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Graduate School of Ajou University
Department of Neuroscience and Technology Course
3. Theses(Master)
P2 Receptor Modulation of IL-10 Expression and Its Effects on the Production of Reactive Oxygen species in Brain Ischemia
| DC Field | Value | Language |
|---|---|---|
| dc.contributor.advisor | Lee, Yong Beom | - |
| dc.contributor.author | 한우정 | - |
| dc.date.accessioned | 2019-10-21T06:46:04Z | - |
| dc.date.available | 2019-10-21T06:46:04Z | - |
| dc.date.issued | 2005 | - |
| dc.identifier.other | 365 | - |
| dc.identifier.uri | https://dspace.ajou.ac.kr/handle/2018.oak/16390 | - |
| dc.description | 학위논문(석사)--아주대학교 대학원 :신경과학협동과정,2005 | - |
| dc.description.abstract | 중추신경계에서 ATP는 신경전달물질로 알려져 왔다. 또한 최근 연구에 따르면 ATP는 P2 퓨린 수용체의 의해서 마이크로글리아에서 iNOS와 TNF-α의 발현을 조절한다고 제기되었다. 뿐만 아니라 우리는 항 염증 조절인자로 알려진 IL-10이 활성화된 마이크로글리아에서 발현되는 P2 퓨린 수용체를 통해 생성된다는 것을 확인하였다. 이에 본 연구에서는 P2 퓨린 수용체를 통해 생성되는 IL-10이 활성산소를 억제시켜 뇌졸중을 막아주는 작용에 대해서 연구하였다. 뇌졸중의 in vitro 모델인 신경세포-아스트로사이트-마이크로글리아 세포배양을 oxygen-glucose deprivation (OGD) 상태에서, 마이크로글리아 세포에서 IL-10, P2 퓨린 수용체, 활산소종의 발현을 확인을 하였다. 더 나아가 IL-10와 활성산소가 어떤 기작에 의해서 생성이 되는지를 확인하기 위해서 P2X7 수용체에 특이 antagonist인 o-ATP 및 P2Y1 의 수용체 측이 antagonist인 MRS2179를 처리하여 확인한 결과, IL-10의 생성에 P2X7, P2Y1 수용체가 모두 관여하는 반면 활산소종의 생성에는 P2X7 수용체만이 관여하는 것을 확인하였다. IL-10이 활산소종의 생성을 억제하는 것을 확인하기 위해서 anti-IL-10을 처리하여 확인한 결과, anti-IL-10 처리시 활산소종의 생성이 증가되는 것을 확인하였다. 더욱이, P2Y1 수용체의 특이 antagonist인 MRS2179에 의해 IL-10의 분비가 억제되어 P2X7에서 분비되는 활산소종을 억제하지 못하여 신경세포사멸이 증가되는 것을 확인하였다. 이로써 본 연구를 통하여 뇌졸중 상태에서 활성화된 마이크로글리아에서 발현된 P2X7 및 P2Y1 수용체에 의해 생성된 IL-10이 활성산소의 생성을 억제함으로써 뇌졸중에서 일어나는 신경세포사멸을 줄일 수 있다는 것을 규명하였다. | - |
| dc.description.tableofcontents | TABLE OF CONTENTS APPROVAL PAGE = ⅰ ACKNOWLEDGEMENT = ⅱ ABSTRACT = ⅲ TABLE OF CONTENTS = ⅴ LIST OF FIGURES = ⅶ LIST OF ABBREVIATIONS = ⅷ INTRODUCTION = 1 MATERIALS AND METHODS = 4 1. Cortical neuron culture. = 4 2. Microglia and Astrocyte culture = 4 3. Neuron-microlgia-astrocyte co-culture = 5 4. Exposure of rat cortical culture to OGD = 5 5. Assessment of neuronal injury = 6 6. ATP efflux measurement = 6 7. Reverse transcription ― PCR (RT-PCR) analysis = 7 8. Determination of IL-10 release = 8 9. Immunocytochemistry = 8 10. Terminal deoxynucleotidyl transferase-mediated fluorescein-dUTP nick-end labeling (TUNEL) assay = 9 11. Detection of ROS = 10 12. Focal cerebral ischemia = 11 13. Tissue preparation = 11 14. Statistical analysis = 12 RESULTS = 13 1. Neuronal cell death and ATP release by OGD stimulation = 13 2. IL-10 expression and ROS production in OGD-stimulated micrglia = 14 3. P2 purinergic receptors expression on microglia in OGD-stimulated microglia = 14 4. IL-10 and ROS expression via P2Y or P2X7 receptors = 15 5. Effects of extracellular ATP on IL-10 and P2 purinergic receptor expression in OGD-stimulated microglia = 16 6. IL-10 protects cortical neurons against OGD-induced neuronal cell death = 17 7. Neuronal cell death after ischemia = 17 8. Expression of the IL-10 and P2 purinergic receptors on microglia after ischemia = 18 DISCUSSION = 19 REFERENCES = 24 국문요약 = 32 | - |
| dc.language.iso | eng | - |
| dc.publisher | The Graduate School, Ajou University | - |
| dc.rights | 아주대학교 논문은 저작권에 의해 보호받습니다. | - |
| dc.title | P2 Receptor Modulation of IL-10 Expression and Its Effects on the Production of Reactive Oxygen species in Brain Ischemia | - |
| dc.title.alternative | 뇌졸중에서 P2 퓨린 수용체에 의한 IL-10 발현조절 및 활산소종 생산에 대한 조절기전 연구 | - |
| dc.type | Thesis | - |
| dc.contributor.affiliation | 아주대학교 일반대학원 | - |
| dc.contributor.alternativeName | 한우정 | - |
| dc.contributor.department | 일반대학원 신경과학기술과정 | - |
| dc.date.awarded | 2005. 2 | - |
| dc.description.degree | Master | - |
| dc.identifier.localId | 564516 | - |
| dc.identifier.url | http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000000365 | - |
| dc.description.alternativeAbstract | Recently, exogenous ATP has been introducted as a potential mediator of inducible nitric oxide synthase (iNOS) expression and TNF-α production in rat microglia via P2 purinergic receptors. In addition, we showed that ATP evokes the expression of IL-10, an important anti-inflammatory modulator in the CNS, in rat microglia via P2 purinergic receptors. Therefore, we hypothesized that extracellular ATP, mediating microglial IL-10 expression, could protect ischemic brain injury by a negative feedback inhibition to limit the production of proinflammatory cytokines and reactive oxygen species (ROS). In the present study, we investigated that the expression patterns of IL-10 and P2 receptors were similarly increased in oxygen?glucose deprivation (OGD)-stimulated microglia, suggesting that both P2X7 and P2Y1 receptors are involved in extracellular ATP-evoked IL-10 release from OGD-stimulated microglia. H₂O₂ assay showed that ROS release from OGD-stimulated microglia was blocked by P2X7 specific antagonist such as o-ATP, whereas it was increased by P2Y1 specific antagonist such as MRS2179. Moreover, MRS2179 pre-treatment augmented OGD-stimulated neuronal cell death by blocking the IL-10 release from microglia. These results suggest that P2X7 receptor is the primary receptor stimulated by ATP to generate ROS in brain ischemia, and ischemic brain injury is regulated by IL-10 released from microglia through P2X7 and /or P2Y1 receptors. | - |
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