發(fā)布時(shí)間：2018-01-30 09:40

  本文關(guān)鍵詞： 放射性腦病 星形膠質(zhì)細(xì)胞 血腦屏障 腫瘤壞死因子 糖皮質(zhì)激素 海馬　出處：《廣西醫(yī)科大學(xué)》2012年碩士論文　論文類(lèi)型：學(xué)位論文

【摘要】：目的放射治療是治療頭頸部等惡性腫瘤的有效手段之一,但是同時(shí)也帶來(lái)了以記憶力減退為主要癥狀的認(rèn)知功能障礙。海馬在人類(lèi)的情緒整合、學(xué)習(xí)、記憶等高級(jí)功能中占有重要地位,放射治療引起海馬損傷已經(jīng)成為日益關(guān)注的重要課題。放射治療引起的腦損傷還涉及到炎性反應(yīng),因此采取抗炎手段防治放射性腦病已經(jīng)成為研究重點(diǎn),而糖皮質(zhì)激素已經(jīng)廣泛應(yīng)用于臨床,但是糖皮質(zhì)激素如何防治放射性腦損傷的作用機(jī)制還未完全明了.本實(shí)驗(yàn)通過(guò)建立低劑量分割照射腦損傷模型,初步探討糖皮質(zhì)激素在放射治療后海馬損傷中的作用,為臨床防治放射性腦損傷提供理論依據(jù)。 方法將48只昆明小鼠隨機(jī)分成空白組、30Gy模型組、模型+地塞米松治療組、空白+地塞米松治療組,共4小組,每組12只。在清醒狀態(tài)下,用自制模具固定小鼠,放置于直線加速器平臺(tái)上,照射源和平臺(tái)之間放置帶孔鉛模,光源透過(guò)鉛�？装l(fā)出光束精確定位照射腦部,2Gy/次,5次/周,連續(xù)照射3周,累計(jì)劑量30Gy。放射后模型+地塞米松治療組及空白+地塞米松治療組.分別給予地塞米松(dexamethasone, DXM)1mg/KG腹腔注射,照射前、地塞米松治療后通過(guò)Morris水迷宮測(cè)試小鼠的空間學(xué)習(xí)和記憶能力。DXM治療后3周每組隨機(jī)抽出6只采用石蠟切片免疫組織化學(xué)法的方法,觀察DXM前、后星形膠質(zhì)細(xì)胞(GFAP+)附性細(xì)胞數(shù)目,來(lái)評(píng)價(jià)神經(jīng)干細(xì)胞分化和膠質(zhì)細(xì)胞對(duì)DXM治療前、后的反應(yīng)情況,石蠟切片免疫組織化學(xué)的方法檢測(cè)腫瘤壞死因子(TNF-α)(?)的陽(yáng)性細(xì)胞數(shù)目來(lái)評(píng)價(jià)DXM治療前、后中樞神經(jīng)系統(tǒng)神經(jīng)炎性反應(yīng)的情況,并做海馬組織尼氏染色和HE染色等病理切片染色。同時(shí)每組隨機(jī)抽取6只通過(guò)尾靜脈注射伊文思藍(lán)(EB),通過(guò)測(cè)定DXM治療前、后腦組織EB含量和腦組織含水量的改變情況來(lái)評(píng)價(jià)照射后血腦屏障破壞程度。 結(jié)果 1.一般情況：放射后模型組和模型+DXM治療組,出現(xiàn)易激惹、相互撕咬等精神癥狀,模型組出現(xiàn)彗星尾征(見(jiàn)附圖)。模型組和模型+DXM治療組照射后3周出現(xiàn)輕度毛發(fā)脫落,經(jīng)糖皮質(zhì)激素治療后模型+DXM治療組上述癥狀有所減輕。照射前各組小鼠體重?zé)o明顯差別,照射后3周,模型組、模型+DXM治療組體重與空白組相比出現(xiàn)明顯差別,治療后2周,模型+DXM治療組小鼠體重恢復(fù)到空白組水平,模型組小鼠體重雖然出現(xiàn)恢復(fù),但與空白組相比尚未出現(xiàn)明顯差別。空白組、空白+DXM治療組小鼠體重逐漸增加,未見(jiàn)有明顯差別。 2.行為學(xué)測(cè)試：照射前6周,空白組雖然穿越平臺(tái)次數(shù)和第一次穿越平臺(tái)時(shí)間未見(jiàn)有明顯差別,但在第一象限和中環(huán)區(qū)活動(dòng)時(shí)間明顯增多,然而采用糖皮質(zhì)激素干預(yù)后,上述的差別恢復(fù)到空白組水平。模型組小鼠在放射后6周不僅穿越平臺(tái)的次數(shù)明顯下降,第1次找到平臺(tái)的時(shí)間也明顯延長(zhǎng),然而采用激素干預(yù)后,穿越平臺(tái)的次數(shù)、第1次穿越平臺(tái)的時(shí)間、在各象限和各環(huán)尋找平臺(tái)的時(shí)間未見(jiàn)有明顯差別�？瞻�+DXM治療組和模型+DXM治療組行為學(xué)測(cè)試未見(jiàn)有明顯差異。 3.尼氏染色：放射治療后模型組與空白組相比在海馬CA1區(qū)尼氏細(xì)胞數(shù)目明顯減少(P0.01)。地塞米松治療組與模型組相比在海馬CA3區(qū)尼氏細(xì)胞數(shù)目增多(P0.05)。空白+DXM治療組與空白組相比在海馬CA1區(qū)細(xì)胞數(shù)目增多(P0.05)。其余各組未見(jiàn)有明顯的差異。 4.蘇木精-伊紅染色法(hematoxylin-eosin staining)簡(jiǎn)稱(chēng)HE染色法：放射后模型組與空白組相比細(xì)胞數(shù)目在海馬CA1、CA2、CA4區(qū)均沒(méi)有明顯差異(P0.05),模型+DXM治療組與模型組相比在海馬各區(qū)也均無(wú)顯著的統(tǒng)計(jì)學(xué)意義(P0.05),空白+DXM治療組與空白組相比未見(jiàn)有顯著差異(P0.05)。 5. GFAP免疫組化：放射后模型組與空白組相比在海馬CA1、CA2、CA3、CA4各區(qū)GFAP陽(yáng)性細(xì)胞表達(dá)上調(diào)(P0.05),模型+DXM治療組與模型組相比在海馬CA1、CA2、CA3、CA4各區(qū)GFAP陽(yáng)性細(xì)胞數(shù)目表達(dá)增加(P0.05),空白+DXM治療組與空白組相比在海馬CA3區(qū)陽(yáng)性細(xì)胞數(shù)目表達(dá)增加(P0.01)。 6. TNF-α免疫組化：模型組與空白組相比在海馬CA2、CA3、CA4區(qū)TNF-α陽(yáng)性細(xì)胞數(shù)目表達(dá)明顯上調(diào)(P0.05),模型+DXM治療組與模型組相比在海馬CA2、CA3、CA4區(qū)TNF-a陽(yáng)性細(xì)胞數(shù)目表達(dá)顯著下降(P0.05)。空白+DXM治療組與空白組相比在CA4區(qū)陽(yáng)性細(xì)胞數(shù)目表達(dá)明顯增加(P0.05)。 7.放射后血腦屏障的改變：放射后模型組與空白組相比腦組織含水量、EB含量顯著增加(P0.01)。模型+DXM治療組與模型組相比腦組織內(nèi)含水量、EB含量明顯降低(P0.05),空白+DXM治療組與空白組相比沒(méi)有明顯的統(tǒng)計(jì)學(xué)意義(P0.05)。 結(jié)論 1.低劑量分割顱腦照射可以引起與海馬損傷有關(guān)的行為學(xué)改變。 2.糖皮質(zhì)激素治療后海馬區(qū)GFAP、TNF-α明顯表達(dá)下調(diào),腦組織內(nèi)EB含量、含水量也有明顯降低。
