지치 (Lithospermum erythrorhizon) 뿌리의 망막세포 보호효과에 대한 연구
- 주제(키워드) Glaucoma , Lithopermum erythrorhizon , R28 cell , Retinal ganglion cell , Optic nerve crush , retinal degeneration
- 발행기관 강릉원주대학교
- 지도교수 전중균, 정상훈
- 발행년도 2018
- 학위수여년월 2018. 2
- 학위명 석사
- 학과 및 전공 일반대학원 KIST강릉분원학.연협동과정
- 세부전공 해양생물공학
- 원문페이지 68
- 본문언어 한국어
- 저작권 강릉원주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
This study aimed to investigate the protective effects of Lithospermum erythrorhizon on the retinal degeneration in vitro and in vivo. Oxidative stress was induced in retinal precursor cells (R28) by L-Buthionine-(S,R)-sulfoximine (BSO) plus glutamate. Among the five systematic fractions tested, dichloromethane fraction of L. erythrorhizon (MCLE) showed a significantly higher cell viability compared to the negative control. Treatment of MCLE reduced the intracellular reactive oxygen species (ROS) and restored glutathione levels in R28 cells. The apoptotic proteins (Bax, apoptosis-inducing factor (AIF), cleaved poly (ADP-ribose) polymerase (PARP), and cleaved caspase-3) were down-regulation by treatment of MCLE. In an animal model, optic nerve crush (ONC) was performed in C57BL/6 mice and MCLE was orally administered throughout the experimental period. Oral administration of MCLE was markedly protective retinal ganglion cells (RGCs) and inhibited loss of the thickness of retinal layers. These results demonstrated that MCLE has protective effects against retinal degeneration both in vitro and in vivo. Therefore, MCLE could be a beneficial agent for treatment of retinal degeneration, such as glaucoma.
more목차
Abstract··························································································1
List of Figures················································································3
List of Table······················································································6
1. 서론····························································································7
1.1. 연구배경···············································································7
1.2. 녹내장·················································································11
1.3. 산화 스트레스······································································12
1.4 지치 (Lithospermum erythrorhizon)·············································14
2. 재료 및 방법··············································································17
2.1. 연구재료··············································································17
2.2. 식물····················································································17
2.3. 추출 및 분리········································································18
2.4. 망막전구세포의 배양····························································25
2.5. 세포생존율 측정··································································· 25
2.6. Propidium iodide (PI)/Hoechst 333422 이중 염색법························26
2.7. 세포 내 ROS 측정································································27
2.8. 세포 내 글루타티온 (Glutathione, GSH)함량 측정·······················28
2.9. 세포 단백질 추출 및 웨스턴블롯·············································38
2.10. 실험동물············································································30
2.11. 시신경 압박 동물 모델·························································30
2.12. 광 간섭성 단층촬영·····························································31
2.13. 망막 안저 이미지 촬영·························································32
2.14. 망막신경절세포 표지와 망막 플랫 마운트·······························32
2.15. 통계 분석···········································································33
3. 결과 및 고찰··············································································34
3.1. LC-MC/MC 분석에 의한 화합물 동정 ·····································34
3.1. 산화스트레스로 유도되는 망막 세포 사멸 억제 효과················36
3.2. 산화스트레스로 유도되는 활성산소종 생성 억제 효과················40
3.3. 산화스트레스로 유도되는 글루타티온 감소 억제 효과················41
3.4. 망막 세포에서 세포사멸 관련 단백질 활성 억제 효과················45
3.5. 시신경압박으로 인한 망막변성 보호효과··································48
3.6. 망막신경절세포와 망막층 보호효과··········································51
4. 결론··························································································55
5. 참고문헌····················································································56
