猪牙髓干细胞体外分离培养与分化研究  

The differentiation,isolation and cultivation of swine dental pulp stem cells

作　　者：郭俊[1] 杨健[1] 贺露[1] 李伟 胡红梅 GUO Jun;YANG Jian;HE Lu;LI Wei;HU Hong - mei(Department of Endodontics, Affiliated Stomatology Hospital of Nanchang University, Nanchang 333006, Jiangxi, Chin)

机构地区：[1]南昌大学附属口腔医院牙体牙髓科,江西南昌333006 [2]井冈山大学医学部口腔系,江西吉安343000

出　　处：《广东医学》2018年第11期1612-1615,共4页Guangdong Medical Journal

基　　金：国家自然科学基金资助项目(编号:81660188)

摘　　要：目的探讨分离培养猪牙髓干细胞(s DPSCs)的方法,为下一步的研究及牙成体干细胞的应用提供理论基础。方法 6月龄猪磨牙获取牙髓;采用酶消化法获取s DPSCs,取第1代细胞为研究对象计算细胞集落形成率(CFU);采用免疫荧光等技术检测s DPSCs标记物;体外诱导细胞成骨、成脂分化。结果酶消化法获取的第1代s DPSCs的CFU-F为2~8 colonies/103;细胞贴壁生长,短梭形;体外可连续培养多代;s DPSCs碱性磷酸酶(ALP)、巢蛋白、波形蛋白阳性表达;Stro-1阳性率为75.4%,CD90为99.8%;CD24为42.6%;不表达CD34和牙本质涎蛋白(DSP);s DPSCs诱导后有DSP和ALP的表达;诱导条件下s DPSCs可向成牙本质细胞和脂肪细胞方向分化。结论采用酶消化培养法能够有效地分离得到s DPSCs;猪牙髓中存在牙髓干细胞并具有克隆繁殖能力和分化能力。Objective To study the cultivation and isolation of swine dental pulp stem cells （sDPSCs）, so as to a basis for future applications and research of sDPSCs. Methods Molars from six months old pig were taken to obtain pulp, and single cell supernatant was obtained by enzyme digestion. The expression of sDPSCs specific proteins was detec- ted by immunofluorescence and flow cytometry. Colony - forming efficiency and differentiation potential were evaluated. Results The average of CFU - F of sDPSCs was 2 - 8 colonies/103 cells plated. The sDPSCs were able to adhere to plas- tic substratum and showed short fibroblast -like shape. Cultured SDPSCs were found to express ALP, Vimentin, Nestin, Stro - 1 （75.4%）, CD90（99. 8% ） and CD24 （42. 6% ）, but not CD34 or DSP. With adipogenic medium induction, some sDPSCs showed adipogenic potential. Long -term cultures of sDPSCs grown in osteo/dentinogenic medium demon- strated dentinogenic potential with high levels of calcium and expression of DSP and ALP. Conclusion sDPSCs survived in the swine dental pulp can be isolated with enzymatic digestion, and induced into muhipotent differentiation.

关 键 词：猪 牙髓干细胞 分离 分化 

分 类 号：R78[医药卫生—口腔医学]