[Abstract]:Radiotherapy is one of effective methods in the treatment of head and neck malignant tumors, but also brought to memory loss and cognitive dysfunction as the main symptoms. In learning in the emotional integration of human, plays an important role in memory and other advanced features, caused by damage to the hippocampus has become an increasingly important topic concerned in radiotherapy. Radiotherapy for brain it is related to the damage caused by inflammation, so take anti-inflammatory means of prevention and treatment of radiation encephalopathy has become a research focus, and glucocorticoids have been widely used in clinical, but the mechanism of glucocorticoid to prevent radiation-induced brain injury is not entirely clear. The low dose radiation brain injury model segmentation, preliminary study the role of glucocorticoids in hippocampus injury after radiation therapy, to provide a theoretical basis for clinical prevention and treatment of radiation brain injury.
Methods 48 Kunming mice were randomly divided into blank group, 30Gy model group, model + dexamethasone treatment group, blank + dexamethasone treatment group, a total of 4 groups, 12 rats in each group. In the awake state, using self-made mold fixed mice, placed on the linear accelerator platform, with Kong Qianmo placed between the radiation source and platform, through the light source lead a die hole precise positioning beam brain, 2Gy/ time, 5 times / week, 3 weeks of continuous irradiation, total dose of 30Gy. after radiation model + dexamethasone treatment group and blank + dexamethasone treatment group were given dexamethasone (dexamethasone, DXM) 1mg/KG intraperitoneal injection before irradiation after dexamethasone treatment through spatial learning and memory ability.DXM treatment of the Morris water maze test in mice after 3 weeks in each group were randomly selected 6 paraffin sections by immunohistochemistry, DXM was observed before, after astrocytes (GFAP+) the number of cells And to evaluate the differentiation of neural stem cells and glial cells of DXM before treatment, after the reaction, method of paraffin section immunohistochemical detection of tumor necrosis factor alpha (TNF- alpha) (?) the number of positive cells of DXM before treatment, after central nervous system inflammatory response, and Nissl staining and HE staining pathological sections of hippocampus tissue staining. At the same time randomly selected 6 rats by intravenous injection of Evans blue (EB), through the determination of DXM before treatment, the changes of brain tissue EB content and water content of brain tissue to evaluate the damage of blood brain barrier after irradiation.
Result
1. general situation: model group and +DXM treatment group after radiation model, irritability, mental symptoms such as biting each other, model group, comet tail sign (see photo). Mild hair loss appeared in model group and +DXM treatment group 3 weeks after irradiation, after the treatment with corticosteroids in the treatment group symptom model +DXM reduced. Body weight of mice in each group had no significant difference before irradiation, irradiation after 3 weeks, the model group, +DXM treatment group weight model compared with the blank group appeared obvious difference, after 2 weeks of treatment, the treatment group +DXM model mice weight recovery to the level of the blank control group, body weight of mice in the model group while the recovery, but not yet compared with the control group there was significantly difference. The blank group, blank +DXM treatment group mice body weight increased gradually, there are obvious differences between the No.
