemia 13
INTRODUCTION 14
MATERIAL AND METHODS 16
Materials and cell culture 16
Purification and transduction of PEP-1-GSTA2 proteins into HT-22 cells 16
Western blot analysis 17
Confocal fluorescence microscopy analysis 18
MTT assay 18
Measurement of reactive oxygen species (ROS) levels 19
TUNEL staining 19
Experimental animals 19
Immunohistochemical analysis 20
Statistical analysis 21
RESULTS 22
Purification and transduction of PEP-1-GSTA2 protein into HT-22 cells 22
Effects of transduced PEP-1-GSTA2 protein against H₂O₂-induced HT-22 cells 27
Effects of PEP-1-GSTA2 proteins on H₂O₂-induced signaling pathways in HT-22 cells 32
Protective effect of PEP-1-GSTA2 protein on ischemic injury 37
DISCUSSION 42
REFERENCES 46
Chapter II. Protective effects of PEP-1-GSTA2 protein against MPP+- and MPTP-induced dopaminergic neuronal cell death(이미지참조) 54
INTRODUCTION 55
MATERIAL AND METHODS 57
Materials and cell culture 57
Purification and transduction of PEP-1-GSTA2 proteins into SH-SY5Y cells 57
Western blot analysis 58
Confocal fluorescence microscopy analysis 59
MTT assay 59
Measurement of reactive oxygen species (ROS) levels 59
TUNEL staining 60
Animal study and immunohistochemistry 60
Statistical analysis 61
RESULTS 63
Purification and transduction of PEP-1-GSTA2 protein into SH-SY5Y cells 63
Effects of transduced PEP-1-GSTA2 protein against MPP+-induced SH-SY5Y cells(이미지참조) 68
Effects of PEP-1-GSTA2 proteins on MPP+-induced signaling pathways in SH-SY5Y cells(이미지참조) 73
Effects of PEP-1-GSTA2 protein against MPTP-induced in a PD animal model 79
DISCUSSION 82
REFERENCES 85
Chapter III. PEP-1-GLRX1 protects against MPP+- and MPTP-induced dopaminergic neuronal cell death by inhibiting MAPK activation and apoptosis(이미지참조) 94
INTRODUCTION 95
MATERIAL AND METHODS 97
Materials and cell culture 97
Expression and purification of PEP-1-GLRX1 proteins 97
Transduction of PEP-1-GLRX1 protein into SH-SY5Y cells 98
Western blot analysis 98
Fluorescence confocal microscopy analysis 99
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay 100
Measurement of intracellular reactive oxygen species (ROS) levels 100
Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay 100
Measurement of apoptotic protein expression 101
PD animal model 101
Immunohistochemistry 102
Statistical analysis 103
RESULTS 104
Construction and purification of PEP-1-GLRX1 protein 104
Transduction of PEP-1-GLRX1 protein into SH-SY5Ycells 106
Effects of transduced PEP-1-GLRX1 protein against MPP+-induced SH-SY5Y cells(이미지참조) 109
Effects of PEP-1-GLRX1 proteins on MPP+-induced signaling pathways in SH-SY5Y cells(이미지참조) 114
Effects of PEP-1-GLRX1 protein against MPTP-induced PD animal model 119
DISCUSSION 122
REFERENCES 126
국문초록 134
PUBLICATIONS 138
I-1. Purification of PEP-1-GSTA2 protein. 23
I-2. Transduction of PEP-1-GSTA2 proteins into HT-22 cells. 26
I-3. Effects of transduced PEP-1-GSTA2 protein against H₂O₂-induced cell viability. 29
I-4. Effects of PEP-1-GSTA2 protein against H₂O₂-induced ROS production and DNA... 31
I-5. Effects of PEP-1-GSTA2 protein on H₂O₂-induced MAPKs activation in HT-22 cells. 35
I-6. Effects of PEP-1-GSTA2 protein against H₂O₂-induced Bcl-2, Bax, Caspase 3 and... 36
I-7. Transduction of PEP-1-GSTA2 protein into gerbil brain and its protective effects. 39
I-8. Neuroprotective effects of PEP-1-GSTA2 protein against ischemic damage. 41
II-1. Purification of PEP-1-GSTA2 protein. 64
II-2. Transduction of PEP-1-GSTA2 proteins into SH-SY5Y cells. 67
II-3. Effects of transduced PEP-1-GSTA2 protein against MPP+-induced cell viability.(이미지참조) 70
II-4. Effects of PEP-1-GSTA2 protein against MPP+-induced ROS production and DNA...(이미지참조) 72
II-5. Effects of PEP-1-GSTA2 protein on MPP+-induced MAPKs activation in SH-SY5Y...(이미지참조) 75
II-6. Effect of PEP-1-GSTA2 protein against MPP+-induced Bcl-2, Bax, Caspase 3 and...(이미지참조) 78
II-7. PEP-1-GSTA2 protein transduction into animal brain. Transduction of PEP-1-GSTA2... 80
II-8. Transduced PEP-1-GSTA2 protein inhibits dopaminergic neuronal cell death in a PD... 81
III-1. Construction and purification of PEP-1-GLRX1 proteins. 105
III-2. Transduction of PEP-1-GLRX1 proteins into SH-SY5Y cells. 108
III-3. Effects of transduced PEP-1-GLRX1 protein against MPP+-induced cell viability.(이미지참조) 111
III-4. Effects of PEP-1-GLRX1 protein against MPP+-induced ROS production and DNA...(이미지참조) 113
III-5. Effects of PEP-1-GLRX1 protein on MPP+-induced MAPKs and NF-κB activation in...(이미지참조) 115
III-6. Effects of PEP-1-GLRX1 protein on MPP+-induced ASK1 and Akt activation in SH-...(이미지참조) 116
III-7. Effects of PEP-1-GLRX1 protein on MPP+-induced apoptotic protein expression in...(이미지참조) 118
III-8. PEP-1-GLRX1 protein transduction into animal brain. 120
III-9. Transduced PEP-1-GLRX1 protein inhibits dopaminergic neuronal cell death in PD... 121