“非病毒基因活化基质”转染ephrinB2促进牙髓血运重建术的实验研究

如何使根管内新生组织快速地形成血管网，并促进根管内干细胞分化为成牙本质细胞，是决定牙髓血运重建术成败的关键。我们前期实验证实：体外转染ephrinB2的牙源性干细胞具备更强的成骨能力，并能促进周围的血管内皮细胞更快地形成血管网。但体外培养干细胞存在细胞来源困难和污染的可能性，且在转染过程中常用的病毒载体也对患者构成潜在的风险。“非病毒基因活化基质”（gene-activated matrices，GAM）可以被植入患者体内自动转染其周围的细胞。在本项目中，我们将通过体外实验来优化非病毒载体聚乙烯亚胺（polyethyleneimine, PEI）的参数以提高其转染效率，检测转染后的干细胞成骨能力和促血管形成能力的变化，以及上下游相关因子分泌的情况。我们将优化的PEI与纳米羟基磷灰石胶原支架混合制成PEI-ephrinB2 GAM，最后使用比格犬根尖周炎模型来验证其在牙髓血运重建术中的效果。

How to form vascular network rapidly in new regenerated tissue in root canals, and promote stem cells in canals differentiating into odontoblasts, determine the success of dental pulp revascularization. Our previous studies demonstrated that, dental stem cells transfected with ephrinB2 have enhanced osteogenic capacity, and promote new vascular networks formation using surrounded vascular endothelial cells. However, it is difficult to obtain dental stem cells and have the possibility of contamination to culture stem cells in vitro. Besides, the virus using in transfection is a potential risk for patients. Non-viral gene-activated matrices could be implanted in patients and transfect surrounded cells automatically. In this project, parameters will be optimized to improve transfection efficiency of polyethyleneimine in vitro. We will detect osteogenic and angiogenic capacity of stem cells after transfection, and the expression of related molecules. PEI-ehrinB2 GAM will be constructed using optimized PEI and collagen-nanohydroxyapatite. Finally, we will evaluate the effect of PEI-ehrinB2 GAM in dental pulp revascularization using periapical periodontitis model of beagle dogs.