人瘦素基因过表达增强大鼠骨髓基质细胞成骨能力  

Human leptin gene overexpression enhances osteogenesis of bone marrow stromal cells from rats

作　　者：郑宝玉[1] 江俊[2] 陈玉玲[2] 林敏魁[2] 骆凯[2] 闫福华[3] ZHENG Bao-yu JIANG Jun CHEN Yu-ling LIN Min-kui LUO Kai YAN Fu-hua(School and Hospital of Stomatology, Wenzhou Medical University, Wenzhou 325027 School and Hospital of Stomatology, Fujian Medical University, Fuzhou 350002 Institute and Hospital of Stomatology, Nanjing University Medical School, Nanjing 210008, China)

机构地区：[1]温州医科大学口腔医学院口腔医院,浙江温州325027 [2]福建医科大学口腔医学院口腔医院,福建福州350002 [3]南京大学医学院附属口腔医院,江苏南京210008

出　　处：《基础医学与临床》2017年第2期169-175,共7页Basic and Clinical Medicine

基　　金：国家自然科学基金(81100761);福建省自然科学基金(2012J05144);温州市科技局医学专项基金(Y20140312)

摘　　要：目的探讨人瘦素(h LEP)基因修饰对大鼠骨髓基质细胞(r BMSCs)成骨能力的影响。方法以转染h LEP腺病毒载体(Ad5-h LEP-EGFP)的r BMSCs为实验组,空载体(Ad5-EGFP)转染的r BMSCs和未转染的r BMSCs为对照组。MTT法检测细胞增殖情况,实时荧光定量PCR法检测Ⅰ型胶原(Col-Ⅰ)和碱性磷酸酶(ALP)mRNA的表达,茜素红染色检测矿化结节形成能力。同时,将各组r BMSCs与β磷酸三钙(β-TCP)复合后移植于裸鼠皮下培养8周,观察新骨形成情况。结果 Ad5-h LEP-EGFP能有效转染r BMSCs,转染后细胞增殖活性无明显改变,但Col-Ⅰ和ALP mRNA的表达较对照组有明显提高(P<0.05),矿化结节形成能力也更强(P<0.05)。而且,h LEP修饰后的r BMSCs能与β-TCP结合形成组织工程化复合物,并能在裸鼠体内形成更多的类骨样组织。结论 h LEP修饰的r BMSCs可增强细胞成骨能力,有望应用于骨及牙周组织再生研究。Objective To investigate the effects of human leptin（ h LEP） gene transfection on rat bone marrow stromal cells（ r BMSCs）. Methods r BMSCs were cultured and transfected with adenoviruses encoding h LEP（ Ad5-h LEP-EGFP） in vitro as experimental group while r BMSCs transfected with Ad5-EGFP and non-transfected were control groups. The proliferation was detected by MTT and the expression of collagen type Ⅰ（ Col-Ⅰ） and alkaline phosphatase（ ALP） were assessed by real-time PCR. The ability of mineralized nodule forming was also examined by Alizarin red staining. The combination of transfected r BMSCs and β-tricalcium phosphate（ β-TCP） was constructed and the osteogenic ability of the construction was evaluated in nude mice. Results h LEP could be transfected into r BMSCs successfully by adenovirous. After transfection,the proliferation was not affected while Col-Ⅰand ALP expressions were more pronounced in r BMSCs transfected with Ad5-h LEP-EGFP（ P〈0. 05）. Alizarin red staining showed the ability of mineralized nodule forming was also up-regulated in Ad5-h LEP-EGFP group（ P〈0. 05）. In addition,the transfected r BMSCs adhered to β-TCP and survived well and the combination showed more new bone like tissue formation in nude mice compared to control groups. Conclusions r BMSCs transfected with h LEP might be potently used in bone or periodontal tissue regeneration.

关 键 词：瘦素 骨髓基质细胞 基因转染 腺病毒 β磷酸三钙 

分 类 号：R781[医药卫生—口腔医学]