人釉原蛋白真核表达载体体外表达产物生物学活性的检测

目的探讨人釉原蛋白基因重组质粒PcDNA3．1-AMG的真核细胞转染表达产物是否具有促进牙周组织再生的作用。方法脂质体介导PcDNA3．1-AMG体外转染COS1细胞．ELISA法检测转染细胞内及其培养上清液中重组釉原蛋白的表达：建立犬牙周组织缺损模型．局部使用转染表达产物冻干粉．8周后通过组织学观察牙周组织再生的情况。结果PcDNA3．1-AMG转染的COS1细胞内重组釉原蛋白浓度为（0．253±0．075）μg／ml。培养液中浓度为（0．065±0．011）μg／ml；使用转染表达产物8周后．牙周缺损区牙骨质、牙槽骨均有显著再生，并且新生牙骨质为有胶原纤维穿通的无细胞牙骨质。结论重组质粒PcDNA3．1－AMG在体外转染哺乳动物细胞后能够表达重组人釉原蛋白．并且表达产物具有促进牙周组织再生的生物学活性。

Experimental study on the effect of expressed products of recombinant plasmid for human amelogenin to promote periodontal regeneration

Objective To investigate whether the expressed products of recombinant plasmid PcDNA3.1-AMG for human amelogenin in COS-1 cell lines could promote periodontal regeneration. Methods Deliver the recombinant plasmid PcDNA3.1-AMG for human amelogenin to COS-1 cell lines in vitro. Recombinant amelogenin in the transfected cells and cultured supernant were detected by ELISA technique. Buccal dehiscence models were made in dogs, and then the lyophilized cultured supernant of PcDNA3.l-AMG trasfected cells was applied to the defected areas. After 8 weeks, the periodontal regeneration was evaluated by histological methods. Results The concentrations of recombinant amelogenin in PcDNA3.1-AMG trasfected cells and cultured supernant were （0.253 ± 0.075） μg/ml and （0.065± 0.011） μg/ml respectively. And application of lyophilized expressed products resulted in significant regeneration of acellular cementum with collagenous fibers extending to newly formed alveolar bone. Conclusions There is expression of recombinant amelogenin in PcDNA3.1-AMG trasfected COS1 cells. And the expressed products possess the ability to promote periodontal regeneration.