發(fā)布時間：2018-05-22 08:36

  本文選題：脊髓損傷 + Taxol�。� 參考：《西南醫(yī)科大學(xué)》2017年碩士論文

【摘要】：目的:研究模擬在脊髓損傷的微環(huán)境中,Taxol與Y27632聯(lián)合應(yīng)用對新生SD (Sprague Dawley,SD)大鼠背根神經(jīng)節(jié)神經(jīng)元的軸突生長的影響。方法:①21只SD大鼠隨機(jī)分為對照組7只,手術(shù)組7只(用改良Allen法制作雌性成年SD大鼠T9平面完全性截癱模型),假手術(shù)對照組7只。制模后7天后取出T8～T10段脊髓,并經(jīng)過分離、過濾形成組織勻液制備脊髓萃取液。②分離、培養(yǎng)以及鑒定新生SD大鼠的背根神經(jīng)節(jié)神經(jīng)元(dorsal root ganglion neurons, DRGNs)。③分組培養(yǎng)、觀察脊髓萃取液對DRG神經(jīng)元軸突生長,分別為:A組DRGNs+50μlPBS,B組DRGNs+50μl假手術(shù)組脊髓萃取液,C組DRGNs+50μl損傷脊髓萃取液,D組DRGNs+50μl對照組脊髓萃取液。培養(yǎng)2d后,各組于倒置顯微鏡下測量軸突生長的平均長度,然后運(yùn)用免疫熒光染色技術(shù),測量軸突遠(yuǎn)端微管熒光平均密度。實驗2: DRGNs、脊髓萃取液分別和共同與Y27632,Taxol培養(yǎng)、DRG神經(jīng)元軸突生長。共分為五組,A組為DRGNs+50μlPBS,B組為DRGNs+50μl脊髓萃取液(假手術(shù)組);C組為DRGNs+50μl完全性截癱大鼠脊髓萃取液+3nM Taxol,D組為DRGNs+50μl完全性截癱大鼠脊髓萃取液+30μM Y27632; E組,DRGNs+50μl完全性截癱大鼠脊髓萃取液+3nM Taxol+30μMY27632。培養(yǎng)1d,2d后于倒置顯微鏡下測量軸突生長的平均長度,然后運(yùn)用免疫熒光染色技術(shù),測量軸突遠(yuǎn)端微管熒光平均密度。結(jié)果:①實驗1:共同培養(yǎng)2d后,各組軸突生長平均長度:A組為 142.1 ±5.7μm,B組為 144.8±3.1μm,C組為47.2±3.5μm,D組為136.9±4.2μm。單因素方差分析示:C組和A、B、D組之間,P0.01,有顯著的統(tǒng)計學(xué)差異;軸突遠(yuǎn)端平均微管熒光密度,A組為175.1±2.9 AFU/μm,B組為 191.6±3.1 AFU/μm,C組為64.5± 1.5 AFU/μm,D組為 174.8±3.1AFU/μm。C組和A、B、D組相比較,P0.01,有明顯的統(tǒng)計學(xué)差異。②實驗2:共同培養(yǎng)1d后,各組軸突平均長度:A組為136.5±4.6μm,B組為47.6±3.9μm,C組為 137.9±5.2μm, D組為 158.6±4.6μm,E組為 181.2±5.2μm,單因素方差分析示: B組和A、C、D及E組相對比,P0.01,有顯明的統(tǒng)計學(xué)上的差異。2d后,軸突平均長度,A組為142.9±1.4μm,B組為44.5±2.2μm,C組為152.6±5.3μm,D組為168.1±3.1μm,E組為194.6±3.2μm。單因素方差分析示:B組與A、C、D及E之間,P0.01,有明顯的統(tǒng)計學(xué)差異。C組和D、E組相比較,P0. 05,有明顯的統(tǒng)計學(xué)差異。E組和A、B、C、D組相比較,P0.01,有明顯的統(tǒng)計學(xué)差異;A組與C組之間,P0. 05,兩者無統(tǒng)計學(xué)差異。軸突遠(yuǎn)端平均微管熒光密度,A組為181.8±3.4AFU/μm,B組為61.6±2.7AFU/μm,C組為205.2±2.1AFU/μM, D 組為 401.2±3.5 AFU/μM,E組為470.2±3.6AFU/μm。B組和A、C、D、E組相比較,P0. 01,有明顯的統(tǒng)計學(xué)差異。E組和A、B、C、D組相比較,P0. 01,有明顯的統(tǒng)計學(xué)差異;C組與E組之間,P0. 05,兩者無統(tǒng)計學(xué)差異。結(jié)論:①DRGNs置于完全性截癱大鼠脊髓萃取液中培養(yǎng),其軸突的再生和延長受到抑制,經(jīng)過前期實驗結(jié)果,證實完全性截癱大鼠脊髓萃取液能模擬脊髓損傷的微環(huán)境,抑制新生SD大鼠DRGNs軸突的生長;②單獨使用適合劑量的Taxol與Y27632能積極地增進(jìn)DRGNs軸突生長,由于生長錐形成。聯(lián)合應(yīng)用Taxol與Y27632對新生大鼠DRGNs軸突生長促進(jìn)作用效果比單一應(yīng)用更為有效,形成多個分支、甚至神經(jīng)網(wǎng)。
[Abstract]:Objective: To investigate the effects of the combined application of Taxol and Y27632 on the axon growth of dorsal root ganglion neurons in neonatal SD (Sprague Dawley, SD) rats. Methods: (1) 21 rats were randomly divided into 7 rats in the control group and 7 in the operation group (using the modified Allen method to make the female adult SD rat T9 plane complete paraplegia model). 7 rats in the sham operation control group were taken out of the T8 ~ T10 segment after 7 days, and the spinal extract was prepared by filtration to form tissue homogenate. (2) isolation, culture and identification of the dorsal root ganglion neurons (dorsal root ganglion neurons, DRGNs) in newborn SD rats. (3) group culture to observe the axon growth of the spinal cord extraction solution to the DRG neuron. A group DRGNs+50 mu lPBS, B group DRGNs+50 Mu l sham group spinal extract fluid, C group DRGNs+50 u l injured spinal extract fluid, D group DRGNs+50 micron l control group spinal extract fluid. After culture, each group measured the average length of the axon growth under the inverted microscope, and then used immunofluorescence staining technique to measure the average density of the distal axon microtubule fluorescence density Degree. 