知母皂苷BⅡ对大鼠神经干细胞向酪氨酸羟化酶阳性神经元分化的影响  

Effects of timosaponin B-II on differentiation of rat neural stem cells into tyrosine hydroxylase positive neurons in vitro

作　　者：张敏娜[1] 王婷婷[2] 钟鸣[3] 王思民 邓雯霞 王光辉[3] 郑公朴 周采菊 黄成钢 Zhang Minna;Wang Tingting;Zhong Ming;Wang Simin;Deng Wenxia;Wang Guanghui;Zheng Gongpu;Zhou Caiju;Huang Chenggang(Department of Pharmaceutical Analysis,Jining Medical Univercity,Rizhao 276826,China;Department of Traditional Chinese Medicine,Rizhao People's Hospital,Rizhao 276800,China;Department of Immunopharmacology,Jining Medical Univercity,Rizhao 276826,China;Shanghai Institute of Pharmaceutical Research,Chinese Academy of Sciences,Shanghai 201203,China)

机构地区：[1]济宁医学院药物分析教研室,日照276826 [2]日照市人民医院中医科,276800 [3]济宁医学院免疫药理学教研室,日照276826 [4]中国科学院上海药物研究所,上海201203

出　　处：《中华行为医学与脑科学杂志》2019年第8期740-745,共6页Chinese Journal of Behavioral Medicine and Brain Science

基　　金：山东省重点研发计划项目(2018GSF119006);济宁医学院青年教师科研扶持基金项目(JY2017KJ050).

摘　　要：目的探讨知母皂苷BII ( timosaponin B-II,TB-II)对大鼠来源的神经干细胞( neural stem cells,NSCs)向酪氨酸羟化酶阳性(tyrosine Hydroxylase,TH)神经元分化的影响.方法原代培养大鼠神经干细胞,采用细胞增殖计数法、形态学观察和免疫组化等方法鉴定NSCs的自我增殖能力和多向分化的生物学功能.取第三代( P3) NSCs,设立无添加 TB-II的血清自然分化组为对照组, TB-II实验组依次添加10 μg/ml、30 μg/ml和100 μg/ml ,作用7 d.采用免疫组织化学方法检测TH在各组的表达情况;Western blot法检测分化细胞TH蛋白表达的程度.结果(1)所培养细胞具备自我增殖能力,表达nestin蛋白,分化为神经元和神经胶质细胞,符合神经干细胞生物学属性.(2)与对照组相比,TB-II 30 μg/ml剂量组和TB-II 100 μg/ml剂量组 TH阳性细胞比率均升高[( 10. 03 ± 1. 36)%,(20. 01±3. 37)%,(31. 32±3. 98)%],差异有统计学意义(t=16. 54,6. 15,均P<0. 05). TB-II 10 μg/ml组与对照组比较差异无统计学意义(t=18. 36,P>0. 05).(3)Western结果显示,与对照组比较,TB-II 30 μg/ml剂量组和TB-II 100 μg/ml组的TH蛋白的相对表达量升高,差异有统计学意义[对照组:( 1. 02 ± 0. 24),TB-II 30 μg/ml 剂量组:( 3. 64 ± 1. 78),TB-II 100 μg/ml 剂量组:( 5. 88 ± 2. 34);t=12. 58,9. 15,均P<0. 05]. TB-II 10 μg/ml组与对照组比较差异无统计学意义( t=16. 42,P>0. 05).结论 TB-II可促进NSCs向TH阳性神经元分化.Objective To explore the effect of timosaponin B-II (TB-II) on the differentiation of neural stem cells (NSCs) into tyrosine hydroxylase (TH) positive neurons in neonatal rats. Methods The biological functions of self-proliferation and multi-differentiation of NSCs were identified by primary culture, cell proliferation counting, morphological observation and immunology. NSCs of SD rats were cultured in vitro and treated with different concentrations of TB-II (10 μg/ml, 30 μg/ml , 100 μg/ml) for 7 days. Immunohistochemistry was used to detect the effect of TB-II on the differentiation of NSCs into TH-positive neurons, and Western blot was used to detect the expression of TH protein in neurons. Results (1)The cultured cells had the ability to self-proliferation, expressed nestin protein and differentiated into neurons and glial cells. So the cultured cells were conformed to the biological function of neural stem cells.(2)Compared with the control group, the TH positive cell ratio of TB-II 30 μg/ml group and TB-II 100 μg/ml group increased ((10.03±1.36)%),(20.01±3.37)%),(31.32±3.98)%), the difference was significant (t=6.15, 16.54, both P<0.05). There was no significant difference between TB-II 10 μg/ml group and control group (P>0.05).(3)Western results showed that the relative expression of TH protein in TB-II 30 g/ml group and TB-II 100 μg/ml group was higher than that in control group, the difference was statistically significant (control group:(1.02±0.24), TB-II 30μg/ml group:(3.64±1.78), TB-II 100 μg/ml group:(5.88±2.34);t=12.58, 9.15, both P<0.05). There was no significant difference between TB-II 10 μg/ml group and control group (P>0.05). Conclusion TB-II can promote the differentiation of NSCs into TH-positive neurons.

关 键 词：知母皂苷BII 神经干细胞 抑郁症 酪氨酸羟化酶 大鼠 

分 类 号：R285.5[医药卫生—中药学]