뇌종양 세포에서 Ophiobolin A에 의해 유도되는 paraptosis 유사 세포사멸의 분자기전
Underlying mechanisms of Ophiobolin A-induced paraptosis-like cell death in glioma cells
- 주제(키워드) Ophiobolin A , Glioblastoma multiforme (GBM) , endoplasmic reticulum , ER vacuolation , paraptosis , thiol homeostasis
- 발행기관 아주대학교
- 지도교수 최경숙
- 발행년도 2017
- 학위수여년월 2017. 8
- 학위명 석사
- 학과 및 전공 일반대학원 의생명과학과
- 실제URI http://www.dcollection.net/handler/ajou/000000025945
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
Ophiobolin A (OP-A), a fungal sesterterpene from Bipolaris oryzae, has recently been shown to have anti-glioma activity. I found that OP-A induces paraptosis-like cell death accompanied by the endoplasmic reticulum (ER) dilation, accumulation of poly-ubiquitinated proteins in glioma cells and CHOP-mediated ER stress plays a critical role in this process. Inhibition of protein synthesis using cycloheximide (CHX) almost completely blocked the vacuolation and glioma cell death, suggesting a requirement of new protein synthesis for this death accompanied by vacuolation. Interestingly, thiol-based antioxidants, including N-acetylcysteine (NAC) and reduced glutathione (GSH) and N-(2-mercaptopropionyl)-glycine (NMPG), but not other non-thiol antioxidants, including Ascorbic acid and Tiron and MnTBAP (Mn(III)tetrakis (4-benzoic acid) porphyrin) very effectively blocked OP-A-induced vacuolation and cell death. In addition, notable generation of reactive oxygen species (ROS) by OP-A was not detected, suggesting that the anti-cancer effects of OP-A may be mediated though its ability to covalently modify free sulfhydryl groups on proteins causing protein misfolding, rather than through ROS generation. Collectively, our results indicate that ER stress due to disruption of thiol proteostasis may be responsible for the potent anti-glioma activity of OP-A.
more목차
I. INTRODUCTION 1
II. MATERIALS AND METHODS 4
1. Chemicals and antibodies 4
2. Cell culture 4
3. Measurement of cellular viability (Live/Dead assay) 5
4. Examination of the stable cell lines expressing the fluorescence specifically in the endoplasmic reticulum 5
5. Western blotting 6
6. Immunocytochemistry (ICC) 6
7. Gene silencing of CHOP (Knockdown experiments using siRNAs and shRNAs) 7
8. Measurement of reactive oxygen species (ROS) production 7
9. Statistical analysis 8
III. RESULTS 9
1. Ophiobolin A induces paraptosis-like cell death in glioma cells via dilation of the ER 9
2. OP-A induces ER stress and CHOP plays a critical role in OP-A-induced paraptosis-like cell death 19
3. Thiol-antioxidants but not other ROS scavengers block OP-A-induced paraptosis-like cell death 28
4. Disruption of thiol homeostasis by OP-A critically contributes to OP-A-mediated anti-cancer effects in various cancer cell lines 36
IV. DISCUSSION 47
V. REFERENCES 56
국문 요약 63
