本文選題：電離輻射 + 放射性腦損傷　； 參考：《揚(yáng)州大學(xué)》2013年碩士論文

【摘要】：隨著醫(yī)學(xué)放射技術(shù)的發(fā)展,放射技術(shù)廣泛應(yīng)用于醫(yī)學(xué)中的診斷和治療。在疾病診斷中,X射線檢查如X線射片、數(shù)字減影血管造影(DSA)、X射線計算機(jī)斷層掃描(CT)和CT血管成像等在醫(yī)學(xué)實(shí)踐中的應(yīng)用非常普遍；在疾病治療中,現(xiàn)代放射治療是治療惡性腫瘤的主要手段之一,有時是唯一的手段。然而隨著放射技術(shù)的不斷改進(jìn)及臨床中廣泛應(yīng)用,電離輻射潛在致癌風(fēng)險及放射性損傷備受關(guān)注,因此研究電離輻射致癌及放射性損傷的機(jī)理對放射學(xué)的發(fā)展有重要的意義。 研究發(fā)現(xiàn)一類新的非編碼RNA—microRNA(miRNA)參與輻射誘導(dǎo)的生物學(xué)反應(yīng)過程,研究表明輻射能在不同的組織和細(xì)胞中誘導(dǎo)miRNA的表達(dá)改變；miRNA在大腦的發(fā)育和功能中有很重要的作用。因此研究miRNA如何參與電離輻射引起的生物效應(yīng)對揭示輻射的分子生物學(xué)機(jī)制及潛在的臨床應(yīng)用價值具有重要意義,miRNA有可能成為放射病診斷和治療的潛在新靶點(diǎn)。 本研究擬探索放射性腦損傷早期miRNA的差異表達(dá),為進(jìn)一步研究提供依據(jù)。而腦組織的放射性損傷主要特征之一是血腦屏障的破壞。腦微血管內(nèi)皮細(xì)胞是血腦屏障的主要組成細(xì)胞,因此本實(shí)驗(yàn)選擇腦微血管內(nèi)皮細(xì)胞模擬體外放射模型。 一、大鼠腦微血管內(nèi)皮細(xì)胞培養(yǎng)的實(shí)驗(yàn)研究 目的獲取miRNA芯片所需的大鼠腦微血管內(nèi)皮細(xì)胞(BMVECs)。 方法采用兩次酶消化法、密度梯度離心法獲取BMVECs;應(yīng)用vWF和GFAP免疫熒光雙重標(biāo)記方法進(jìn)行BMVECs鑒定。 結(jié)果成功在體外培養(yǎng)出BMVECs,免疫熒光標(biāo)記為vWF標(biāo)記陽性、GFAP標(biāo)記陰性。 二、電離輻射后大鼠腦微血管內(nèi)皮細(xì)胞的miRNA表達(dá)譜變化的實(shí)驗(yàn)研究 目的觀察電離輻射后BMVECs中miRNA表達(dá)的差異。 方法對融合的BMVECs采用6MV-X線照射,照射劑量為10Gy,用miRNA芯片分析照射組和對照組miRNA表達(dá)差異,選取目標(biāo)miRNA進(jìn)行RT-qPCR驗(yàn)證。 結(jié)果基因芯片結(jié)果中顯示總共有18個miRNA在照射后發(fā)生差異表達(dá),其中7個上調(diào),11個下調(diào)；miR-34a上調(diào)10.8倍,miR-347上調(diào)約17.9倍,miR-106b約下調(diào)3.6倍。RT-qPCR驗(yàn)證顯示照射組BMVECs的miR-34a、miR-347和miR-106b表達(dá)譜改變和芯片結(jié)果相符合。 三、大鼠單次全腦照射后早期大腦皮層miRNA表達(dá)改變的實(shí)驗(yàn)研究 目的分別觀察在0、2、10和30Gy大鼠單次全腦照射后,1天和7天時大腦皮層目標(biāo)miRNA表達(dá)的改變。 方法建立0、2、10和30Gy照射劑量大鼠單次全腦照射模型,分別在1天和7天時取大腦皮層,使用RT-qPCR觀察miR-106b、miR-34a和miR-34表達(dá)譜改變。 結(jié)果與OGy照射組比較,照射后1天,2Gy組miR-106b表達(dá)無顯著差異、miR-34a約上調(diào)6.4倍、miR-347約上調(diào)2倍；10Gy組miR-106b約下調(diào)1.7倍、miR-34a約上調(diào)5.3倍、miR-347表達(dá)無顯著差異；30Gy組miR-106b約下調(diào)2.5倍、miR-34a表達(dá)無顯著差異、miR-347約下調(diào)1.6倍。照射后1周,2Gy組miR-106b表達(dá)無顯著差異、miR-34a約上調(diào)1.4倍、miR-347約上調(diào)2.3倍；10Gy組miR-106b表達(dá)無顯著差異、miR-34a約上調(diào)2.6倍、miR-347約上調(diào)5.3倍；30Gy組miR-106b約下調(diào)1.4倍、miR-34a約上調(diào)1.8倍、miR-347約上調(diào)3.3倍。 結(jié)論 1.通過兩次酶消化法和密度梯度離心法可獲得較理想的BMVECs,經(jīng)vWF免疫熒光標(biāo)記陽性、GFAP標(biāo)記陰性。 2.照射后早期即發(fā)生miRNA表達(dá)譜的改變,miRNA芯片提示總共有18個miRNA在照射后發(fā)生差異表達(dá),其中7個上調(diào),11個下調(diào)；應(yīng)用RT-qPCR對芯片結(jié)果進(jìn)行驗(yàn)證,結(jié)果顯示照射組BMVECs的]miR-34a、miR-106b和miR-347表達(dá)譜改變和芯片結(jié)果相符合。 3.使用RT-qPCR觀察大鼠大腦皮層放射后不同時間點(diǎn)和不同照射劑量下目標(biāo)miRNA表達(dá)譜的改變,照射后miR-106b表達(dá)下調(diào)和miR-34a表達(dá)上調(diào)可能與放射性腦損傷的損傷機(jī)制相關(guān)；miR-347表達(dá)上調(diào)可能與放射性腦損傷的保護(hù)機(jī)制相關(guān)。
[Abstract]:With the development of medical radiation technology, radiation technology is widely used in the diagnosis and treatment of medicine. In the diagnosis of disease, X ray examination, such as X-ray radiography, digital subtraction angiography (DSA), X ray computed tomography (CT) and CT angiography, is very common in medical practice; in the treatment of disease, modern radiation therapy is One of the main means for the treatment of malignant tumors is sometimes the only means. However, with the continuous improvement of the radiation technology and the wide application of the clinic, the potential carcinogenic risk and radiation damage of ionizing radiation are concerned. Therefore, it is of great significance to study the mechanism of ionizing radiation carcinogenesis and radioactivity damage to the development of radiology.
A new class of non coded RNA - microRNA (miRNA) has been found to be involved in radiation induced biological reactions. Research shows that radiation can induce changes in the expression of miRNA in different tissues and cells; miRNA plays an important role in the development and function of the brain. Therefore, the study of how miRNA participates in the biological effects caused by ionizing radiation. It is of great significance to reveal the molecular biological mechanism of radiation and its potential clinical value. MiRNA may be a potential new target for the diagnosis and treatment of radiation sickness.
