【摘要】： 目的:研究硫酸鎂對(duì)神經(jīng)干細(xì)胞輻射損傷的保護(hù)作用。 方法:取新生24h內(nèi)的SD大鼠腦組織,進(jìn)行神經(jīng)干細(xì)胞的原代培養(yǎng)并進(jìn)行鑒定。將神經(jīng)干細(xì)胞分為空白對(duì)照組、實(shí)驗(yàn)對(duì)照組和實(shí)驗(yàn)用藥組,分別用2Gy、4Gy ~(60)Coγ射線對(duì)實(shí)驗(yàn)組及實(shí)驗(yàn)對(duì)照組進(jìn)行照射,照射后24h、48h、72h用CCK-8檢測(cè)神經(jīng)干細(xì)胞的增殖,流式細(xì)胞儀檢測(cè)凋亡及周期變化,透射電鏡觀察細(xì)胞核的形態(tài)變化,并與實(shí)驗(yàn)對(duì)照組進(jìn)行比較。用SAS統(tǒng)計(jì)軟件包進(jìn)行數(shù)據(jù)處理。 結(jié)果: 1.分別用絲裂元進(jìn)行神經(jīng)干細(xì)胞培養(yǎng)傳代后, EGF培養(yǎng)的神經(jīng)干細(xì)胞主要分化為GFAP陽(yáng)性的神經(jīng)膠質(zhì)細(xì)胞。而EGF及bFGF聯(lián)合進(jìn)行培養(yǎng)的神經(jīng)干細(xì)胞,分化為NSE陽(yáng)性的神經(jīng)元細(xì)胞比例明顯較單用EGF組增多,GFAP陽(yáng)性神經(jīng)膠質(zhì)細(xì)胞也明顯減少。 2.在神經(jīng)干細(xì)胞增殖情況的研究中,發(fā)現(xiàn)不同濃度的硫酸鎂對(duì)神經(jīng)干細(xì)胞增殖情況有不同的影響。當(dāng)濃度大于5mg/ml時(shí)將導(dǎo)致神經(jīng)干細(xì)胞的死亡,當(dāng)其濃度為1.25mg/ml、2.5mg/ml時(shí),硫酸鎂將能促進(jìn)神經(jīng)干細(xì)胞的增殖。 3.將神經(jīng)干細(xì)胞傳代后,分別用2Gy及4Gy ~(60)Coγ射線照射神經(jīng)干細(xì)胞,照射后24h、48h、72h時(shí)間點(diǎn)發(fā)現(xiàn),實(shí)驗(yàn)對(duì)照組與空白對(duì)照組比較其增殖明顯降低(p0.05),實(shí)驗(yàn)用藥組與實(shí)驗(yàn)對(duì)照組比較其增殖情況明顯好轉(zhuǎn),除24h時(shí)間點(diǎn)外,各時(shí)間點(diǎn)比較均有統(tǒng)計(jì)學(xué)意義(p0.05)。 4.神經(jīng)干細(xì)胞經(jīng)2Gy、4Gy ~(60)Coγ射線照射后24h、48h、72h研究發(fā)現(xiàn),與空白對(duì)照組比較實(shí)驗(yàn)對(duì)照組其凋亡率明顯增加(p0.05),細(xì)胞核固縮、核仁邊集,而各時(shí)間點(diǎn)實(shí)驗(yàn)用藥組與實(shí)驗(yàn)對(duì)照組比較,其凋亡率明顯好轉(zhuǎn)(p0.05),細(xì)胞核形態(tài)明顯好轉(zhuǎn),更接近空白對(duì)照組。 5.神經(jīng)干細(xì)胞經(jīng)2Gy、4Gy ~(60)Coγ射線照射后培養(yǎng)24h、48h、72h,與空白對(duì)照組比較,實(shí)驗(yàn)對(duì)照組細(xì)胞G1期阻滯明顯增加(p0.05),同時(shí)G2期也有輕微的阻滯,且4Gy組較2Gy組,G2期阻滯更明顯。實(shí)驗(yàn)用藥組與實(shí)驗(yàn)對(duì)照組比較,各時(shí)間點(diǎn)G1期阻滯明顯降低(p0.05),更接近空白對(duì)照組。 結(jié)論: 1. EGF具有促進(jìn)神經(jīng)干細(xì)胞向膠質(zhì)細(xì)胞分化的趨勢(shì)。bFGF與EGF同時(shí)培養(yǎng)時(shí),能使神經(jīng)干細(xì)胞分化為神經(jīng)元的比例增加。 2.當(dāng)硫酸鎂的濃度為1.25mg/ml、2.5mg/ml時(shí)能促進(jìn)神經(jīng)干細(xì)胞增殖。 3.早期運(yùn)用硫酸鎂可阻止電離輻射對(duì)神經(jīng)干細(xì)胞增殖的損傷;使電離輻射所致神經(jīng)干細(xì)胞凋亡率降低,對(duì)電離輻射所致神經(jīng)干細(xì)胞損傷起到保護(hù)作用;同時(shí)能夠降低電離輻射所致神經(jīng)干細(xì)胞G1期阻滯,減輕電離輻射對(duì)神經(jīng)干細(xì)胞的損傷。
[Abstract]:Objective: to study the protective effect of magnesium sulfate on radiation injury of neural stem cells. Methods: the brain tissues of SD rats within 24 hours were collected and the primary culture and identification of neural stem cells were carried out. Neural stem cells were divided into blank control group, experimental control group and experimental drug group. The experimental group and experimental control group were irradiated with 2 Gy ~ (4) Gy ~ (60) Co 緯 -ray, respectively. The proliferation of neural stem cells was detected by CCK-8 at 24 h, 48 h and 72 h after irradiation. Apoptosis and cycle changes were detected by flow cytometry and morphological changes of nucleus were observed by transmission electron microscope and compared with experimental control group. The data are processed by SAS statistical software package. Results: 1. After the neural stem cells were cultured and subcultured with mitogen, the neural stem cells cultured with EGF mainly differentiated into GFAP positive glial cells. However, the proportion of neural stem cells differentiated into NSE positive neurons in EGF and bFGF co-culture group was significantly higher than that in EGF group alone, and the number of GFAP positive glial cells was also significantly decreased. 