枸杞多糖对氯化甲基汞诱导海马神经干细胞损伤的保护作用

Protective effect of lycium barbarum polysaccharides on methyl mercury chloride-induced hippocampal neural stem cells injury

目的观察分析枸杞多糖对氯化甲基汞所诱导神经干细胞(NSCs)损伤的影响。方法取孕16 dSD大鼠胚胎的海马组织,分离纯化得到海马NSCs。将纯化后的海马NSCs增殖、生长10 d后,按随机分组方法将其分为空白对照组(DMEM/F12培养基);枸杞多糖组(DMEM/F12培养基+枸杞多糖);氯化甲基汞组(DMEM/F12培养基+氯化甲基汞);氯化甲基汞+枸杞多糖组(DMEM/F12培养基+氯化甲基汞+枸杞多糖)。分别测定各组生长环境中的海马NSCs分化、生长的情况,观察分析氯化甲基汞对海马NSCs的损伤作用,以及枸杞多糖干预后对海马NSCs的影响。结果氯化甲基汞组和空白对照组比较,加入氯化甲基汞后的海马NSCs分化后的神经元比例(3.63%±0.62%)和神经元的周长(63.36μm±5.57μm)都较空白对照低;枸杞多糖组和其他各组比较,海马NSCs分化后的神经元比例(7.75%±0.59%)和神经元的周长(253.3μm±11.21μm)都较其他各组高;氯化甲基汞+枸杞多糖组和氯化甲基汞组比较,前者生长环境下的海马NSCs分化后的神经元比例(5.92%±0.98%)和神经元的周长(111.9μm±6.07μm)较氯化甲基汞组高。结论氯化甲基汞对海马NSCs的分化、生长具有损伤作用;枸杞多糖可以减轻氯化甲基汞对海马NSCs的损伤作用,并能促进海马NSCs向神经元的分化及神经元的生长。

Objective To determine the protective roles of lycium barbarum polysaceharide on methyl mercury chloride-induced injury for neuronal differentiation of hippocampal neural stern cells （NSCs）. Methods NSCs were collected from the hippocampus of 16-embryonic day Sprague-Dawley rats. The cells were cuhivated in neural stem cell- specific medium for 10 days. The neurospheres were dissociated into single cells and cultured in the 24-well plates with poly-L-lysine-coated cover glass. Cells were divided into four groups： a control group in which the cells were cultured in DMEM/F12 medium; a lycium barbarum polysaccharide group-the cells were cultured in DMEM/F12 with lycium barbarum polysaccharide; a methyl mercury chloride group-the cells were cultured in DMEM/F12 with methyl mercury chloride; Methyl mercury chloride ＋ lycium barbarum polysaccharide group-the cells were cultured in DMEM/F12 with methyl inereury chloride and lycium barbarum polysaccharide. The cell growth and differentiation were determined by immunostaining against M AP-2 or GFAP antibody. Results The percentage （ 3.63% ± 0.62% ） and average perimeter（63. 36μm ± 5. 57 μm） of the differentiated neurons in the methyl mercury chloride group were lower than that in the control group, while the percentage （7.75% ±0. 59% ） and average perimeter （253.31μm ± 11. 211μm） of them in lyeium barbarum polysaecharide group were much more than all other groups. Compared with the methyl mercury chloride group, after methyl mercury chloride treatment the lyeium barbarum polysaccharide increased the neuronal differentiation （5.92% ± 0.98% ） to the level in control group and their average perimeter （ 111.9μm ± 6.07 μm） in this group also showed 2-fold increase. Conclusion Lyeium barbarum polysaceharide promotes the NSCs to differentiate into neurons and improves the growth of them. Methyl mercury chloride exertes the damage on NSCs differentiation and growth of neurons that can be rescued by lycium barbarum polysaccharide.