Astrocytes의 hemeoxygenase-1의 발현을 통한 microglia의 활성화조절
Astrocytes reduce microglial activation through the expression of hemeoxygenase-1
- 주제(키워드) astrocyte , adenosine , reactive oxygen species , hemeoxygenase-1
- 주제(KDC) 511
- 주제(DDC) 616
- 발행기관 아주대학교
- 지도교수 조은혜
- 발행년도 2007
- 학위수여년월 2007. 8
- 학위명 박사
- 학과 및 전공 일반대학원 신경과학기술과정
- 본문언어 영어
초록/요약
In damaged brain, inflammation occurs as a host defense mechanism. However, brain inflammation has a double-edged effect on brain damage. Inflammation protects the brain from infection, but aggravates the injury of surrounding tissue. Therefore, negative regulatory mechanisms of brain inflammation exist to prevent prolonged and extensive inflammation. The first part of this thesis showed that astrocytes, the most abundant cells in the brain modulate microglial activation by regulating the microglial levels of reactive oxygen species (ROS). Astrocyte culture-conditioned media (ACM) suppressed IFN-gamma- nor LPS-induced ROS production, leading to reduced iNOS expression and NO release in microglia. Treatment of microglia with ACM increased the expression level and activity of hemeoxygenase-1 (HO-1) through the activation of nuclear factor E2-related factor 2 (Nrf2) transcription factor. The active component(s) in ACM was/were heat-labile and smaller than 3 kD. Furthermore, phosphoinositide-3 kinase (PI3K) mediates ACM-induced HO-1 expression. Mimickers of HO-1 products such as bilirubin, ferrous iron and a carbon monoxide (CO)-releasing molecule also reduced IFN-gamma-induced iNOS expression and/or NO release. Damaged astrocytes and neurons also down-regulate microglial activation through HO-1 expression. Therefore, astrocytes and neuron appears to cooperate with microglia to prevent excessive inflammatory responses in the brain by regulating microglial ROS production. The second part of this thesis showed that adenosine could modulate microglial activation. In response to adenosine, Nrf2 was translocated from the cytosol to the nuclei, bound to ARE, and then HO-1 promoter activity increased. PI3K and its downstream, Akt, appeared to mediate adenosine-induced HO-1 expression since adenosine induced Akt phosphorylation and inhibitors of PI3K (LY294002 and wortmanin) reduced adenosine-induced HO-1 expression. These results suggest that adenosine could be an endogenous regulator of brain inflammation through the modulation of microglial ROS production. Taken together, these results suggest that astrocytes and neuron, and an endogenous factor, adenosine, could regualte microglia-mediated brain inflammation to minimize inflammation-induced brain damage.
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ACKNOWLEGEMENTS
ABSTRACT
LIST OF FIGURES
LIST OF TABLE
LIST OF ABBREVIATION
I.INTRODUCTION
A.Inflammation
B.Glia : microglia and astrocytes as main regulators of brain inflammation
1.Microglia
2.Astrocytes
C.Positive regulation of brain inflammation
1.Stimulators of brain inflammation
2.Proinflammatory mediators
D.Negative regulation of brain inflammation
1.Anti-inflammatory cytokines
2.Anti-oxidant enzymes
3.Astrocytes
4.Neuron
5.Adenosine
E.Signaling molecules of inflammation
F.Aims of this study
II.MATERIALS AND METHODS
1.Materials
2.Methods
2.1 Cell culture
2.2 Western blot analysis
2.3 Reverse Transcription and Polymerase Chain Reaction (RT-PCR)
2.4 Electrophoresis Mobility Shift Assay
2.5 Measurement of ROS
2.6 Immunocytochemistry
2.7 HO-1 activity assay
2.8 Measurement of NO
2.9 Transfection and luciferase assay
2.10 Preparation of conditioned media
2.11 Oxygen-glucose deprivation (OGD)
2.12 Fractionation of ACM and OGD
2.13 Statistical analysis
III.RESULTS
Part A. Astrocytes induce hemeoxygenase-1 expression in microglia
1.Astrocytes inhibit microglial inflammatory responses
2.HO-1 is important on anti-inflammatory roles of astrocytes 4
3.ACM increases HO-1 expression through activation of Nrf2/ARE signaling and PI3K/Akt pathway
4.The active component(s) in ACM is/are heat-labile and smaller than 3kD
Part B. Adenosine induces hemeoxygenase-1 via PI3K pathway in microglia
1.Adenosine reduces microglial inflammatory response
2.Adenosine activates nuclear factor E-related factor 2 (Nrf2)
3. PI3K is involeved in adenosine-induced HO-1 expression
4.Adenosine induces HO-1 expression in adenosine receptors independent manner
IV.DISCUSSION
V.SUMMARY AND CONCLUSION
VI.BIBLIORAPHY
국문초록