2: DRGNs, spinal extract fluid and CO and Y27632, Taxol culture, and DRG neuron axon growth, divided into five groups, A group DRGNs+50 mu lPBS, B group DRGNs+50 Mu l spinal extract fluid (sham operation group), and C group was the spinal extract of complete paraplegia rats Liquid +30 mu M Y27632; E group, DRGNs+50 Mu l complete paraplegic rat spinal extract +3nM Taxol+30 micron MY27632. 1D, 2D after the inverted microscope to measure the average length of the axon growth, and then using immunofluorescence staining technique to measure the average fluorescence average density of the distal axon microtubule. Average length: A group was 142.1 + 5.7 mu m, B group was 144.8 + 3.1 mu m, C group was 47.2 + 3.5 m, D group was 136.9 + 4.2 u M. single factor analysis of variance: C group and A, B, D groups, there were significant statistical differences; the average microtubule fluorescence density of the distal axon was 175.1 + 2.9 The average length of axon in each group was 136.5 + 4.6 mu m, B group was 47.6 + 3.9 mu m, 137.9 + 5.2 micron, 158.6 + 4.6 micron, and 181.2 + 5.2 micron, and single factor analysis of variance analysis showed that after 2: co culture of 1D, the average axon length of each group was compared with that of A, B and D group. 0.01, after statistically significant difference.2d, the average length of axon, A group was 142.9 + 1.4 mu m, B group was 44.5 + 2.2 mu m, C group was 152.6 + 5.3 mu m, D group was 168.1 + 3.1 m, E group was 194.6 + 3.2 M. single factor variance analysis. The difference between the.E group and the group of A, B, C, and D, there were significant statistical differences. There was no statistical difference between the A group and the C group. There was no statistical difference between the group A and the C group. The average microtubule fluorescence density of the distal axon was 181.8 + 3.4AFU/ micron, the group was 61.6 + 205.2 mu, and the group was 401.2 + 3.5. E group compared, P0. 01, there was significant statistical difference between group.E and A, B, C, D group compared, P0. 01, there were significant statistical differences; C group and E group, P0. 05, there was no statistical difference. The spinal extract solution of complete paraplegic rats can simulate the microenvironment of spinal cord injury and inhibit the growth of DRGNs axon in newborn SD rats. (2) the use of suitable dose of Taxol and Y27632 can actively promote the growth of DRGNs axon and form the growth cone. The effect of combined application of Taxol and Y27632 on the growth of DRGNs axon of neonatal rats is more than single. Applications are more effective, forming multiple branches or even neural networks.
【學(xué)位授予單位】：西南醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R651.2

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