本文選題：高濃度氧 + 新生小鼠��； 參考：《重慶醫(yī)科大學》2014年碩士論文

【摘要】：目的：探討小膠質(zhì)細胞在新生小鼠高氧暴露腦損傷中的功能變化及其作用。方法：清潔級健康野生型新生3d小鼠C57BL/10J（PND3）48只，隨機分為空氣組（A組）、高氧組(O組)；90%高濃度氧暴露48h，PND5斷頭取腦：（1）蛋白質(zhì)印跡技術(shù)（western-blot，WB）檢測capsase-3蛋白表達。（2）免疫熒光雙標檢測Caspase-3凋亡蛋白表達情況及分析NG2+標記的少突膠質(zhì)祖細胞凋亡；（3）免疫組化染色觀測腦室周圍白質(zhì)小膠質(zhì)細胞形態(tài)變化及Toll樣4受體（Toll-like receptor4,TLR4）的表達；（4） Real-time Quantitative PCR（Q-PCR）檢測TLR4和TNF-α(Tumor necrosis factor α)mRNA表達變化； 新生3d清潔級健康野生型小鼠C57BL/10J（PND3）24只，采用小膠質(zhì)細胞活化抑制劑米諾環(huán)素（Minocycline），腹腔注射（45mg/kg）30min后，90%高濃度氧暴露48h，，分為米諾環(huán)素+空氣組（AM組）、米諾環(huán)素+高氧組（OM組）,PND5斷頭取腦：（1）Q-PCR檢測TLR4和TNF-αmRNA表達變化；（2）免疫熒光雙標檢測Caspase-3凋亡蛋白表達情況及分析NG2+標記的少突膠質(zhì)前體細胞凋亡； 結(jié)果：免疫熒光顯示Caspase-3凋亡蛋白在O組的表達增高，O組NG2+標記的少突膠質(zhì)前體細胞出現(xiàn)細胞凋亡；O組免疫組化染色觀測腦室周圍白質(zhì)活化的小膠質(zhì)細胞數(shù)量及TLR4表達明顯增高；同時Q-PCR檢測結(jié)果中，O組中TLR4和TNF-α的mRNA表達明顯高于A組（P0.05）； OM組TLR4和TNF-α的mRNA表達低于相對應(yīng)的O組（P0.05），但高于A組（P0.05）；OM組腦組織Caspase-3明顯低于單純O組；但高于AM組和A組。各處理組間NG2表達無明顯差異，O組與OM組可見Caspase-3與NG2+共表達。 結(jié)論：新生鼠高濃度氧暴露誘導(dǎo)小膠質(zhì)細胞活化，TLR4受體表達增加，并最終通過氧化應(yīng)激與炎癥反應(yīng)（TNF-α表達增加）導(dǎo)致未成熟腦膠質(zhì)細胞凋亡，且凋亡細胞包括少突膠質(zhì)前體細胞。小膠質(zhì)細胞參與了新生鼠高濃度氧暴露導(dǎo)致的腦損傷。
[Abstract]:Aim: to investigate the functional changes and effects of microglia on brain injury in neonatal mice exposed to hyperoxia. Methods: C57BL/10J(PND3)48 mice of clean grade healthy wild-type newborn mice were used. The rats were randomly divided into two groups: air group A and hyperoxia group (90% high concentration oxygen exposure 48h). Western blot technique was used to detect the expression of capsase-3 protein. 2) Immunofluorescence double labeling was used to detect the expression of Caspase-3 apoptotic protein and to analyze the oligospermia of NG2 labeled by PND5. The Western blot technique was used to detect the expression of Caspase-3 protein. The morphological changes of microglia and the expression of Toll-like receptor 4 (TLR4) in the white matter around the ventricle were observed by immunohistochemical staining. The expression of TLR4 and TNF- 偽 Tumor necrosis factor were detected by Real-time Quantitative PCR Q-PCR4). Three days after birth, C57BL/10J(PND3)24 of clean grade healthy wild-type mice was found. Minocycline, an inhibitor of microglial activation, was used to detect the changes of TLR4 and TNF- 偽 mRNA expression in TLR4 and TNF- 偽 mRNA after intraperitoneal injection of 45 mg 路kg ~ (-1) 路min ~ (-1) min for 48 h after exposure to 90% high concentration oxygen, which was divided into two groups: minocycline air group (AM group), minocycline hyperoxia group (OM group) and minocycline group (OM group) were treated with minocycline 1 Q-PCR to detect the changes of TLR4 and TNF- 偽 mRNA expression. The expression of Caspase-3 apoptotic protein and the apoptosis of oligodendrocyte precursor cells labeled with NG2 were detected by immunofluorescence double labeling. Results: immunofluorescence showed that the expression of Caspase-3 apoptosis protein was increased in group O. The number of activated microglia and the expression of TLR4 in the white matter around the ventricle were observed by immunohistochemical staining in oligodendrocyte precursor cells labeled with NG2 in O group. At the same time, the mRNA expression of TLR4 and TNF- 偽 in Q-PCR group was significantly higher than that in group A (P 0.05). The mRNA expression of TLR4 and TNF- 偽 in OM group was lower than that in O group, but higher than that in group A (P 0.05) group, but higher than that in group A group (P 0.05) group, but higher than that in AM group and A group. There was no significant difference in the expression of NG2 between O and OM groups. Caspase-3 and NG2 were coexpressed in O group and OM group. Conclusion: the expression of TLR4 receptor in microglia activated by high oxygen exposure in neonatal rats is increased, and the expression of TNF- 偽 is increased through oxidative stress and inflammatory response), which leads to the apoptosis of immature brain glial cells. The apoptotic cells include oligodendrocytes. Microglia are involved in brain damage induced by high oxygen exposure in newborn rats.
【學位授予單位】：重慶醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R722.1

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本文編號：1920106