This study is to explore the differential expression of miRNA in the early stage of radiation brain injury and provide a basis for further research. One of the main characteristics of the brain tissue damage is the destruction of the blood brain barrier. The brain microvascular endothelial cells are the main components of the blood brain barrier, so this experiment chooses the model of the brain microvascular endothelial cells to simulate the radiological model in vitro.
Experimental study on culture of rat brain microvascular endothelial cells
Objective to obtain rat brain microvascular endothelial cells (BMVECs) required for miRNA chip.
Methods BMVECs was obtained by two enzyme digestion and density gradient centrifugation. BMVECs was identified by vWF and GFAP immunofluorescence double labeling.
Results BMVECs was successfully cultured in vitro. Immunofluorescence labeling was positive for vWF and negative for GFAP.
Two, the change of miRNA expression profile in rat brain microvascular endothelial cells after ionizing radiation
Objective To observe the difference of miRNA expression in BMVECs after ionizing radiation.
Methods the fusion BMVECs was irradiated with 6MV- X-ray and the dose of irradiation was 10Gy. The difference of miRNA expression between the irradiated group and the control group was analyzed with miRNA chip, and the target miRNA was selected for RT-qPCR verification.
Results the results showed that 18 miRNA were expressed differently after irradiation, including 7 up-regulated and 11 down-regulation, miR-34a up 10.8 times up, miR-347 up-regulated about 17.9 times, and miR-106b down regulation 3.6 times.RT-qPCR verification that the miR-34a of BMVECs in the irradiated group, miR-347 and miR-106b expression profiles were in accordance with the results of the chip.
Three, the change of miRNA expression in the early cerebral cortex after single whole brain irradiation in rats
Objective To observe the changes of miRNA expression in cerebral cortex of rats in 0,2,10 and 30Gy after 1 days and 7 days respectively.
Methods a single whole brain irradiation model of 0,2,10 and 30Gy irradiated rats was established. The cerebral cortex was taken at 1 days and 7 days respectively. The expression of miR-106b, miR-34a and miR-34 were observed by RT-qPCR.
Results compared with the OGy group, there was no significant difference in miR-106b expression in group 2Gy 1 days after irradiation, miR-34a was up to about up to 6.4 times, miR-347 was about 2 times up, miR-106b in 10Gy group was about 1.7 times down, miR-34a was up to 5.3 times, miR-347 expression was no significant difference, 30Gy miR-106b was about 2.5 times down, miR-34a expression was 1.6 times down. After 1 weeks, there was no significant difference in miR-106b expression in group 2Gy, miR-34a about up 1.4 times, miR-347 up to 2.3 times up, miR-106b expression in 10Gy group was not significantly different, miR-34a was about up 2.6 times, miR-347 was about up 5.3 times, miR-106b of 30Gy group was about 1.4 times down, miR-34a was about 1.8 times up, miR-347 approximately up 3.3 times.
conclusion
1. the ideal BMVECs can be obtained by two enzyme digestion and density gradient centrifugation. VWF is positive by immunofluorescence and negative by GFAP.
The expression profiles of miRNA were changed at the early stage of 2. irradiation. The miRNA chip suggested that 18 miRNA were expressed differently after irradiation, of which 7 were up and 11 down, and RT-qPCR was used to verify the results of the chip. The results showed that the]miR-34a of BMVECs in the irradiated group, miR-106b and miR-347 expression profiles were in accordance with the results of the chip.
3. RT-qPCR was used to observe the changes of target miRNA expression profiles at different time points and different doses of radiation in the cerebral cortex of rats. Down regulation of miR-106b expression and up regulation of miR-34a expression after irradiation may be related to the damage mechanism of radioactive brain injury, and the up-regulation of miR-347 expression may be related to the protective mechanism of radiation brain injury.
【學(xué)位授予單位】：揚(yáng)州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R818

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 陳勝利,黃齊好,朱棟梁,鄒蓉珠,黃子誠,陳國東,慕容愛;腦血管介入放射診治中患者的X射線輻射評價[J];中國輻射衛(wèi)生;2004年02期

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本文編號：1970841