Zhang Wen, Liu Wei, Ling Junqi, Lin Zhengmei, Gao Yan, Mao Xueli, Jian Yutao
Department of Operative Dentistry and Endodontics, Guanghua School and Hospital of Stomatology and Guangdong Province Key Laboratory of Stomatology, Sun Yat‑sen University, Guangzhou, Guangdong 510055, P.R. China.
Mol Med Rep. 2014 Oct;10(4):1899-906. doi: 10.3892/mmr.2014.2481. Epub 2014 Aug 11.
Dental pulp stem cells (DPSCs) can be induced towards odontogenic differentiation. Previous studies have shown that vascular endothelial growth factor (VEGF) is able to induce the osteogenic differentiation of cells, but the effectiveness of VEGF in the odontogenic differentiation of DPSCs remains unclear. This study aimed to investigate the effects of lentivirus‑mediated human VEGF gene transfection on the proliferation and odontogenic differentiation of human DPSCs in vitro. DPSCs were transfected with either lentiviral pCDH‑CMV‑MCS‑EFI‑copGFP (pCDH) vector or recombinant pCDH‑VEGF vector, and the growth characteristics of the resulting DPSCs/Vector and DPSCs/VEGF were subsequently assessed. The odontogenic differentiation genes of the two groups of cells, including alkaline phosphatase, osteocalcin, dentin sialophosphoprotein and dentin matrix protein 1 (DMP1), were evaluated by quantitative polymerase chain reaction (qPCR). The specific proteins of odontogenic differentiation, including dentin sialoprotein and DMP1, were analyzed by western blotting. DPSCs/VEGF showed similar proliferation characteristics to DPSCs/Vector during the observation period. qPCR results showed that the relative VEGF gene expression was significantly higher in DPSCs/VEGF than that in DPSCs/Vector two days after transfection (P<0.01). Similarly, western blot analysis showed that the protein expression levels of VEGF were higher in DPSCs/VEGF than those in DPSCs/Vector. On the first, fourth, eighth and 16th days after lentivirus-mediated transfection, the expression of odontogenic differentiation-specific genes and proteins was higher in DPSCs/VEGF than that in DPSCs/Vector. These results indicated that lentivirus-mediated VEGF gene transfection promoted the odontogenic differentiation of human DPSCs in vitro.
牙髓干细胞(DPSCs)可被诱导向牙源性分化。先前的研究表明,血管内皮生长因子(VEGF)能够诱导细胞的成骨分化,但VEGF在DPSCs牙源性分化中的有效性仍不清楚。本研究旨在探讨慢病毒介导的人VEGF基因转染对人DPSCs体外增殖和牙源性分化的影响。用慢病毒pCDH-CMV-MCS-EFI-copGFP(pCDH)载体或重组pCDH-VEGF载体转染DPSCs,随后评估所得DPSCs/载体和DPSCs/VEGF的生长特性。通过定量聚合酶链反应(qPCR)评估两组细胞的牙源性分化基因,包括碱性磷酸酶、骨钙素、牙本质涎磷蛋白和牙本质基质蛋白1(DMP1)。通过蛋白质免疫印迹法分析牙源性分化的特异性蛋白,包括牙本质涎蛋白和DMP1。在观察期内,DPSCs/VEGF与DPSCs/载体表现出相似的增殖特性。qPCR结果显示,转染后两天,DPSCs/VEGF中VEGF基因的相对表达明显高于DPSCs/载体(P<0.01)。同样,蛋白质免疫印迹分析显示,DPSCs/VEGF中VEGF的蛋白表达水平高于DPSCs/载体。在慢病毒介导的转染后的第1、4、8和16天,DPSCs/VEGF中牙源性分化特异性基因和蛋白的表达高于DPSCs/载体。这些结果表明,慢病毒介导的VEGF基因转染促进了人DPSCs的体外牙源性分化。