摘要
目的:观察在体外培养条件下骨髓间充质干细胞对缺氧-复氧新生大鼠大脑皮质神经元活性的影响。方法:实验于2005-01/2006-12在兰州大学第一医院中心实验室完成。健康Wistar大鼠成鼠5只,健康新生Wistar大鼠20只,无菌条件下分离、培养、传代Wistar大鼠骨髓间充质干细胞,细胞融合达90%时更换培养基培养24h,收集细胞培养液即为骨髓间充质干细胞条件培养基;无菌条件下培养新生Wistar大鼠大脑皮质神经元,第8天随机分为正常对照组、缺氧组、间充质干细胞条件培养基处理组,即用骨髓间充质干细胞条件培养基进行缺氧神经元的修复。分别于缺氧0h、缺氧6h、复氧24h用噻唑蓝盐比色法测定吸光度值和BeckMan全自动生化仪测定培养液乳酸脱氢酶漏出量,分别检测各组神经元的活性和细胞损伤后修复的程度。结果:各实验组均全部进入结果分析。①各组神经元的活性:鼠大脑皮质神经元缺氧6h及复氧24h后缺氧组噻唑蓝明显低于正常对照组,骨髓间充质干细胞条件培养基处理组复氧24h后噻唑蓝高于缺氧组(缺氧6h:缺氧组0.42±0.14,正常对照组0.81±0.12;复氧24h:缺氧组0.35±0.15,正常对照组0.82±0.21,骨髓间充质干细胞条件培养基处理组0.74±0.25,P<0.01或0.001)。②各组细胞损伤后修复的程度:缺氧6h和复氧24h后,缺氧组乳酸脱氢酶漏出量比正常对照组明显增多,骨髓间充质干细胞条件培养基处理组比缺氧组明显减少[缺氧6h:缺氧组(790.16±34.51)nkat/L,对照组(340.07±25.17)nkat/L,骨髓间充质干细胞条件培养基处理组(570.11±53.18)nkat/L;复氧24h:缺氧组(1790.36±252.38)nkat/L,对照组(340.07±19.00)nkat/L,骨髓间充质干细胞条件培养基处理组(860.17±40.17)nkat/L,P<0.001]。结论:在体外培养条件下骨髓间充质干细胞能增加缺氧神经元的细胞活性,促进缺氧神经元的修复。
AIM： To observe the influence of bone marrow mesenchymal stem cells （BMMSCs） on the activity of cortical neurons of newborn rats after anoxia-reoxygenation under the condition of in vitro culture. METHODS： From January 2005 to December 2006, the experiment was conducted in the Central Laboratory of the First Hospital of Lanzhou University. Five healthy adult male Wistar rats and 20 healthy newborn Wistar rats were selected. The BMMSCs were separated, cultured and expanded sterilely, and the culture medium was changed when the cells grew to 90%, in which the cells were cultivated for 24 hours. The cell culture medium was collected as the culture medium for BMMSCs. The cortical neurons of newborn rats were cultured in asepsis condition. On day 8, normal control group, anoxic group and BMMSCs conditioned-medium （B-CM） group used to repair the hypoxic neurons were divided randomly. Absorbance value and the leakage of lactate dehydrogenase （LDH） were measured with MTF and BeckMan automatic biochemistry instrument, respectively at anoxia 0, 6 hours and reoxygenation 24 hours respectively to observe the activity of neurons in each group and the recovery after damage. RESULTS： All the groups were involved in the result analysis. ①Activity of neurons： MTT of rat cortical neurons after anoxia 6 hours and resume oxygen 24 hours were obviously lower than the normal control group, and the B-CM group was higher than the anoxia group after reoxygenation 24 hours （anoxia 6 hours： anoxia group： 0.42±0.14, control group： 0.81±0.12; reoxygenation 24 hours： anoxia group： 0.35±0.15, control group： 0.82±0.21, B-CM group： 0.74±0.25, P 〈 0.01 or 0.001）. ②Recovery of cell after damage： After anoxia 6 hours and resume oxygen 24 hours, the leakage of LDH in the anoxia group were significantly increased compared with the control group, and the B-CM group were markedly decreased compared with the anoxia group [anoxia 6 hours： anoxia group （790.16±34.51） nkat/L, normal control group （340.07±25.17） nkat/L, B-CM group （570.11 ±53.18） nkat/L; reoxygenation 24 hours： anoxia group （1 790.36±252.38） nkat/L, normal control group （340.07±19.00） nkat/L, B-CM group （860.17±40.17） nkat/L, P 〈 0.001]. CONCLUSION： BMMSCs cultured in vitro can improve the activity of anoxic neurons and promote the recovery of anoxic neurons.
出处
《中国临床康复》
CSCD
北大核心
2006年第45期18-20,I0004,共4页
Chinese Journal of Clinical Rehabilitation
