【摘要】：目的：研究自發(fā)性腦出血（spontaneous intracerebral hemorrhage, SICH）后紅細(xì)胞對(duì)腦含水量及水通道蛋白-4（aquaporin-4, AQP4）表達(dá)的影響；SICH后腦水腫形成過(guò)程中，AQP4的表達(dá)與腦含水量之間的關(guān)系。 方法：選用成年雄性SD大鼠130只，隨機(jī)分為正常組（NG）、假手術(shù)組（SO）、全血組（WB）、紅細(xì)胞組（RBC），其中SO、WB、RBC組按處死時(shí)間6h、1d、3d、5d再分為四個(gè)亞組，每個(gè)亞組10只，其中5只用于腦含水量、RT-PCR及Western Blotting的檢測(cè),另外5只用于HE染色及免疫組化檢測(cè)AQP4。使用立體定向技術(shù)向SD大鼠右側(cè)尾狀核位置注射自體尾動(dòng)脈血或自體紅細(xì)胞建立腦出血?jiǎng)游锬Ｐ�；采用干濕重法測(cè)腦含水量；應(yīng)用免疫組化和Western blotting法檢測(cè)AQP4蛋白的表達(dá)情況，RT-PCR法檢測(cè)AQP4mRNA的變化情況； 結(jié)果：腦含水量在RBC組6h、1d、3d及5d的組內(nèi)比較，RBC-3d組明顯高于6h、1d、5d組，各組間比較RBC-6h后腦含水量開始增多（P0.05），在出血后3天到達(dá)高峰（P0.01），以后逐漸減少，在出血后5天仍高于正常水平（P0.01）。WB組與RBC組在各時(shí)間段的腦含水量變化規(guī)律相似，但是均低于RBC組；免疫組化結(jié)果WB組及RBC組3d均可見(jiàn)AQP4表達(dá)于神經(jīng)膠質(zhì)細(xì)胞或血管周邊的神經(jīng)膠質(zhì)細(xì)胞足突，胞漿著色，呈棕黃色。Western blotting結(jié)果顯示RBC組內(nèi)比較，RBC-3d組的AQP4蛋白量明顯高于RBC-6h、1d、5d組，各組間比較RBC-6h后AQP4蛋白量開始增多（P0.05），在出血后3天到達(dá)高峰（P0.01），以后逐漸減少，在出血后5天仍高于正常水平（P0.01）；WB組與RBC組在各時(shí)間段的AQP4蛋白量變化規(guī)律相似，但是均低于RBC組。RT-PCR結(jié)果顯示RBC組6h、1d、3d及5d的組內(nèi)比較， RBC-3d組AQP4mRNA明顯高于6h、1d、5d組，RBC-6h后血腫周圍組織AQP4mRNA表達(dá)開始增高（P0.05），在出血后3天到達(dá)高峰（P0.01），以后逐漸下降，在出血后5天仍高于正常水平（P0.01），WB組AQP4mRNA表達(dá)水平與RBC組規(guī)律相似，但均低于RBC組；NG組、SO組的各時(shí)間段腦含水量、AQP4蛋白量及AQP4mRNA表達(dá)水平無(wú)明顯差異。 結(jié)論： 1. AQP4在WB組和RBC組第3天時(shí)陽(yáng)性表達(dá)最強(qiáng)，，提示紅細(xì)胞可刺激AQP4大量表達(dá)； 2. WB組和RBC組在第3天時(shí)腦含水量最高，提示紅細(xì)胞在SICH后腦水腫形成中起著重要作用，即紅細(xì)胞可能是SICH后第3天腦水腫形成的重要物質(zhì)基礎(chǔ)； 3.綜合AQP4及腦含水量的結(jié)果，AQP4的陽(yáng)性表達(dá)與腦含水量的變化相一致，即WB組和RBC兩組在第3天時(shí)腦含水量最高，AQP4的陽(yáng)性表達(dá)也最強(qiáng)，說(shuō)明AQP4的表達(dá)與腦含水量的變化趨勢(shì)一致，提示SICH后第3天所形成的腦水腫可能是通過(guò)AQP4的表達(dá)上調(diào)來(lái)實(shí)現(xiàn)的。
[Abstract]:Aim: to study the effect of erythrocyte on brain water content and aquaporin-4 (aquaporin-4, AQP4) expression after spontaneous intracerebral hemorrhage (spontaneous intracerebral hemorrhage, SICH) and the relationship between AQP4 expression and brain water content during the formation of cerebral edema after SICH. Methods: 130 adult male SD rats were randomly divided into normal (NG), sham-operation group, (SO), whole blood group, (WB), erythrocyte (RBC), group and SO,WB,RBC group, which were divided into four subgroups according to the time of death for 6 h, 1 d, 3 d and 5 d, respectively. Of the 10 rats in each subgroup, 5 were used for the detection of brain water content, RT-PCR and Western Blotting, and 5 were used for HE staining and immunohistochemical detection of AQP4.. Intracerebral hemorrhage model was established by injecting autologous caudate blood or autologous red blood cells into the right caudate nucleus of SD rats by stereotactic technique. Immunohistochemical and Western blotting methods were used to detect the expression of AQP4 protein, and RT-PCR method was used to detect the changes of AQP4mRNA. Results: the brain water content in the RBC group was significantly higher than that in the RBC-3d group (P 0.05). The brain water content in the RBC-3d group was significantly higher than that in the RBC-3d group at 6 h / 1 d and 5 d after RBC-6h (P0.05), and the brain water content in the RBC-3d group was significantly higher than that in the control group (P < 0.05). The cerebral water content in P0.01). WB group and RBC group was similar to that in RBC group, but lower than that in RBC group, and reached the peak 3 days after bleeding (P0.01), then decreased gradually, and remained higher than that of normal level 5 days after hemorrhage (P0.01). WB group and RBC group in each time period). The results of immunohistochemistry showed that AQP4 was expressed in the foot process of glial cells or peripheral glial cells in WB group and RBC group on the 3rd day, and the cytoplasm was stained. The results of brown. Western blotting showed that the AQP4 expression in RBC group was higher than that in RBC group. The amount of AQP4 protein in RBC-3d group was significantly higher than that in RBC-6h,1d,5d group. Compared with RBC-6h, the amount of AQP4 protein began to increase (P0.05), reached the peak at 3 days after bleeding (P0.01), and then decreased gradually. It was still higher than normal level 5 days after bleeding (P0.01). The changes of AQP4 protein content in WB group and RBC group were similar, but lower than those in RBC group. RT-PCR results showed that AQP4mRNA in RBC-3d group was significantly higher than that in 6 h / 1 day group and 5 d group, compared with 6 h / 1 day group and 5 d group, RT-PCR results showed that AQP4mRNA in RBC-3d group was significantly higher than that in 6 h / 1 day group. After RBC-6h, the expression of AQP4mRNA began to increase (P0.05), reached the peak 3 days after hemorrhage (P0.01), then gradually decreased, still higher than the normal level 5 days after bleeding (P0.01). The expression of AQP4mRNA in WB group was similar to that in RBC group, but lower than that in RBC group. There was no significant difference in brain water content, AQP4 protein content and AQP4mRNA expression in NG group and SO group. Conclusion: 1. The positive expression of AQP4 in WB group and RBC group was the strongest on the 3rd day, suggesting that erythrocytes could stimulate AQP4 expression in large amount; 2. WB and RBC groups had the highest brain water content on the third day, suggesting that red blood cells play an important role in the formation of brain edema after SICH, that is, red blood cells may be an important material basis for the formation of brain edema on the third day after SICH; 3. According to the results of AQP4 and brain water content, the positive expression of AQP4 was consistent with the change of brain water content, that is, WB and RBC groups had the highest brain water content and the strongest positive expression of AQP4 on the third day. The results suggest that the expression of AQP4 is consistent with the change of brain water content, suggesting that the brain edema formed on the 3rd day after SICH may be realized by the up-regulation of AQP4 expression.
【學(xué)位授予單位】：河南科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2014
【分類號(hào)】：R743.34

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