人脐带间充质干细胞条件培养液对Aβ损伤原代海马神经元凋亡的影响

Effects of human umbilical cord mesenchymal stem cells conditioned medium on the apoptosis of hippocampus neuron induced by amyloid β in cultured rat hippocampus neuron in vitro

目的探讨人脐带间充质干细胞条件培养液(human umbilical cord mesenchymal stem cells conditioned medium,h UCMSCs CM)对体外培养的Aβ(Amyloidβ,β-样淀粉蛋白)损伤海马神经元凋亡的影响。方法分离、培养人脐带间充质干细胞,第3~4代的细胞用于收集条件培养液;分离、培养大鼠海马神经元,培养7~8 d的神经元用于实验分为h UCMSCs CM对神经元细胞活力的影响:分为溶剂对照组、Aβ损伤组、Aβ+h UCMSCs CM低剂量组、Aβ+h UCMSCs CM中剂量组、Aβ+h UCMSCs CM高剂量组。通过MTT检测神经元活力;通过DAPI染色观察5组神经元凋亡情况;通过Western Blot分析神经元cyt-c蛋白水平。结果与溶剂对照组相比,Aβ可降低体外培养的原代海马神经元细胞活力,作用呈浓度、时间依赖性(P<0.05);与Aβ损伤组相比,h UCMSCs CM可对抗Aβ引起的细胞活力下降(P<0.05)。与溶剂对照组相比,Aβ(终浓度10μmol/L)处理12 h显著增加体外培养的原代海马神经元细胞凋亡;与Aβ损伤组相比,h UCMSCs CM可减少Aβ诱导的神经元凋亡,抑制Aβ引起的cyt-c释放,作用呈浓度依赖性(P<0.05)。结论人脐带间充质干细胞条件培养液可抑制Aβ引起的原代培养海马神经元凋亡,发挥神经保护作用,其机制可能与抑制cyt-c释放有关。

Objective To investigate the effects of human umbilical cord mesenchymal stem cells conditioned medium(hUCMSCsCM)on the apoptosis of hippocampus neuron induced by amyloidβ(Aβ)in cultured rat hippocampus neuron in vitro.Methods The neurons cultured for 7~8d were treated with Aβand/or hUCMSCsCM at different concentrations,respectively.The experiment was divided into four groups:solvent control group,Aβinjury group,low dose Aβ+hUCMSCsCM group and median dose Aβ+hUCMSCsCM.The neuron viability was determined by MTT assay.The neuron apoptosis was detected by DAPI staining.The levels of cyt-c protein were measured by Western Blot.Results As compared with solvent control group,Aβcould decrease the neuron viability of primary generation hippocampus neuron in a dose and time-dependent manner(P<0.05).As compared with Aβinjury group,hUCMSCsCM could alleviate the increase of cell vitality caused by Aβ(P<0.05).As compared with solvent control group,Aβwith final concentration of 10μmol/L for 12h increased obviously the apoptosis of primary culture hippocampus neuron.As compared with Aβinjury group,and hUCMSCsCM could decrease neuron apoptosis induced by Aβ,which could inhibit cyt-c release caused by Aβin a dose-dependent manner(P<0.05).Conclusion The human umbilical cord mesenchymal stem cells conditioned medium can inhibit the apoptosis of primary culture hippocampus neuron caused by Aβ,which has neuroprotective effects,and its actin mechanism may be correlated with inhibiting the release of cyt-c in vitro.