2. In the study of the proliferation of neural stem cells, it was found that different concentrations of magnesium sulfate had different effects on the proliferation of neural stem cells. When the concentration is higher than 5mg/ml, the neural stem cells will die. When the concentration is 1.25 mg / ml or 2.5 mg / ml, magnesium sulfate can promote the proliferation of neural stem cells. 3. The neural stem cells were irradiated with 2Gy and 4Gy ~ (60) Co 緯 -rays respectively. The proliferation of neural stem cells in the experimental control group was significantly lower than that in the blank control group (p0.05). Compared with the experimental control group, the proliferation of the experimental group was improved obviously, except 24 h time point, the comparison of each time point had statistical significance (p0.05). 4. Compared with the control group, the apoptosis rate of NSCs was significantly increased (p0.05), nuclear pyknosis and nucleolar edge aggregation were observed in the experimental control group after 24 h ~ (48 h) irradiation with 2 Gy ~ (4) Gy ~ (60) Co 緯 -ray. Compared with the control group, the apoptotic rate and nuclear morphology of the experimental drug group were significantly improved (p0.05), which were closer to the blank control group. 5. Neural stem cells were irradiated with 2 Gy ~ (4) Gy ~ (60) Co 緯 -ray for 24 h or 48 h and 72 h after irradiation. Compared with the control group, the G _ 1 phase arrest was significantly increased in the experimental control group (p0.05), and the G _ 2 phase was also slightly blocked in the 4Gy group compared with the 2Gy group. G 2 arrest was more obvious. Compared with the experimental control group, the G1 phase block in the experimental group was significantly decreased (p0.05), which was closer to the blank control group. Conclusion: 1. EGF could promote the differentiation of neural stem cells into glial cells. When bFGF and EGF were cultured at the same time, the proportion of neural stem cells differentiated into neurons was increased. 2. When the concentration of magnesium sulfate is 1.25 mg / ml, 2.5 mg / ml, it can promote the proliferation of neural stem cells. 3. Early application of magnesium sulfate could prevent the proliferation of neural stem cells induced by ionizing radiation, decrease the apoptosis rate of neural stem cells induced by ionizing radiation, and play a protective role on the injury of neural stem cells induced by ionizing radiation. At the same time, it can reduce the G 1 arrest of neural stem cells induced by ionizing radiation, and alleviate the injury of neural stem cells induced by ionizing radiation.
【學(xué)位授予單位】：蘇州大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R363

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