切割穹窿海马伞海马提取液和银杏叶提取物联合诱导海马NSCs向胆碱能神经元的分化

The differentiation of hippocampal neural stem cells(NSCs) into cholinergic neurons induced by the combination of extract from fimbria-fornix transected hippocampus and extract of ginkgo biloba(EGb)

为探讨切割穹窿海马伞海马提取液和银杏叶提取物(extract of ginkgo bilobo,EGb)在海马NSCs向胆碱能神经元定向分化中的作用,分别制备大鼠穹窿海马伞切割侧和正常侧海马提取液;将从鼠胚海马中分离扩增的NSCs球分成4组,应用不同的培养液促其分化:(1)联合组:含切割侧海马提取液和银杏内酯的DMEM/F12培养基;(2)提取液组:含切割侧海马提取液的DMEM/F12培养基;(3)EGb组:含银杏内酯的DMEM/F12培养基;(4)对照组:含正常侧海马提取液的DMEM/F12培养基。培养14d后行ChAT免疫荧光检测,计算ChAT阳性神经元的分化率,图像处理细胞面积和周长。结果显示联合组各项指标均明显优于其它各组(P<0.01);提取液组、EGb组各项指标也均优于对照组(P<0.05);细胞面积提取液组优于EGb组(P<0.05),细胞周长EGb组优于提取液组(P<0.05),两组ChAT阳性神经元分化率无明显差异(P>0.05)。上述提示切割穹窿海马伞的海马提取液和EGb联合应用可诱导海马NSCs分化为更多、更为成熟的胆碱能神经元。

To investigate the effects of extract from fimbria-fornix transected hippocampus and EGb on the proper differentiation of hippocampal NSCs into cholinergic neurons, the hippocampal extracts of fimbria-formix transected side and normal side were prepared respectively. The neurospheres isolated and expanded from hippocampus of fetal rats were divided into 4 groups with different culture media prorooting tbe differentiation. （ 1 ）Combined group： the DMEM/F12 euhure medium containing fimbria-fornix transected hippocampal extract and EGb ; （2） Extract group ： the DMEM/F12 culture medium containing fimbria-fornix transected hippocampal extract ; （3） EGb group ： the1）MEM/F12 culture medium containing EGb; （4）Control group： the DMEM/F12 culture medium containing hippocampal extract of normal side. After 14 days of culture, the immuuofluorescence of ChAT was detected, and then the differential rate of ChAT positive neurons were calculated, the cellular size and perimeter were acquired by image processing. The results showed that all indicators of combined group were superior to those of other groups （ P 〈0.01 ）. The indicators of extract group and EGb group were better than the control group ＇s （ P 〈 0. 05 ）. The extract group was better than the EGb group on cellular area （ P 〈 0.05 ）, but worse than the EGb group on the cellular pcrimeter（ P 〈 0.05 ）. And between the two groups, there was no significant difference for the differential rate of ChAT positive neurons （P 〉0.05 ）. All resnlts suggest that the combination of fimbria-fornix transected hippocampal extract and EGb can induce the hippocampal NSCs differentiating into more and more mature cholinergic neurons.