重组甲状旁腺激素相关蛋白1-36及其在促进骨原细胞分化中的作用
陈玲1，孙雯2
基金项目：高等学校博士学科点专项科研基金-新教师类（20123234120007）；国家自然科学基金（81200755）；江苏省自然科学基金（BK2012440） 作者简介：陈玲（1970年出生），女，副主任医师，主要研究方向：分子生物学 通信联系人：孙雯（1983年出生），女，讲师，主要研究方向：分子生物学.

（1. 南京医科大学附属江宁医院 口腔科，南京 211100；2. 南京医科大学基础医学院 人体解剖教研室，南京 210029）
摘要：目的：构建重组甲状旁腺激素相关蛋白(Parathyroid hormone related protein, PTHrP)1-36并观察其在促进骨原细胞分化中的作用。方法：通过构建pGEX-2TK/ PTHrP1-36原核表达载体，表达重组小鼠PTHrP1-36蛋白，并通过处理小鼠原代骨髓间充质细胞，观察重组PTHrP1-36在促进骨原细胞分化中的作用。结果：实验结果显示，pGEX-2TK/ PTHrP1-36原核表达载体构建成功，IPTG可以诱导新生蛋白表达。通过IPTG诱导液的浓度和时间优化，确定1mM IPTG诱导3小时为最佳诱导条件，并利用此条件进行大量蛋白表达。通过处理小鼠骨髓细胞，发现重组PTHrP1-36蛋白可以增加总成纤维细胞克隆形成单位(CFU-f)、碱性磷酸酶阳性的CFU-f和钙化结节的数目。 结论：重组PTHrP1-36可以促进骨原细胞的分化，提示PTHrP1-36在促进骨形成中的潜在作用。
关键词：甲状旁腺激素相关蛋白；骨原细胞；重组蛋白
中图分类号：Q78

Synthetic PTHrP1-36 and its role for promoting osteoprogenitor commitment
CHEN Ling1, SUN Wen2
(1. Department of Stomatology, Jiangning Affiliated Hospital of Nanjing Medical University, Nanjing, 211100; 25 2. Department of Anatomy, Histology and Embryology, Nanjing Medical University, Nanjing, 210029)
Abstract: Objective: To synthetise mouse Parathyroid hormone related protein 1-36 (PTHrP1-36) and study its role for promoting osteoprogenitor commitment. Method: pGEX-2TK/ PTHrP1-36 was constructed and PTHrP1-36 was expressed. The role of 30 PTHrP1-36 for osteoprogenitor commitment was observed by treating bone marrow cells.Result: The results of PCR analysis showed that the pGEX-2TK/ PTHrP1-36 plasmid was correct templates for expressing PTHrP1-36. The molecular weights obtained by SDS-PAGE were the same as the theoretical value after IPTG induction. Scale up production was carried out by 3-hour-induction with 1mM IPTG which is the optimal induction factors. Cell culture revealed that Synthetic PTHrP1-36 can increase the number of total colony forming units fibroblastic (CFU-f), ALP-positive CFU-f and calcific nodules. Conclusion: Synthetic PTHrP1-36 can promote osteoprogenitor commitment which points out its role for enhancing bone formation.
Key words: parathyroid hormone related protein; osteoprogenitor; synthetic protein