杜仲水提物诱导大鼠肾上腺嗜铬细胞瘤细胞向神经元样细胞分化  

Induction of Neuron-Like Cell Differentiation in PC12 Cells by Water Extract of Duzhong(Eucommia ulmoides Oliver)

作　　者：朱晓岩 李辞霞 周文泰 潘佳蓉 赵善廷[1] ZHU Xiaoyan;LI Cixia;ZHOU Wentai;PAN Jiarong;ZHAO Shanting(College of Veterinary Medicine,Northwest A&F University,Yangling 712100,Shaanxi,China;College of Life Sciences,Northwest A&F University,Yangling 712100,Shaanxi,China)

机构地区：[1]西北农林科技大学动物医学院,陕西杨凌712100 [2]西北农林科技大学生命科学学院,陕西杨凌712100

出　　处：《中华中医药学刊》2023年第5期5-9,I0011,I0012,共7页Chinese Archives of Traditional Chinese Medicine

基　　金：国家重点研发计划-政府间国际科技创新合作重点专项(2018YFE0127000);陕西省重点研发项目(2021NY-021)。

摘　　要：目的 探讨杜仲水提物对大鼠肾上腺嗜铬细胞瘤细胞(Rat pheochromocytoma cells, PC12细胞)向神经元样细胞分化的作用。方法 用0~10 mg·mL^(-1)杜仲水提物培养PC12细胞4 d,双抗噻唑蓝法(MTT法)检测吸光度(A值)并选取最佳浓度配置杜仲诱导培养基;诱导4 d后通过免疫荧光标记神经丝蛋白H(neurofilament-H,NF-H)检测诱导情况;通过免疫蛋白印迹检测细胞外信号调节激酶(extracellular signal-regulated kinase, ERK)及磷酸化细胞外信号调节激酶(phosphorylated extracellular signal regulated kinases, p-ERK)表达情况。结果 不同浓度杜仲培养4 d的PC12细胞,与空白对照A值相比,1~3 mg·mL^(-1)剂量组有增高趋势;4~7 mg·mL^(-1)剂量组有降低趋势;8~10 mg·mL^(-1)剂量组显著降低(P<0.05)。4 mg·mL^(-1)含杜仲诱导培养基培养PC12细胞4 d, 90%以上分化为神经元样细胞,细胞胞体变大,突起伸长,互相连接成网,约20%突起长度> 30μm,显著高于对照组。随着诱导时间的延长,NF-H表达逐渐增多增强,诱导4 d后,约90%以上的细胞NF-H强阳性表达,p-ERK蛋白表达水平较对照组明显降低。结论 杜仲水提物能够诱导PC12细胞分化为神经元样细胞,表达神经元特异蛋白神经丝蛋白,其机制可能与ERK信号通路调节密切相关。Objective To investigate the effect of water extract of Duzhong(Eucommia ulmoides Oliver,WEU)on the differentiation of PC12 cells to neuron-like cells.Methods PC12 cells were cultured with 0-10 mg·mL^(-1) WEU for 4 d.The A value was measured by MTT and the optimal concentration was selected to configure WEU induction medium.The induction was detected by immunofluorescence labeling of neurofilament-H(NF-H)after 4 d of induction.The expressions of extracellular signal-regulated kinase(ERK)and phosphorylated extracellular signal-regulated kinase(p-ERK)were detected by Western blot.Results The A values of PC12 cells cultured for 4 d with different concentrations of WEU were higher in the 1~3 mg·mL^(-1) dose group,lower in the 4~7 mg·mL^(-1) dose group,and significantly lower in the 8~10 mg·mL^(-1) dose group compared with the control(P<0.05).4 mg·mL^(-1) of WEU-induced medium for 4 d showed that more than 90%of PC12 cells differentiated into neuron-like cells,and the cell bodies became larger,the protrusions elongated and interconnected to form a network,and about 20%of the protrusions were>30μm in length,which was significantly higher than that of the control group.With the extension of induction time,NF-H expression gradually increased and enhanced,and after 4 d of induction,more than 90%of the cells were strongly positive for NF-H expression,and the p-ERK protein expression level was significantly lower than that of the control group.Conclusion WEU can induce PC12 cells to differentiate into neuron-like cells and express neuron-specific protein neurofilament protein,the mechanism of which may be closely related to the regulation of ERK signaling pathway.

关 键 词：杜仲水提物 PC12细胞 神经元样细胞 诱导分化 

分 类 号：R285.5[医药卫生—中药学]