新型培养基促进牙乳头细胞成骨分化及其在牙周骨组织再生中的应用  

A Novel Chemically Defined Medium Enhanced the Osteogenic Potential and Periodontal Bone Regeneration of Dental Papilla Cells

作　　者：王涛 王世恺 陈国庆 田卫东[1,2,3] WANG Tao;WANG Shi-kai;CHEN Guo-qing;TIAN Wei-dong(State Key Laboratory of Oral Diseases,National Clinical Research Center for Oral Diseases,West China Hospital of Stomatology,Sichuan University,Chengdu 610041,China;Department of Oral and Maxillofacial Surgery,West China Hospital of Stomatology,Sichuan University,Chengdu 610041,China;National Engineering Laboratory for Oral Regenerative Medicine,West China Hospital of Stomatology,Sichuan University,Chengdu 610041,China)

机构地区：[1]口腔疾病研究国家重点实验室国家口腔疾病临床医学研究中心四川大学华西口腔医院,成都610041 [2]四川大学华西口腔医院创伤与整形外科,成都610041 [3]口腔再生医学国家地方联合工程实验室四川大学华西口腔医院,成都610041

出　　处：《四川大学学报（医学版）》2020年第6期735-741,共7页Journal of Sichuan University(Medical Sciences)

基　　金：国家重点研发计划(No.2017YFA0104800)资助。

摘　　要：目的 探讨新型化学成分明确培养基(chemically defined medium,CDM)对牙乳头细胞(dental papilla cells,DPCs)成骨分化潜能及体内牙周骨再生的影响.方法 分离大鼠DPCs,按培养基不同分为传统培养基(conventional medium,CM)组和CDM组,分别在CM和新型CDM中培养,评估两组DPCs细胞表面标记、增殖、迁移、成骨分化潜能;制备SD大鼠牙周骨缺损模型,CM和CDM两组DPCs分别复合胶原凝胶植入缺损区域内,评估两组DPCs在牙周骨缺损中的骨再生修复能力.结果 在CM和CDM组中,DPCs细胞均显示相似的细胞表面标记;与CM相比,CDM可促进DPCs增殖、克隆形成及细胞迁移(P<0.05);定量PCR显示CDM组DPCs成骨相关基因Runx2、Alp和Opn上调(P<0.05);碱性磷酸酶(alkaline phosphatase,ALP)染色显示CDM组DPCs细胞ALP活性高于CM组(P<0.05);细胞经成骨诱导后,茜素红染色显示CDM组DPCs细胞成骨分化能力高于CM组(P<0.05).在大鼠体内牙周骨缺损模型中,移植CDM组DPCs细胞8周后,HE染色显示缺损区域皮质骨连续的新生骨,而CM组新生骨较少,骨皮质仍未愈合,microCT扫描定量分析显示CDM组骨体积分数(BV/TV)高于CM组(P<0.05).结论 CDM培养基可促进DPCs增殖和成骨分化能力,为未来细胞治疗中干细胞培养基的选择提供了新的方案.Obejective To investigate the role of a novel chemically defined medium(CDM) in the regulation of dental papilla cells(DPCs) functional phenotype in vitro and periodontal bone regeneration in vivo. Methods DPCs were isolated and cultured in conventional medium(CM) or CDM. The surface makers, and the proliferation, migration and osteogenic differentiation abilities of DPCs were evaluated. In vivo, the DPCs that mixed with collagen gel were implanted into the model rats in the defect of periodontal to repair the periodontal tissue. Regeneration of the tissues was examined by microcomputed tomography and histological observation. Results DPCs in the CM group and CDM group showed similar surface markers. Compared to the CM group, the CDM significantly enhanced the proliferation,colony-forming efficiency and migration of DPCs in vitro. In addition, real time PCR showed that the expression levels of osteogenesis-related genes, Runx2, Alp and Opn. were significantly enhanced in DPCs in the CDM group. DPCs cells treated with CDM also exhibited higher alkaline phosphatase activity and stronger ability of formation of mineralized nodules in vitro. In vivo, DPCs from CDM group significantly enhanced the periodontal bone regeneration and the reconstruction of periodontal bone tissues in rat periodontal defect model. Conclusion CDM is a suitable medium to culture DPCs for periodontal bone regeneration. This research provided a substitute for basic research and set the stage for future clinical application of stem cell transplantation.

关 键 词：牙周骨再生 化学成分明确培养基 牙乳头细胞 组织工程 

分 类 号：R781[医药卫生—口腔医学]