红景天多糖对精原干细胞体外增殖的影响  

Rhodiola polysaccharide effect on spermatogonial stem cell proliferation in vitro

作　　者：李俊涛[1] 张培海[2] 曲晓伟[3] 李郑生[1] 李松伟[4] 

机构地区：[1]河南省中医学院第一附属医院男科,河南省郑州市450000 [2]四川省中西医结合医院男科,四川省成都市610000 [3]郑州人民医院泌尿科,河南省郑州市450000 [4]河南省中医学院第一附属医院风湿科,河南省郑州市450000

出　　处：《中国组织工程研究》2016年第10期1501-1507,共7页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金面上项目(81573952)~~

摘　　要：背景:体外建立快速、有效的培养方法以获得满足临床治疗应用的精原干细胞,是进行精原干细胞移植有待解决的主要问题。目的:探讨红景天多糖对精原干细胞体外增殖的影响。方法:无菌条件下从小鼠睾丸组织中分离精原干细胞和Sertoli细胞,将精原干细胞接种于Sertoli细胞饲养层上共培养后分为3组,分别为实验组1(单纯细胞培养基),实验组2(细胞培养基+150 mg/L红景天多糖)和实验组3(细胞培养基+150 mg/L红景天多糖+1 U/L白血病抑制因子+10μg/L胶质细胞源神经营养因子)。共培养7 d后,采用流式细胞术检测细胞体外增殖状态,计算细胞活性率以及精原干细胞已知抗原标志物GFRa-1、Thy-1和C-kit的阳性表达率。结果与结论:(1)共培养7 d后,细胞生长迅速,细胞生长形态呈集落样和葡萄串样,细胞团数量增加明显,符合精原干细胞的增殖特征。(2)精原干细胞中的GFRa-1、Thy-1和C-kit蛋白均表达于细胞膜和细胞质中,主要表达于细胞膜中。(3)精原干细胞活性率、GFRa-1阳性率、Thy-1阳性率:实验组3>实验组2>实验组1,组间两两比较,差异均有显著性意义(P<0.05)。(4)C-kit阳性率:实验组1与实验组2之间相比较,差异无显著性意义(P>0.05),实验组3高于实验组1及实验组2,差异均有显著性意义(P<0.05)。(5)结果表明,红景天多糖单独或与白血病抑制因子、胶质细胞源神经营养因子联合应用,能增强精原干细胞体外增殖能力。BACKGROUND： To establish a rapid and effective method to obtain sufficient spermatogonial stem cells that can meet the clinical need is urgent to be solved in the spermatogonial stem cell transplantation. OBJECTIVE： To study the effect of rhodiola polysaccharide on the proliferation of spermatogonial stem cells in vitro. METHODS： Under sterile conditions, spermatogonial stem cells and Sertoli cells were isolated from the testis of mice, and spermatogonial stem cells were seeded onto the feed layer of Sertoli cells. Then, the co-cultured cells were assigned into experimental group 1（simple cell culture medium）, experimental group 2（cell culture medium containing 150 mg/L rhodiola polysaccharide） and experimental group 3（cell culture medium containing 150 mg/L rhodiola polysaccharide, 1 U/L leukemia inhibitory factor and 10 μg/L glial cell line-derived neurotrophic factor）. After 7 days of co-culture, flow cytometry was used to detect cell proliferation in vitro, and cell viability and positive expression of GFRa-1, Thy-1 and C-kit were calculated. RESULTS AND CONCLUSION： After 7 days of co-culture, the cells grew rapidly and presented with colony and thyrsiform growth, and the number of cell masses increased significantly, all of which were in line with the proliferative features of spermatogonial stem cells. The GFRa-1, Thy-1 and C-kit proteins were expressed in the cell membrane and cytoplasm, mainly in the cell membrane. The viability of spermatogonial stem cells and positive expression of GFRa-1 and Thy-1 were ranked as follows： experimental group 3 experimental group 2 experimental group 1, and there were significant differences between groups（P〈0.05）. The positive expression of C-kit had no difference between experimental groups 1 and 2, but it was significantly higher in the experimental group 3 than the other two groups（P〈0.05）. These findings indicate that rhodiola polysaccharide used alone or combined with leukemia inhibitory factor and glial cell line-derived neurotrop

关 键 词：干细胞 培养 红景天多糖 精原干细胞 Sertoli细胞饲养层 细胞增殖 国家自然科学基金 

分 类 号：R394.2[医药卫生—医学遗传学]