STRO-1 selected rat dental pulp stem cells transfected with adenoviral-mediated human bone morphogenetic protein 2 gene show enhanced odontogenic differentiation.

Xuechao Yang, Peter M van der Kraan, Juliette van den Dolder, X Frank Walboomers, Zhuan Bian, Mingwen Fan, John A Jansen
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引用次数: 68

Abstract

Dental pulp stem cells harbor great potential for tissue-engineering purposes. However, previous studies have shown variable results, and some have reported only limited osteogenic and odontogenic potential.Because bone morphogenetic proteins (BMPs) are well-established agents to induce bone and dentin formation,in this study STRO-1-selected rat dental pulp-derived stem cells were transfected with the adenoviral mediated human BMP-2 gene. Subsequently, the cells were evaluated for their odontogenic differentiation ability in medium not containing dexamethasone or other stimuli. Cultures were investigated using light microscopy and scanning electron microscopy (SEM) and evaluated for cell proliferation, alkaline phosphatase(ALP) activity, and calcium content. Real-time polymerase chain reaction (PCR) was performed for gene expression of Alp, osteocalcin, collagen type I, bone sialoprotein, dentin sialophosphoprotein, and dentin matrix acidic phosphoprotein 1. Finally, an oligo-microarray was used to profile the expression of odontogenesis-related genes. Results of ALP activity, calcium content, and real-time PCR showed that only BMP2-transfected cells had the ability to differentiate into the odontoblast phenotype and to produce a calcified extracellular matrix. SEM and oligo-microarray confirmed these results. In contrast, the non-transfected cells represented a less differentiated cell phenotype. Based on our results, we concluded that the adenovirus can transfect STRO-1 selected cells with high efficacy. After BMP2 gene transfection, these cells had the ability to differentiate into odontoblast phenotype, even without the addition of odontogenic supplements to the medium.

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经STRO-1筛选的大鼠牙髓干细胞转染腺病毒介导的人骨形态发生蛋白2基因后,其成牙分化增强。
牙髓干细胞在组织工程方面具有巨大的潜力。然而,先前的研究显示了不同的结果,有些研究仅报道了有限的成骨和牙源性潜力。由于骨形态发生蛋白(BMPs)是一种公认的诱导骨和牙本质形成的物质,因此本研究将stro -1选择的大鼠牙髓源性干细胞转染腺病毒介导的人BMP-2基因。随后,在不含地塞米松或其他刺激的培养基中评估细胞的成牙分化能力。利用光镜和扫描电镜(SEM)研究培养物,并评估细胞增殖、碱性磷酸酶(ALP)活性和钙含量。采用实时聚合酶链反应(Real-time polymerase chain reaction, PCR)检测Alp、骨钙素、I型胶原、骨唾液蛋白、牙本质唾液磷酸蛋白、牙本质基质酸性磷酸蛋白1的基因表达。最后,利用寡核苷酸芯片分析牙形成相关基因的表达。ALP活性、钙含量和实时PCR结果显示,只有转染bmp2的细胞能够分化成成牙髓细胞表型并产生钙化的细胞外基质。SEM和oligo-microarray证实了这些结果。相反,未转染的细胞表现为分化程度较低的细胞表型。基于我们的结果,我们得出结论,腺病毒可以高效地转染STRO-1选择的细胞。在BMP2基因转染后,即使培养基中没有添加成牙源性补充剂,这些细胞也有能力分化成成牙细胞表型。
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来源期刊
Tissue engineering
Tissue engineering CELL & TISSUE ENGINEERING-BIOTECHNOLOGY & APPLIED MICROBIOLOGY
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