人牙周膜细胞在左旋聚乳酸/羟基磷灰石生物材料上的生长观察  

OBSERVATION OF HUMAN PERIODONTAL LIGAMENT CELLS CULTURED ON ELECTROSPUN PLLA/HA BIOMATERIAL

作　　者：王琳[1] 冯海兰[1] 梅芳[2] 胡晓阳[3] 邓旭亮[3] 杨小平[4] 唐军民[2] 王新知[1] 

机构地区：[1]北京大学口腔医学院修复科 [2]北京大学基础医学院组织学与胚胎学教研室,北京100083 [3]北京大学口腔医学院特诊科,北京100081 [4]北京化工大学,北京100029

出　　处：《解剖学报》2008年第4期573-577,共5页Acta Anatomica Sinica

基　　金：国家自然科学基金资助项目(30471907)

摘　　要：目的观察电纺左旋聚乳酸/羟基磷灰石(PLLA/HA)生物材料(简称生物材料)的生物相容性,并以人牙周膜细胞作为种子细胞与生物材料复合培养,探索该生物材料用于人牙周组织再生的可能性。方法用电纺法制备PLLA/HA纤维生物材料;组织块法分离培养人牙周膜细胞,并用免疫组织化学方法鉴定;MTT法评价生物材料的生物相容性;将人牙周膜细胞与生物材料复合培养,用扫描电镜进行观察;将转染人腺病毒介导的绿色荧光蛋白的人牙周膜细胞与生物材料复合培养,用激光扫描共焦显微镜观察。结果成功分离了人牙周膜细胞,波形蛋白染色阳性确定该细胞来源于中胚层;MTT法检测人牙周膜细胞在生物材料上的增殖状况与正常培养基本一致;扫描电镜下可见细胞在生物材料上生长旺盛、充分伸展;激光扫描共焦显微镜下可见人牙周膜细胞沿生物材料呈纤丝生长,表现出一定的三维结构。结论电纺PLLA/HA生物材料具有三维空间网状结构和良好的生物相容性,与人牙周膜细胞复合培养有利于细胞的生长、贴附和增殖,并呈现一定的立体结构,为人牙周组织再生提供了实验依据。Objective To observe the attachment and growth of human periodontal ligament cells （hPDLCs） on electrospun poly （L-lactic acid）/hydroxyapatite （PLLA/HA） fibrous biomaterial （called ＂ biomaterial＂ for short）, and to investigate the feasibility of applying this new material in human periodontal tissue regeneration. Methods PLLA/HA fibrous biomaterial was fabricated by electrospinning technique, the hPDLCs were harvested from healthy premolars extracted for orthodontics, and the ceils were identified by immunohistochemical method. The compatibility of the biomaterial was evaluated by the MTT assay method, the hPDLCs were seeded onto the PLA/HA fibrous biomaterial, and then the compound was observed under scanning electron microscope ; the hPDLCs transfected by the recombinant adenovirus containing green fluorescent protein was co-cultured with the biomaterial and observed under confocal laser scanning microscope. Results The hPDLCs were successfully separated, and immunohistochemistry staining of vimentin protein showed that the cells came from the mesoderm; the MTF assay results showed that the cells propagated on the biomaterial well compared with cultured normally; the scanning electon microscope （SEM） results showed the cells on the biomaterial grew and stretched well; the confocal laser scanning microscope results showed that the cells grew along the fiber of the biomaterial, and presented a certain three dimensional structure. ConcIusion The results suggested that electrospun PLLA/HA fibrous biomaterial possesses a three dimensional net-like structure and a good cellular compatibility with hPDLCs; the hPDLCs can adhere and propagate well on it. This experiment can provide some basic theory for the use of electrospun PLLA/HA fibrous biomaterial in the regeneration of human periodontal tissue.

关 键 词：组织工程 电纺 左旋聚乳酸 羟基磷灰石 牙周膜细胞 免疫组织化学 细胞培养 人 

分 类 号：R392[医药卫生—免疫学]