Abstract



BACKGROUND: The peripheral blood stem cell is a multi-differentiation precursor cell, and it can differentiate into osteoblasts. Tissue engineered bone, which is regarded as a vector of cell transplantation, has good compatibility with receptor tissue and seed cells. Transforming growth factor-β (TGF-β) is an important regulatory factor for repairing bone injury. Additionally, TGF-β can induce peripheral blood stem cells to differentiate and proliferate into osteoblasts.
OBJECTIVE: To study TGF-β expression in repairing alveolar bone during synergetic transplantation of peripheral blood stem cells and tissue engineered bone.
DESIGN: Observational study.
SETTING: Stomatology Hospital of Xi’an Jiaotong University.
MATERIALS: This study was performed at the Stomatology Hospital of Xi’an Jiaotong University from 2003 to 2006. Experimental animals were provided by the Animal Experimental Center, Medical College of Xi’an Jiaotong University (original Xi’an Medical University). All animals were intramuscularly induced with ketamine, intramuscularly anesthetized with sumianxin, and then sacrificed for surgery. The experiment was approved by the local ethics committee.
METHODS: Peripheral blood stem cells were extracted from dog and prepared as a cell suspension. Iliac bone was obtained from healthy pig to prepare decalcifying-deproteinic tissue engineered bone. The tissue engineered bone was then dipped into peripheral blood stem cell suspension. Ten healthy hybrid dogs were randomly divided into an experimental group and a control group, with 5 dogs in each group. An incision was made from left to right along the canine teeth of the lower mandible, along the lip, lateral to the gingival sulcus, to the alveolar crest, and then along the bilateral vestibular groove to form a trapezoid segment. Subsequently, the segment was turned downward to expose the bone lamella lateral to the lip. In addition, a bone defect region of 2 cm × 2 cm × 1 cm was drilled between the lateral incisor of lower mandible using a turbine drill. Peripheral blood stem cell-tissue engineered bone was implanted in the experimental group but tissue engineered bone only was implanted in the control group. At 2, 3, 4, 8 and 12 weeks after surgery, during the differentiation and proliferation of peripheral blood stem cell into osteoblasts, TGF-β expression was measured using immunohistochemistry.
MAIN OUTCOME MEASURES: ① Morphological changes of peripheral blood stem cells differentiating into osteoblasts and structural function of organoid were observed under optical microscopy and by transmission electron microscopy. ② TGF-β expression was measured using immunohistochemistry during the differentiation and proliferation of peripheral blood stem cells into osteoblasts.
RESULTS: Two weeks after peripheral blood stem cell-tissue engineered bone transplantation in the experimental group, TGF-β expression was mildly positive at the fringe of the bone defect. Four to eight weeks after the transplantation, high numbers of osteoblasts, fibroblasts and collagenous fibers were found at the center of the bone defect region, and TGF-β expression was strongly positive. The bone defect was completely repaired after 12 weeks. In the control group, 8-12 weeks after tissue engineered bone transplantation, TGF-β expression was mildly positive only at the fringe of the bone defect.
CONCLUSION: During dog alveolar bone defect repair, TGF-β can induce peripheral blood stem cells, in combination with tissue engineered bone, to differentiate and to proliferate into osteoblasts.

Department of Oral Medicine, Stomatology Medical College, Xi’an Jiaotong University, Xi’an 710004, Shaanxi Province, China

Zhao Shu-xian, Associate professor, Associate chief physician, Department of Oral Medicine, Stomatology Medical College, Xi’an Jiaotong University, Xi’an 710004, Shaanxi Province, China
zhaoshu@mail.
xjtu.edu.cn

Supported by: the Science and Technology Foundation of Shaanxi Province, No. 2004-K16-G16*

Received:2007-08-20
Accepted:2008-02-21
(07-50-9-5143/GW)

