巴戟天多糖和淫羊藿苷促进视磺酸诱导的P19细胞增殖和分化  

Morinda officinalis polysauharide and Icariin promote the Proliferation and differentiation of P19 Cells induced by retionic acid

作　　者：牛长敏 夏蒙蒙 钟亚楠 徐文华 郑英[1] NIU Chang-min;XIA Meng-meng;ZHONG Ya-nan;XU Wen-hua;ZHENG Ying(Yangzhou University Medical College,Jiangsu 225000,China)

机构地区：[1]扬州大学医学院,江苏扬州225000

出　　处：《时珍国医国药》2020年第12期2881-2886,共6页Lishizhen Medicine and Materia Medica Research

基　　金：国家自然科学基金(81871205)。

摘　　要：目的探讨巴戟天多糖和淫羊藿苷(ICA)对视磺酸(RA)诱导P19细胞向生精细胞方向分化的影响。方法 MTT法分析巴戟天多糖和ICA对RA诱导的P19细胞增殖的影响,明确两者的最佳作用浓度及作用时间;流式细胞术检测巴戟天多糖和ICA对RA诱导的P19细胞凋亡的影响;实时定量PCR分析两者对RA诱导P19细胞向生精细胞方向分化进程的影响;Western Blot从蛋白水平进一步进行验证。结果终浓度4μmol/L的RA可诱导P19向生精细胞方向分化,表达生精细胞特异性未分化精原细胞标志物Plzf和精原细胞分化减数分裂启动标志物Stra8;MTT检测显示巴戟天多糖和ICA可以促进RA诱导的P19细胞增殖,最佳作用浓度分别为0.5 mg·L^(-1)和7.5μmol/L;Annexin V-FITC/PI标记后流式细胞仪检测细胞凋亡水平,未见巴戟天多糖和ICA对RA诱导的P19细胞凋亡有显著影响;转录和蛋白水平分析显示巴戟天多糖和ICA可以促进RA诱导P19细胞向生精细胞分化进程,胚胎干细胞(ES)标志物Oct4表达显著降低,Plzf和Stra8表达显著增高后下降;但是不添加RA时,巴戟天多糖和ICA不能直接诱导P19细胞向生精细胞方向分化。结论巴戟天多糖和ICA可以促进RA诱导的P19细胞增殖和向生精细胞方向分化的进程。Objective To investigate the effects of Morinda officinalis polysaccharide and Icariin(ICA) on the differentiation of P19 cells into germ cells induced by retinoic acid(RA).Methods MTT assay was performed to determine the effect of Morinda officinalis polysaccharide and ICA on P19 cell proliferation induced by RA, optimal concentration and time of action;flow cytometry was used to detect the effect of Morinda officinalis polysaccharide and ICA on P19 cell apoptosis induced by RA;real-time quantitative PCR was used to analyze the effect of them on RA induced differentiation of P19 cells into germ cells;western blot was carried out to further verify the protein expression levels.Results RA with a final concentration of 4 μmol/L could induce P19 cells to differentiate into germ cells, which expressed the undifferentiated spermatogonial marker Plz, spermatogonial differentiation and meiosis initiation marker Stra8. MTT assay showed that Morinda officinalis polysaccharide and ICA could promote the proliferation of P19 cells induced by RA, and the most appropriate concentrations were 0.5 mg·L^(-1) and 7.5μmol/L, respectively. Apoptosis rate was detected by flow cytometry after annexin V-FITC/PI labeling, we found no significant effect of Morinda officinalis polysaccharide and ICA on RA induced P19 cells. Transcriptional and protein level analysis showed that Morinda officinalis polysaccharide and ICA could promote the process of RA induced P19 to differentiate into germ cells, which showed that the expression level of Oct4, an embryonic stem cell(ES) marker, decreased significantly, the expression levels of Plzf and Stra8 increased significantly. However, they could not directly induce the differentiation of P19 cells into germ cells without RA. Conclusion Morinda officinalis polysaccharide and ICA could promote the proliferation and differentiation of P19 cells into germ cells induced by RA.

关 键 词：巴戟天多糖 ICA RA P19 细胞增殖 分化 

分 类 号：R285.5[医药卫生—中药学]