从JAK/STAT信号转导通路探讨益肾化浊方对神经干细胞定向分化的影响  

Effects of Yishen Huazhuo Fang( 益肾化浊方) on Directional Differentiation of Neural Stem Cells Based on JAK / STAT Signal Transduction Pathway

作　　者：崔远武[1] 周震[1] 张琳琳[1] 宋宛珊[2] 韩文文[1] 王凯[2] 张玉莲[1] 

机构地区：[1]天津中医药大学第二附属医院,天津市河北区真理道816号300150 [2]天津中医药大学

出　　处：《中医杂志》2016年第6期512-515,共4页Journal of Traditional Chinese Medicine

基　　金：国家自然科学基金项目(81102577);国家重点基础研究发展计划(2010CB530405)

摘　　要：目的探讨益肾化浊方治疗阿尔茨海默病的可能作用机制。方法从孕14天昆明小鼠体外提取并培养神经干细胞,建立分化系统中可溶性Aβ25-35(5μmol/L)介导的具有神经毒性的阿尔茨海默病细胞模型。实验分为对照组(等量培养基)、模型组(5μmol/L Aβ25-35100μl)和益肾化浊中药组(5μmol/L Aβ25-35100μl+0.1μg/ml益肾化浊方100μl)。常规消化细胞,计数后以细胞密度每毫升2×105/个(每孔2ml)接种于6孔板,培养48 h。Western Blot法检测神经胶质酸性蛋白(GFAP)、微管蛋白(Tubulin)和转录激活子3(STAT3)、磷酸化转录激活子3(P-STAT3)、Smad1蛋白表达;RT-PCR法检测GFAP、STAT3、Smad1、Tubulin基因的表达。结果与对照组比较,模型组GFAP、STAT3基因及蛋白表达显著增加、PSTAT3蛋白表达显著增加,Tubulin基因及蛋白表达显著减少(P<0.05),Smad1基因及蛋白表达增加不明显(P>0.05);与模型组比较,益肾化浊中药组GFAP、STAT3、Smad1基因及蛋白表达均显著下降、PSTAT3蛋白表达亦下降,而Tubulin基因及蛋白表达显著增加(P<0.05)。结论益肾化浊方可能通过调控JAK/STAT信号通路中相关蛋白及基因表达,从而抑制神经干细胞向星形胶质细胞方向分化来延缓阿尔茨海默病发展。Objective To study possible mechanism of Yishen Huazhuo Fang（ 益肾化浊方） to treat Alzheimer＇s disease. Methods The researchers extracted and cultured neural stem cells in vitro from Kunming mice with 14-day pregnancy. Then the researchers established neurovirulent cell models of Alzheimer＇s disease mediated by soluble Aβ25- 35（ 5 μmol/L） in differentiation system. Samples were divided into the control group（ equivalent medium）,the model group（ Aβ25- 35,100 μl,5 μmol/L） and the Chinese medicine of Yishen Huazhuo group（ Aβ25- 35,100 μl,5 μmol / L ＋ Yishen Huazhuo Fang 100 μl,0. 1 μg / ml）. After conventional digesting those cells and counting,the researchers inoculated them in 6-hole plates according to cell density 2 × 105/ ml（ 2 ml in per hole） and cultured for 48 h. The researchers used western blot method to detect expression of glial fibrillary acidic protein（ GFAP）,microtubulin（ Tubulin） and transcriptional activator 3（ STAT3）,phosphorylated transcriptional activator 3（ P-STAT3） and Smad1 protein. They also used RT-PCR method to detect gene expression of GFAP,STAT3,Smad1 and Tubulin. Results Compared with the control group,expression of GFAP,STAT3 gene and protein and P-STAT3 protein in the model group significantly increased. However expression of Tubulin gene and protein significantly decreased（ P〈0. 05）. Expression of Smad1 gene and protein unconspicuously increased（ P〈0. 05）.Compared with the model group,expression of GFAP,STAT3,Smad1 gene and protein and P-STAT3 protein in the Chinese medicine of Yishen Huazhuo group significantly decreased. However expression of Tubulin gene and protein significantly increased（ P〈0. 05）. Conclusion Yishen Huazhuo Fang may restrain neural stem cells from differentiation towards astrocytes through regulating expression of related proteins and gene in the JAK / STAT signaling pathway to delay the development of Alzheimer＇s disease.

关 键 词：阿尔茨海默病 益肾化浊方 Β淀粉样肽 细胞定向分化 神经干细胞 星形胶质细胞 

分 类 号：R285[医药卫生—中药学]