软骨寡聚基质蛋白对骨形态发生蛋白-2诱导的间充质干细胞分化的影响
Effects of cartilage oligomeric matrix protein on bone morphogenetic protein-2-induced differentiation of mesenchymal stem cells.
作者信息
Guo Peng, Shi Zhong-li, Liu An, Lin Tiao, Bi Fanggang, Shi Mingmin, Yan Shi-Gui
机构信息
Department of Orthopaedic Surgery, The Second Affiliated Hospital, Medical College of Zhejiang University, Hangzhou, China.
出版信息
Orthop Surg. 2014 Nov;6(4):280-7. doi: 10.1111/os.12135.
OBJECTIVE
To investigate the effect of overexpression of cartilage oligomeric matrix protein (COMP) on bone morphogenetic protein-2 (BMP-2) induced osteogenic and chondrogenic differentiation of mesenchymal stem cells (MSCs). In this study, we used liposomes to transfect MSCs with plasmid encoding COMP and then induced the transfected MSCs to differentiate in osteogenic and chondrogenic differentiation media containing BMP-2.
METHODS
MSCs transfected with plasmid DNA encoding recombinant human COMP were induced to differentiate into osteocytes and chondrocytes by BMP-2. Real-time polymerase chain reaction (PCR) assays of osteogenesis-related markers (collagen type I alpha 1, runt-related transcription factor 2, osteopontin, bone gla protein) and chondrogenesis-related markers (collagen type II alpha 1, sry-related high-mobility group box 9, Aggrecan) was performed to evaluate the process of cell differentiation. Cell differentiation was evaluated by alkaline phosphatase (ALP) and Alizarin red S stains for osteogenic differentiation and alcian blue staining for chondrogenic differentiation.
RESULTS
Real-time PCR assay showed significantly greater COMP expression by MSCs when COMP gene had been transfected into the cells (P < 0.01). Overexpression of COMP down-regulated expression of osteogenesis-related markers and up-regulated expression of chondrogenesis-related markers. ALP staining and Alizarin red S staining were weakened, whereas alcian blue staining was enhanced.
CONCLUSION
Overexpression of COMP inhibits BMP-2-induced osteogenic differentiation and promotes BMP-2-induced chondrogenic differentiation. These findings may provide new insights for cartilage tissue engineering. The experiments in the present study were all in vitro, which has potential limitations. Further in vivo studies to investigate the effects of COMP in animal models are necessary, which will be the next step in our research.
目的
研究软骨寡聚基质蛋白（COMP）过表达对骨形态发生蛋白-2（BMP-2）诱导的间充质干细胞（MSC）成骨及成软骨分化的影响。在本研究中，我们使用脂质体将编码COMP的质粒转染至MSC，然后在含BMP-2的成骨及成软骨分化培养基中诱导转染后的MSC分化。
方法
用编码重组人COMP的质粒DNA转染MSC，然后用BMP-2诱导其分化为骨细胞和软骨细胞。通过实时聚合酶链反应（PCR）检测成骨相关标志物（I型胶原蛋白α1、 runt相关转录因子2、骨桥蛋白、骨钙蛋白）和成软骨相关标志物（II型胶原蛋白α1、 sry相关高迁移率族蛋白9、聚集蛋白聚糖），以评估细胞分化过程。通过碱性磷酸酶（ALP）和茜素红S染色评估成骨分化，通过阿尔新蓝染色评估成软骨分化。
结果
实时PCR检测显示，当COMP基因转染至细胞后，MSC中COMP表达显著增加（P < 0.01）。COMP过表达下调了成骨相关标志物表达，上调了成软骨相关标志物表达。ALP染色和茜素红S染色减弱，而阿尔新蓝染色增强。
结论
COMP过表达抑制BMP-2诱导的成骨分化，促进BMP-2诱导的成软骨分化。这些发现可能为软骨组织工程提供新的见解。本研究中的实验均为体外实验，存在潜在局限性。有必要进一步开展体内研究以探究COMP在动物模型中的作用，这将是我们下一步的研究方向。
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