人骨形态发生蛋白质7基因转染兔骨髓基质干细胞向成骨细胞的分化

人骨形态发生蛋白质7基因转染兔骨髓基质干细胞向成骨细
胞的分化
康焱;廖威明;盛蹼义;张珑涓
【期刊名称】《中国组织工程研究》
【年(卷),期】2006(010)021
【摘要】BACKGROUND: At present, bone morphogenetic protein (BMP) is the only cytokine characterized as the ability of osteoblast, and it can promote marrow mesenchymal stem cells (MSCs) differentiating into chondroblast and osteoblast.OBJECTIVE: To investigate the differentiation into osteoblast from MSCs of rabbits transfected by human bone morphogenetic protein-7 (hBMP-7) gene with genetic engineering technique. DESIGN: Single sample observation. SETTING: Department of Orthopaedics, the First Hospital Affiliated to Sun Yat-sen University. MATERIALS: pcDNA1.1/AMP-hBMP7 plasmid.METHODS: The experiment was completed at the Surgery Laboratory of the First Hospital Affiliated to Sun Yat-sen University from May 2004 to March 2005. MSCs of rabbits were cultured with whole bone marrow technique, transfected with pcDNA1.1/AMP-hBMP7 and pcDNA1.1/AMP plasmids respectively in vitro, and left the blank controls. Transcription and expression of transfected genes were detected so as to observe form and growth of cells. Component of calcium content in cytoplasm was measured and osteogenic phenotype was identified with alkaline phosphatase (ALP)
staining, calcium salts staining and transmission electron microscopy (TEM).MAIN OUTCOME MEASURES: ① Form and growth of MSCs of rabbits; ② evaluation of expressive product; ③ results of ALP staining (Ca-Co technique); ④ results of chinalizarin staining; ⑤ results of TEM; ⑥ results of scanning electron m icroscopy (SEM); ⑦ assay of osteocalcin.RESULTS: hBMP-7 gene was contained in MSCs after transfection and expressed the relevant mRNA. Cellular form changed a little after expression of amid genes, but growth curve did not changed as compared with that in non-transfected group. Expression of osteocalcin was increased as compared with that in non-transfected group, and ALP staining and calcium salts staining of transfected cells were
positive.CONCLUSION: Transfection of hBMP-7 gene can promote MSCs of rabbits differentiating into osteoblast.%背景:骨形态发生蛋白质是目前发现惟一的一类具有成骨能力的细胞因子,它能刺激基质干细胞向成软骨细胞和成骨细胞方向分化.目的:应用基因工程方法,观察人骨形态发生蛋白质7基因转染兔骨髓基质干细胞向成骨细胞的定向分化.设计:单一样本观察.单位:中山大学附属第一医院骨科. 材料:pcDNA1.1/AMP-hBMP7质粒.方法:实验于2004-05/2005-03在中山大学附属第一医院外科实验室完成.全骨髓培养法培养兔骨髓基质干细胞,在体外分别转染pcD-NA1.1/AMP-hBMP7和pcDNA1.1/AMP并留置空白对照.检测转染基因的转录和表达,观察细胞形态和生长情况,通过对感染细胞的碱性磷酸酶染色、钙盐染色、电镜观察胞质中钙质成分以及检测骨钙素鉴定其成骨分化表型.主要观察指标:①兔骨髓基质干细胞的形态及生长.②表达产物的鉴定.③碱性磷酸酶染色(钙钴法)结果.④茜素红染色结果.⑤透射电镜观察结果.⑥扫描电镜观察结果.⑦骨钙素的测定结果.结果:人骨形态发生蛋白质7基因存在于转染后的骨髓基质干细胞中并表
达相应的mRNA.目的基因表达后的细胞形态略有变化,但生长曲线与未转染组无明显变化.骨钙素表达较未转染组明显增强.转染组细胞的碱性磷酸酶染色和钙盐染色均为阳性.结论:人骨形态发生蛋白质7基因转染可促进兔骨髓基质干细胞向成骨细胞表型分化.
【总页数】4页(P161-163,封三)
【作者】康焱;廖威明;盛蹼义;张珑涓
【作者单位】中山大学附属第一医院骨科,广东省广州市,510080;中山大学附属第一医院骨科,广东省广州市,510080;中山大学附属第一医院骨科,广东省广州
市,510080;中山大学附属第一医院骨科,广东省广州市,510080
【正文语种】中文
【中图分类】R3
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