应用要点：本实验以体外扩增培养的髂骨生长板软骨细胞为种子细胞，结合同种异体脱钙骨基质体外构建组织工程生长板，观察到细胞和材料有良好的生物相容性，生长板软骨细胞可于材料增殖、分泌基质，为自体髂骨生长板细胞和组织工程方法应用于长骨生长板损伤治疗作出了有益的探索。

同行评价：骨骺板是未成年人很重要的结构，损伤后导致生长受阻，组织工程的兴起有望通过组织工程的方法来解决这一难题。作者初步构建了组织工程化骨骺板，为解决这一难题作了有益的探索，具有一定创新性。文章如果能跟正常骨骺板进行对照会更好，也希望作者能进一步深入研究并进入动物试验研究，检测组织工程骨骺板的治疗效果。

相关链接：生长板损伤修复是小儿矫形外科实验研究和临床治疗面临的一大难题。游离生长板软骨移植，因缺乏血供而应用困难；血管化生长板移植解决了血循环问题，但是自体移植供体来源有限；异体移植存在免疫排斥；非软骨组织材料只能阻止小面积生长板缺损内骨桥形成，无再生修复能力。软骨细胞及组织工程软骨移植可能是生长板损伤修复的最佳选择。

摘要
目的：由创伤、感染等引起的长骨生长板损伤会导致肢体的短缩与成角畸形,组织工程学科的兴起为治疗生长板损伤提供了契机。采用组织工程技术，旨在探索兔髂骨生长板细胞和同种异体脱钙骨基质共培养构建组织工程生长板的可行性。　
方法：实验于2005-06/2006-06在解放军第四军医大学西京医院全军骨科研究所完成。①实验材料：清洁级3周龄新西兰白兔2只，雌雄不限，体质量2.0~2.5 kg。②实验过程：自兔髂嵴处切取部分髂骨生长板软骨经机械剪切和Ⅱ型胶原酶消化后培养生长板软骨细胞，收集传代培养的第3代兔髂骨生长板细胞体外扩增后接种到同种异体脱钙骨基质上，混合培养。③实验评估：于联合培养24 h，第7，14，21天进行扫描电镜、组织学、免疫组化染色，观察细胞在材料中的生长情况。
结果：①体外单层培养的兔髂骨生长板细胞呈多角形，Ⅱ型胶原免疫组化染色阳性。②扫描电镜检查显示体外复合培养24 h,软骨细胞即开始贴附于脱钙骨基质网架上；第7天分布于支架材料上的生长板细胞迅速分化增殖,分泌细胞外基质；第21天细胞长满支架，重叠生长。③苏木精-伊红染色示，复合培养14 d后，生长板细胞很好地贴附于脱钙骨基质网架上，细胞多为球形，胞浆丰富。
结论：同种异体脱钙骨基质和髂骨生长板细胞共同培养可成功构建出组织工程生长板。
关键词：组织工程；生长板；脱钙骨基质；细胞培养；组织构建

王德元，高文魁，李智钢，邓永忠. 兔髂骨生长板细胞和脱钙骨基质共培养构建组织工程生长板[J].中国组织工程研究与临床康复，2008，12(2):227-230 [www.zglckf.com/zglckf/ejournal/upfiles/08-2/2k-227(ps).pdf]

Constructing tissue-engineered growth plate by allograft demineralized bone matrix cocultured with rabbit iliac growth-plate cells

Abstract

AIM： Long bone growth plate injury induced by wound and infection may cause limb reduction or angular deformity. Tissue engineering provides a promising treatment of growth plate injuries. In this study, we investigated the feasibility of establishing tissue-engineered growth-plate by allograft demineralized bone matrix (DBM) co-cultured with rabbit iliac growth-plate cells.
METHODS: The experiment was performed at Department of Orthopaedics, Fourth Military Medical University of Chinese PLA from June 2005 to June 2006. ①Two 3-week-old New Zealand rabbits irrespective of gender (clean grade, 2.0-2.5 kg) were selected. Growth plate cells were harvested from iliac crest epiphyseal cartilage of the rabbits by dissection and digestion with type Ⅱ collagenase. The third passage cells cultured were collected and incubated on allograft demineralized bone matrix. ② Histology, immunohistochemical staining and electronic scanning microscope (SEM) examinations were performed to observe cell growth on DBM 24 hours, 7, 14 and 21 days after culture.
RESULTS: ①Growth plate chondrocytes exhibited polygonal in monolayer culture, and immunohistochemical staining for type Ⅱ collagen was positive. ②SEM examination showed that twenty-four hours after coculture, the cells adhered to DBM scaffolds; Seven days after culture, the growth-plate chondrocytes rapidly proliferated and began to secret extracellular matrix; cells covered the whole scaffold and became overlapped on the 21st day. ③HE staining showed after 14 days of culture, growth plate cells adhered DBM scaffold in spherical shape with abundant cytoplasm.
CONCLUSION: Tissue-engineered growth-plate is successfully constructed by allograft mineralized bone matrix co-cultured with rabbit iliac growth-plate cells.

Wang DY, Gao WK, Li ZG, Deng YZ. Constructing tissue-engineered growth plate by allograft demineralized bone matrix cocultured with rabbit iliac growth-plate cells.Zhongguo Zuzhi Gongcheng Yanjiu yu Linchuang Kangfu 2008;12(2):227-230(China) [www.zglckf.com/zglckf/ejournal/upfiles/08-2/2k-227(ps).pdf]