Protective effects of Tat-ATOX1 on MPP+-induced cellular toxicity and MPTP-induced Parkinson's disease
MPP+로 유도한 세포 독성과 MPTP로 유도한 파킨슨 병에서 Tat-ATOX1의 보호 효과
- 주제(키워드) 도움말 Tat-ATOX1 , Parkinson’s disease , Dopaminergic neuron , MAPK , Protein therapy
- 발행기관 강릉원주대학교 일반대학원
- 지도교수 도움말 김대원, 박순호
- 심사위원 유기연, 김대원, 박순호
- 사용가능일 20190222
- 발행년도 2018
- 학위수여년월 2019. 2
- 학위명 석사
- 학과 및 전공 도움말 일반대학원 치의학과
- 실제URI http://www.dcollection.net/handler/kangnung/000000010326
- UCI I804:42001-000000010326
- 본문언어 영어
초록/요약 도움말
신경퇴행성 질환인 파킨슨 병(Parkinson’s disease, PD)은 뇌 흑질(substantia nigra, SN)의 도파민 뉴런 소실로 일어난다. Antioxidant 1(ATOX 1) 단백질은 항산화물질이자 샤페론(chaperone)으로써 다양한 질병에서 중요한 역할을 한다. 본 연구에서는 1-methyl-4-phenylpyridinium ion (MPP+)으로 유도된 SH-SY5Y 세포 사멸과 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)으로 유도된 PD 동물 모델에서 Tat-ATOX 1의 보호 효과를 알아보았다. Tat-ATOX 1은 MPP+에 노출 된 SH-SY5Y 세포의 사멸과 독성을 현저히 억제하였다. 또한 Tat-ATOX 1은 Akt와 mitogen activated protein kinases (MAPKs)의 활성, cleaved caspase-3와 Bax의 발현 수준을 유의미하게 억제하였다. Tat-ATOX1은 MPTP로 유도된 PD 동물 모델의 뇌 흑질로 형질도입 되어 도파민 뉴런 세포 소실에 대해 보호한다. 결론적으로 Tat-ATOX1이 MAPKs와 세포 사멸 신호 경로 억제를 통하여 도파민 뉴런 세포 사멸을 억제한다는 것을 실험 결과로 알 수 있다. 따라서 Tat-ATOX1을 PD의 잠재적 치료제로 제시한다.
more초록/요약 도움말
Parkinson’s disease (PD), a progressive neurodegenerative diseases, is characterized by the loss of dopaminergic neurons in the substantia nigra (SN) par compacta and symptomized by bradykinesia, rigidity, and postural instability. Although the pathogene- sis of PD is not clearly understood yet, PD is associated with several factors including oxidative stress, inflammation, and mitochondrial dysfunction in the pathogenesis proces-ses [1, 2]. It has been well described that 1-Methyl-4-phenylpyidinium ion (MPP+), transformed from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), can cross blood brain barrier (BBB) leading to PD-like symptoms. Therefore, MPP+ and MPTP have been used to induce PD-like symptoms in both in vitro and in vivo models of PD [2-4]. Antioxidant 1 (ATOX1), as a copper chaperone protein, is highly expressed in the brain and plays important roles as an antioxidant against oxidative stress induced by superoxide and hydrogen peroxide [5-7]. In addition, several studies have reported that ATOX1, as an antioxidant and copper regulator protein, is highly involved promoting cell survival in cancer and neuronal diseases [8-13]. In a previous study, we demonstrated that cell permeable Tat-ATOX1 protein markedly inhibited HT-22 and RINm5F cell death caused by oxidative stress [14, 15]. Tat peptides are known as common protein transduction domains (PTDs) and can deliver macromolecules including proteins into cells and tissues [16-18]. Therefore, PTD has been widely used to deliver therapeutic agents into cells or tissues in protein therapy experiments and many studies have reported that PTD fusion proteins transduce into cells and inhibited cell death [19-25]. In this study, we examined the roles of Tat-ATOX1 protein on MPP+-induced SH-SY5Y cell death and in an animal model of PD. Tat-ATOX1 protein transduced into dopaminergic neuronal cells in vitro and in vivo, where it markedly inhibited dopaminergic neuronal cell death induced by MPP+-or MPTP.
more목차 도움말
Ⅰ. Introduction 1
Ⅱ. Materials and Methods 3
2.1. Cell culture and Materials 3
2.2. Transduction of Tat-ATOX1 protein into SH-SY5Y cells 3
2.3. Western blot analysis 4
2.4. Fluorescence microscopy analysis 4
2.5. Cell viability assay 4
2.6. Measurement of intracellular ROS and DNA damage levels 5
2.7. Experimental PD animal model study 5
2.8. Immunohistochemistry and cell counting 6
2.9. Statistical analysis 8
Ⅲ. Results 9
3.1. Transduction of Tat-ATOX1 protein into SH-SY5Y cells 9
3.2. Effects of Tat-ATOX1 protein against MPP+-induced SH-SY5Y cell death 13
3.3. Effects of Tat-ATOX1 protein against MPP+-induced MAPKs and apoptosis signaling pathways 16
3.4. Effects of Tat-ATOX1 protein in MPTP-induced PD animal model 19
Ⅳ. Discussion 22
Ⅴ. Reference 25
Ⅲ. Korean Abstract 31
