本文選題：紅細(xì)胞生成素 + 許旺細(xì)胞��； 參考：《鄭州大學(xué)》2009年碩士論文

【摘要】：目的觀察和評價(jià)紅細(xì)胞生成素(erythropoietin,EPO)對體外培養(yǎng)許旺細(xì)胞(Schwann cells,SCs)分泌神經(jīng)營養(yǎng)因子和細(xì)胞周期的影響,探討EPO促進(jìn)周圍神經(jīng)再生的機(jī)制。 方法選用6只兩周齡的新西蘭兔作為實(shí)驗(yàn)動(dòng)物,經(jīng)75%酒精浸泡20min消毒,無菌條件下取雙側(cè)坐骨神經(jīng)和臂叢神經(jīng),在顯微鏡下剝干凈神經(jīng)外膜,將神經(jīng)剪成0.5cm×0.5cm×0.5cm的小植塊,用植塊法+酶消化法+差速貼壁法培養(yǎng)純化許旺細(xì)胞。實(shí)驗(yàn)設(shè)對照組(A組):20%胎牛血清DMEM液;實(shí)驗(yàn)組1(B組):20%胎牛血清DMEM液+0.5U/ml EPO;實(shí)驗(yàn)組2(C組):20%胎牛血清DMEM液+5U/ml EPO。 1.于細(xì)胞培養(yǎng)第24h、48h、72h分別對各組許旺細(xì)胞應(yīng)用酶聯(lián)免疫吸附試驗(yàn)(ELISA)測定培養(yǎng)上清液中的神經(jīng)生長因子(NGF)和睫狀神經(jīng)營養(yǎng)因子(CNTF)。 2.于細(xì)胞培養(yǎng)第24h分別對各組許旺細(xì)胞進(jìn)行流式細(xì)胞術(shù)檢測細(xì)胞周期分布,比較SCs處于S期百分比(S%)和增殖指數(shù)PI值(S+G_2M)%。 結(jié)果 1.許旺細(xì)胞培養(yǎng)第24h、48h、72h,在各時(shí)間點(diǎn)細(xì)胞培養(yǎng)液中NGF的含量,A組與B組、A組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05),B組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05);在各時(shí)間點(diǎn)細(xì)胞培養(yǎng)液中CNTF的含量,A組與B組、A組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05),B組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05)。 2.許旺細(xì)胞培養(yǎng)第24h,細(xì)胞處于S期百分比(S%),A組與B組、A組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05),B組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05):細(xì)胞增殖指數(shù)PI值(S+G_2M)%,A組與B組、A組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05),B組與C組之間差異有統(tǒng)計(jì)學(xué)意義(P＜0.05)。 結(jié)論 1.EPO可促進(jìn)體外培養(yǎng)許旺細(xì)胞分泌NGF和CNTF,且具有劑量依賴性; 2.EPO可影響體外培養(yǎng)許旺細(xì)胞的細(xì)胞周期,提高許旺細(xì)胞的增殖活性; 3.EPO可提高體外培養(yǎng)許旺細(xì)胞分泌神經(jīng)營養(yǎng)因子的水平,并且能提高許旺細(xì)胞的增殖活性,這可能是其促進(jìn)周圍神經(jīng)再生的作用機(jī)制之一。
[Abstract]:Objective to observe and evaluate the effect of erythropoietin erythropoietin (EPO) on the secretion of neurotrophic factor and cell cycle in Schwann cells of Schwann cells in vitro, and to explore the mechanism of EPO promoting peripheral nerve regeneration. Methods six two-week-old New Zealand rabbits were used as experimental animals. The sciatic nerve and brachial plexus nerve were removed under sterile condition by 75% alcohol immersion in 20min. The nerve was cut off the epineurium under the microscope, and the nerve was cut into the 0.5cm 脳 0.5cm 脳 0.5cm graft. Schwann cells were cultured and purified by enzyme digestion method. The experimental group was divided into two groups: control group (n = 20), control group (n = 20), control group (n = 20), control group (n = 1), group B (n = 1), group B (n = 20), 0.5U/ml (n = 20), and group 2 (n = 2), group C (n = 20), 5U/ml (n = 20). 1. The nerve growth factor NGF (NGF) and ciliary neurotrophic factor (CNTFN) in the supernatant of Schwann cells were determined by enzyme-linked immunosorbent assay (Elisa) at 24 h and 48 h and 72 h after cell culture respectively. 2. The cell cycle distribution of Schwann cells in each group was detected by flow cytometry at 24 hours after cell culture. The percentage of SCs in S phase was compared with the Pi value of proliferative index (Pi). Result 1. The content of NGF in the culture medium of Schwann cells was significantly different between group A and group B at 24 h and 48 h and 72 h after culture. There was significant difference between group A and group B and group C (P < 0.05) and between group B and group C (P < 0.05), and between group A and group B (P < 0.05), and between group B (P < 0.05) and group C (P < 0.05). The content of CNTF was significantly different between group A and group B (P < 0.05) and between group B (P < 0.05) and group C (P < 0.05). 2. At the 24h of Schwann cell culture, the percentage of cells in S phase was significantly different between group A and group B (P < 0.05) and group C (P < 0.05). The Pi value of cell proliferation index (Pi) was higher than that of group A and B (P < 0.05). The difference between group C and group C was statistically significant (P < 0.05) and the difference between group B and group C was statistically significant (P < 0.05). Conclusion 1.EPO can promote the secretion of NGF and CNTF by Schwann cells in vitro in a dose-dependent manner. 2.EPO can affect the cell cycle of Schwann cells cultured in vitro and increase the proliferation activity of Schwann cells. 3.EPO can increase the level of neurotrophic factor secreted by Schwann cells in vitro and increase the proliferative activity of Schwann cells which may be one of the mechanisms of promoting peripheral nerve regeneration.
【學(xué)位授予單位】：鄭州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2009
【分類號】：R329

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