Test 2. behavior: 6 weeks before irradiation, the blank group although the number of cross platform and cross platform of the first time there was no significant difference, but significantly increased in the first quadrant and the central area of activity time, however with glucocorticoid intervention, the difference back to the level of the blank group. The mice in the model group after radiation 6 not only the number of weeks of crossing the platform was significantly decreased, the first time to find the platform is also significantly prolonged, but the use of hormone intervention, times of passing through the platform, the first cross platform time in each quadrant and the ring of the time to find the platform did not have significant difference. The blank +DXM treatment group and +DXM treatment group behavior model the test did not have obvious difference.
3. Nissl staining: after radiotherapy in model group compared with blank group decreased significantly in the hippocampal CA1 region of Nissl cell number (P0.01). Dexamethasone treatment group compared with the model group in the CA3 region of the hippocampus Nissl cells (P0.05). The blank +DXM treatment group compared with the control group in the hippocampus CA1 number (P0.05). While there was no significant difference.
4. hematoxylin eosin staining (hematoxylin-eosin staining) HE staining: after radiation model group compared to control group the number of cells in the hippocampus CA1, CA2, there are no significant differences between the CA4 region (P0.05), model +DXM treatment group compared with model group, statistically significant in the hippocampus also were not significant (P0.05) blank, +DXM treatment group compared with the control group there was no significant difference (P0.05).
5. GFAP immunohistochemical staining: after radiation model group compared with blank group in hippocampus CA1, CA2, CA3, CA4 expression of the GFAP positive cells (P0.05), model +DXM treatment group compared with model group in hippocampus CA1, CA2, CA3, CA4 the number of GFAP positive cells increased expression of +DXM (P0.05), blank treatment group compared with the control group the number of positive cells in the hippocampus CA3 expression increased (P0.01).
6. alpha TNF- immunohistochemistry: model group in hippocampus CA2, CA3 compared with the control group, CA4 TNF- alpha positive cells were significantly up-regulated (P0.05) model, +DXM treatment group compared with model group in hippocampus CA2, CA3, CA4, TNF-a positive cell number was significantly decreased (P0.05). The blank treatment +DXM group compared with the control group the number of positive cells in CA4 area was significantly increased (P0.05).
Changes of blood brain barrier after radiotherapy: 7. after radiotherapy of brain tissue compared to the blank group and the model group water content, EB content increased significantly (P0.01). Model +DXM treatment group compared with model group, the brain water content, EB content decreased significantly (P0.05), blank +DXM treatment group and blank group compared with no obvious statistical significance the (P0.05).
conclusion
1. low dose fractionated craniocerebral irradiation can cause behavioral changes associated with hippocampal damage.
After 2. glucocorticoid treatment, the expression of GFAP, TNF- - alpha in the hippocampus, the content of EB in the brain tissue and the water content were also decreased.

【學(xué)位授予單位】：廣西醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R818

【參考文獻(xiàn)】

相關(guān)期刊論文 前4條

1 王琛,謝紅,田野,陳列松,趙培峰;清醒狀態(tài)下大鼠放射性腦損傷實(shí)驗(yàn)?zāi)Ｐ偷慕J];蘇州大學(xué)學(xué)報(bào)(醫(yī)學(xué)版);2004年04期

2 田野,劉春風(fēng),陳列松,張志琳,包仕堯;大鼠早期放射性腦損傷中星形膠質(zhì)細(xì)胞的病理學(xué)改變[J];江蘇醫(yī)藥;2001年05期

3 阮林;韋力;廉春蓉;張文佳;莫立根;李小妹;;全腦照射后血腦屏障改變對(duì)放射性腦損傷的影響[J];中國(guó)神經(jīng)精神疾病雜志;2011年10期

4 葉欽勇,鄭安;甲基強(qiáng)的松龍沖擊治療放射性腦病的臨床研究[J];中國(guó)神經(jīng)免疫學(xué)和神經(jīng)病學(xué)雜志;2003年04期

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本文編號(hào)：1475921