中文摘要：

目的研究甲状旁腺相关肽全长及部分肽段PTHrp（107．139）对体外培养的骨骺干细胞的调控作用并检测调控后stathmin的表达。方法利用免疫磁珠分选细胞的方法，分离出原代骨骺干细胞。转化、提取、鉴定所需的三种质粒。用脂质体介导法将质粒转染入骨骺干细胞，通过G418筛选得到抗性克隆并将其分别扩增培养，并加入1．0pg/ml的强力霉素进行诱导，并设置空白对照。转染后第4小时以及24、72、96小时对各组细胞用Western．blot检测stathmin的表达水平，并抽提各组细胞的RNA进行逆转录，然后通过RT—PCR的方法检测细胞内stathmin的mRNA表达变化，流式细胞仪检测细胞周期和凋亡。结果Tet-on基因表达系统能准确的调控骨骺干细胞中的基因表达。质粒转染后骨骺干细胞数量明显增多。PTHrp全长及部分肽段质粒转染实验组，其骨骺干细胞中stathmin的蛋白水平和mRNA水平均逐渐增加，且增高程度大致相当，空白对照组无明显变化。结论甲状旁腺相关肽及其肽段PTHrp（107-139）均可能有促进骨骺干细胞增殖的作用，stathmin在调控中出现高表达，提示细胞增殖。

英文摘要：

Objective To study the modulating effects of PTHrP and PTHrP（107-139） plamid in cultured precartilaginous stem cells in vitro and the expression level of stathmin after the transfection of the plasmid. Methods Obtain precartilaginous stem cells ofLa Croix ring of neonate SD rats and purify them. Transformation, isolation and identification of the plasmids ,the plasmids were co-transfected by lipofectamine 2000 into precartilaginous stem cells. The preeartilaginous stem cells were firstly cultured in the medium and were induced by 1.0lag/ ml Doxycycline. At the fourth, twenty-fourth, seventy-second and ninty-sixth hour after being transferred, the RNA of the cells of each group were extracted and then were examined by RT-PCR to investigate the changes of expression of mRNA .Acording to the experiment, the cells were examined with Western blot to detect the expression/eve/ofstathmin in the transfected cells at the designed shedule. Then the cell cycle and Apoptosis was examined by flow cytometry. Results The objective gene could be modulated precisely by the control of pTet-on gene aiding system in the precartilaginous stem cells. The quantitive level of precartilaginous stem cells increased significantly after the transfection of the plasmid of PTHrP. In the subject group of PTHrP（107-139） and PTHrp,the increasing quantitive level of the protein and mRNA of stathmin in the cells were detected, meanwhile, there were no siginificant change of the cells in the control group. Conclusion Parathyroid hormone-related protein and its functional fragment like PTHrP（107-139） could effectively modulate the proliferation of precartilaginous stem cells and the auantitive level of stathmin would increase following, the oroliferation ofnreeartilac, inous stem cells.