Zhao SX, Wang M, Zhang JG, Dong K, Zhang L, Zhao DF. Transforming growth factor-beta expression in repairing dog alveolar bone defect during synergetic transplantation of peripheral blood stem cells and tissue engineered bone.Zhongguo Zuzhi Gongcheng Yanjiu yu Linchuang Kangfu 2008;12(8):
1573-1576(China)

[www.zglckf.com/
zglckf/ ejournal/
upfiles/08-8/
8k-1573(ps).pdf]

外周血干细胞与组织工程骨复合移植
过程中转化生长因子β在修复犬牙
周骨缺损的表达*

摘要
背景：外周血干细胞是一类具有多分化潜能的前体细胞,有分化为成骨的潜能,组织工程骨作为细胞移植的载体,与受体组织和种子细胞有良好的相容性。转化生长因子β是骨缺损修复的重要调节因子，有诱导外周血干细胞分化增殖为成骨细胞的作用。
目的：观察外周血干细胞与组织工程骨复合移植修复牙周骨缺损中转化生长因子β的表达。
设计：以细胞为对象的观察实验。
单位：西安交通大学口腔医院。
材料：实验于2003/2006年在西安交通大学口腔医院完成。实验动物由西安交通大学医学院（原西安医科大学）动物试验中心提供。实验动物均采用氯胺酮肌肉诱导后,肌内注射速眠新麻醉后进行手术或处死取材，对动物处置符合动物伦理学标准。
方法：抽取犬外周血干细胞，制成细胞悬液备用。取健康仔猪髂骨制作脱钙脱蛋白生物组织工程骨,浸入犬外周血干细胞细胞悬液中备用。将10 只健康杂种犬分成实验组和对照组,每组5 只。自犬下颌左右侧尖牙之间沿唇侧牙龈沟处达牙槽嵴,再转向双侧前庭沟切开,形成一个梯形瓣，向下翻瓣暴露唇侧骨板，在下颌侧切牙之间用涡轮钻制备 2 cm×2 cm×1 cm 的骨缺损区,实验组植入外周血干细胞-组织工程骨，对照组不移植外周血干细胞,仅移植组织工程骨。术后2，3，4，8，12周采用免疫组织化学方法观察外周血干细胞分化增殖成为成骨细胞过程中转化生长因子β的表达。
主要观察指标：①光镜和透射电镜观察外周血干细胞转化成为成骨细胞的形态学变化和细胞器的结构功能。②免疫组织化学方法测量外周血干细胞在转化成为成骨细胞的过程中转化生长因子β的表达。
结果：实验组在外周血干细胞-组织工程骨移植后2 周,即见骨缺损区边缘转化生长因子β呈弱阳性表达,4~8 周骨缺损区中心区域可见大量的成骨细胞、成纤维细胞和胶原纤维呈现强阳性表达,12 周骨缺损区已完全修复。对照组组织工程骨移植后8~12 周,仅在骨缺损边缘区域呈转化生长因子β弱阳性表达。
结论：当用外周血干细胞和组织工程修复犬牙周骨缺损时，转化生长因子β能诱导外周血干细胞分化和增殖为成骨细胞。
关键词：外周血干细胞; 免疫组织化学; 转化生长因子β;犬

赵淑贤,王　敏,张建国,董 凯,张 兰,赵东方
西安交通大学口腔医学院口腔内科教研室,陕西省西安市 710004
赵淑贤，女，1950年生，陕西省西安市人，汉族，1977年四川医学院毕业，副教授，副主任医师，主要从事牙周病的病因及临床治疗学研究。
陕西省科技攻关基金资助项目(2004-K16-G16)*

中图分类号: R394.2 文献标识码: A 文章编号: 1673-8225(2008)08-1573-04
赵淑贤,王敏,张建国,董凯,张兰,赵东方.外周血干细胞与组织工程骨复合移植修复犬牙周骨缺损过程中转化生长因子β的表达[J].中国组织工程研究与临床康复，2008，12(8):1573-1576
[www.zglckf.com/zglckf/ejournal/upfiles/08-8/8k-1573(ps).pdf]
(Edited by Asok Mukhopadhyay/Ji H/Wang L)