改良酶消化法分离培养人乳牙牙髓干细胞  

Isolation and culture of dental pulp stem cells from human deciduous teeth by modified enzyme digestion

作　　者：别利克孜.卡德尔 刘奕杉[1] 王璇[1] 李伯琦[1] 马艳[1] 毕晓娟[1] 努斯来提.哈里克 

机构地区：[1]新疆医科大学第一附属医院儿牙预防保健科,新疆维吾尔自治区乌鲁木齐市830054 [2]新疆吉木萨尔县人民医院口腔科,新疆维吾尔自治区吉木萨尔县831700

出　　处：《中国组织工程研究》2013年第10期1793-1800,共8页Chinese Journal of Tissue Engineering Research

基　　金：新疆维吾尔自治区自然科学基金面上项目(2011211A067)~~

摘　　要：背景:乳牙牙髓干细胞具有极强的增殖活力,可以在短期内获得组织工程需要的细胞量。目的:比较传统酶消化法和改良酶消化法获取人乳牙牙髓干细胞的成功率及生物学特性。方法:取无龋滞留乳切牙12颗,随机数字表法均分为两组,分别应用传统酶消化法和改良酶消化法分离获取乳牙牙髓细胞,在不同代次细胞中检测细胞扩增天数、收获量、生长曲线、倍增时间、克隆形成率、细胞表面特异标记物STRO-1、CD146、CD34和CD45的表达。细胞成骨、成脂诱导分化能力及通过检测碱性磷酸酶和牙本质涎蛋白表达水平,判定细胞牙向分化潜能。结果与结论:两组细胞生长曲线,扩增天数、细胞收获量及细胞倍增时间相比,差异有显著性意义(P<0.05);牙向分化实验结果显示,改良酶消化法碱性磷酸酶和牙本质涎蛋白较传统酶消化法和对照组而言呈强阳性表达。然而,两组在克隆形成率、细胞表面特异标记物及多项诱导分化等方面差异无显著性意义(P>0.05)。说明改良酶消化法与传统酶消法相比,可在较短时间内完成同源乳牙牙髓干细胞体外扩增培养,可为牙齿再生治疗提供高质量种子细胞,是乳牙牙髓干细胞体外培养的可靠方法。BACKGROUND： Dental pulp stem cells from human deciduous teeth greatly proliferate and cell number required by tissue engineering can be easily acquired within a short time period. OBJECTIVE： To compare the cell harvest success rate and biological characteristics of dental pulp stemcells from human deciduous teeth between conventional enzyme digestion and modified enzyme digestion. METHODS： Twelve caries-free deciduous incisors were selected in this study. Dental pulp stem cells were harvested from these human deciduous teen by conventional enzyme digestion method （n=6 incisors） and modified enzyme digestion method （n=6 incisors）. Cell expansion days, total cell harvest, growth curves, cell doubling time, cell colony formation rate, cell surface specific markers STRO-1, CD146, CD34 and CD45 were determined among different generations of cells. Osteogenic and adipogenic differentiation of cells were determined by detecting alkaline phosphatase and dentin sialoprotein expression levels. RESULTS AND CONCLUSION： There were significant differences in growth curves, cell expansion days, total cell harvest and cell doubling time between conventional enzyme digestion and modified enzyme digestion （P＆lt;0.05）. Alkaline phosphatase and dentin sialoprotein expression levels were significantly higher in dental pulp stem cells cultured by modified enzyme digestion than by conventional enzyme digestion method. However, there were no significant differences in cell colony formation rate, cell surface specific marker expression and induced cell differentiation between modified enzyme digestion and conventional enzyme digestion （P＆gt;0.05）. Compared to conventional enzyme digestion, modified enzyme digestion can assist in in vitro expansion and culture of dental pulp stem cells from homologous deciduous teeth within a short time period and provide high-quality seed cells for tooth regeneration and is a reliable method for in vitro culture of dental pulp stem cells from deciduous teeth.

关 键 词：干细胞 干细胞培养与分化 乳牙牙髓细胞 改良酶消化法 牙本质涎蛋白 体外培养 细胞鉴定 牙向分化 省级基金 干细胞图片文章 

分 类 号：R394.2[医药卫生—医学遗传学]