CEMP1基因修饰骨质疏松大鼠BMSCs与β-TCP体外复合培养实验  

Co-culture of CEMP1 gene modified osteoporotic rat BMSCs with β-TCP in vitro

作　　者：江俊[1] 吴玉铭 何梦娇[1,3] 郑宝玉 许雄程[1,3] 李艳芬 陈玉玲[1] 骆凯[1] JIANG Jun;WU Yuming;HE Mengjiao;ZHENG Baoyu;XU Xiongcheng;LI Yanfen;CHEN Yuling;LUO Kai(School and Hospital of Stomatology,Fujian Medical University,Fuzhou 350002;The First Affiliated Hospital of USTC Anhui Provincial Hospital,Hefei 230000;Institute of Stomatology,Fujian Medical University,Fuzhou 350002;School and Hospital of Stomatology,Wenzhou Medical University,Wenzhou 325027,China)

机构地区：[1]福建医科大学附属口腔医院,福建福州350002 [2]中国科学技术大学附属第一医院安徽省立医院,安徽合肥230000 [3]福建医科大学口腔医学研究院,福建福州350002 [4]温州医科大学附属口腔医院,浙江温州325027

出　　处：《中国骨质疏松杂志》2020年第2期157-161,共5页Chinese Journal of Osteoporosis

基　　金：国家自然科学基金项目(81870766);福建省自然科学基金项目(2017J01522);福建省卫生系统中青年骨干人才培养项目(2015-ZQN-ZD-28);福建医科大学启航基金项目(2016QH079)

摘　　要：目的探讨牙骨质蛋白1(cementum protein 1,CEMP1)基因修饰骨质疏松大鼠骨髓基质细胞(bone marrow stromal cells,BMSCs)与β-磷酸三钙(β-tricalcium phosphate,β-TCP)体外复合培养的可行性。方法体外培养骨质疏松大鼠BMSCs,将携带有CEMP1基因的慢病毒LV-CEMP1-EGFP转染BMSCs,采用荧光显微镜观察转染情况,通过流式细胞仪检测转染率,通过实时荧光定量PCR检测转染后BMSCs中CEMP1基因的表达,Western Blot及免疫细胞化学染色检测BMSCs中CEMP1蛋白的表达情况。将CEMP1基因修饰的BMSCs与β-TCP体外复合培养,在荧光显微镜和扫描电镜下观察细胞的生长状况。结果LV-CEMP1-EGFP具有较高的转染率,CEMP1基因修饰的BMSCs可高表达CEMP1,且在β-TCP上保持良好的生长状态。结论CEMP1基因修饰骨质疏松大鼠BMSCs与β-TCP复合培养,有望应用于骨质疏松状态下的牙周组织再生研究。Objective To evaluate the feasibility of co-culture of cementum protein 1(CEMP1) gene modified osteoporotic rat bone marrow stromal cells(BMSCs) with β-TCP in vitro. Methods BMSCs of osteoporotic rats were isolated and cultured in vitro and transfected with lentiviral vector carrying CEMP1 gene(LV-CEMP1-EGFP) and detected under inverted fluorescence microscope.Transfection efficiency was detected by flow cytometry 72 hours after transfection. Expression of CEMP1 in BMSCs was detected by real time PCR,Western blot and immunohistochemistry. LV-CEMP1-EGFP transfected cells were co-cultured with β-TCP and were observed under inverted fluorescence microscope and scanning electron microscope(SEM). Results LV-CEMP1-EGFP has high transfection rate and CEMP1 was overexpressed in BMSCs. Fluorescent microscope and SEM demonstrated that CEMP1 gene modified BMSCs could migrate into β-TCP and secrete extracellular matrix. Conclusion Recombinant lentiviral vector containing human CEMP1 gene can be transfected into osteoporotic rat BMSCs efficiently. CEMP1 gene modified BMSCs can be combined with β-TCP and grow well. The composite might be beneficial for periodontal tissue regeneration with osteoporosis.

关 键 词：牙骨质蛋白1 骨髓基质细胞 骨质疏松 慢病毒 Β-磷酸三钙 

分 类 号：R681[医药卫生—骨科学]