大鼠颌骨与髂骨来源骨髓基质干细胞体外生物学特性的比较研究

发布时间：2018-04-02 13:19

本文选题：骨髓基质细胞　切入点：颌骨　出处：《南京医科大学》2008年硕士论文

【摘要】： 因创伤、感染、肿瘤及先天性疾病等造成的颌骨缺损十分常见,目前针对颌骨缺损的主要修复方法是自体骨移植,其中以髂骨作为骨供体最为常见。胚胎学研究表明大部分颅颌面骨发生于颅神经嵴来源外胚间充质,而躯干四肢骨来源于中胚层;从成骨方式而言颌骨为膜内成骨,髂骨则为软骨内成骨。大量临床与动物实验表明截断性颌骨缺损采用颌骨供体比其他部位骨供体成功率更高,主要表现为移植供体存活率更高,远期吸收率更低。这些已有的临床、实验室、生物学研究提示我们在不同部位骨骨髓内的骨祖细胞可能具有各自生物学特性;而关于骨组织修复的细胞学模型主要来自长骨,对颌骨来源骨髓细胞的针对性研究甚少。为此,我们采用大鼠颌骨与髂骨来源骨髓基质干细胞(Bone Marrow StromalCells,BMSCs)进行体外扩增培养,通过相应的诱导条件,观察这两种不同来源的BMSCs各自在细胞增殖,集落形成,分化成骨,矿化形成及分化成脂方面是否存在差异,希望借此为进一步探索颌骨在发生、修复上是否具有其独特生物学机制提供一定的依据和参考。第一部分大鼠颌骨与髂骨来源骨髓基质干细胞的体外培养目的:从大鼠的颌骨与髂骨骨髓中运用全骨髓细胞贴壁培养法分别培养出两位点BMSCs,为探讨颌骨与髂骨来源BMSCs的生物学特性是否具有位点性差异提供细胞模型。方法:取4～6周龄150克±10克雄性SD大鼠,首先解剖分离出下颌骨及髂骨,无菌条件下培养基冲出骨髓细胞制成细胞悬液,采用全骨髓贴壁法培养细胞,相差显微镜下观察细胞生长情况及特点。结果:原代细胞接种24h后镜下可见瓶底有细胞贴壁,首次换液去除大部分外周血细胞后可见散在的梭形细胞聚集;随反复换液,瓶内未贴壁细胞逐步被清除,细胞生长形成典型的成纤维细胞样-集落形成单位(colony-forming unit-fibroblast,CFU-F)。细胞首次传代约需2周左右,传代细胞于第6代(髂骨)-第9代(颌骨)前细胞增殖能力比较旺盛,以后细胞活力逐渐减弱,出现不同的分化倾向,部分细胞胞浆中出现空泡及颗粒样物质,最终死亡。结论:全骨髓细胞贴壁培养法能够培养出满足实验要求的BMSCs。第二部分大鼠颌骨与髂骨来源骨髓基质干细胞体外生物学特性的比较研究目的:探讨体外环境下大鼠颌骨与髂骨来源BMSCs体外生物学性状是否存在差异,为进一步深入体内研究提供依据。方法:取第二代正常生长的颌骨与髂骨BMSCs,分别从细胞增殖活性,诱导成骨,矿化及成脂能力等多方面比较两者的体外生物学特性的差异。结果:颌骨来源的BMSCs与髂骨相比,在低浓度接种条件下具有更强的增殖能力,诱导成骨后更高的碱性磷酸酶阳性表现,矿化诱导后形成更多的钙结节;而髂骨来源BMSCs在成脂诱导条件下分化形成更多脂质。结论:颌骨与髂骨来源BMSCs在体外具有不同的生物学特点,提示颌骨来源的BMSCs可能具有其独特生物学特性,为进一步研究颌骨病变与再生修复机制提供实验依据和参考。
[Abstract]:The main repair method for jaw defects is autogenous bone grafting , which is the most common bone donor in the bone graft . The study of embryology shows that most of the cranio - maxillofacial bones are derived from the ectodermal mesenchyme of the cranial nerve ridge , and the bone of the trunk is derived from the endoderm . The large number of clinical and animal experiments show that the bone donor has higher success rate than other parts of bone . The main expression is higher survival rate and lower long - term absorption rate of the graft donor .

These existing clinical , laboratory and biological studies suggest that bone progenitor cells in bone marrow of different parts may have their own biological characteristics ; the cytologic model for bone tissue repair is mainly derived from long bones . Bone marrow stromal cells derived from bone marrow derived from bone marrow are cultured in vitro .

In vitro culture of bone marrow stromal cells derived from bone marrow stromal cells from the first part of the rat ' s jaw and ilium

Objective : To study the biological characteristics of bone marrow cells from the bone marrow of the bone and ilium of the rat , and to provide a cell model .

Methods : Male Sprague - Dawley rats aged from 4 to 6 weeks old were dissected and isolated from bone marrow cells .

Results : After 24 hours of primary cell inoculation , there was a cell adherent on the bottom of the cell , and most of the peripheral blood cells were removed for the first time to remove most of the spindle cells which were scattered in the peripheral blood cells . The cells were removed gradually with the repeated liquid exchange , and the cells were gradually removed and the cells were grown into a typical fibroblast - colony forming unit ( CFU - F ) . After the first passage of the cells about 2 weeks , the cell proliferation ability of the passaged cells in the 6th generation ( ilium ) - 9th generation ( jaw ) was relatively strong , the cell viability was gradually weakened , and the cells appeared different differentiation tendency , and vacuoles and granular substances appeared in the cytoplasm of some cells , and finally died .

Conclusion : The whole bone marrow cell adherent culture method can be used to cultivate the bone marrow cells which meet the experimental requirements .

Comparative study on the in vitro biological characteristics of bone marrow stromal cells derived from bone marrow stromal cells from the second part of the rat ' s jaw and ilium

Objective : To investigate the difference of biological characters of bone and iliac bone in vitro in vitro and to provide a basis for further in vivo study .

Methods : To compare the biological characteristics of bone , bone , mineralization and lipid - forming ability of the second - generation normal - growing jaw and ilium , respectively .

Results : Compared with ilium , bone marrow derived from bone marrow had stronger proliferative ability under low concentration inoculation condition , higher alkaline phosphatase positive expression after induction of bone formation , and formed more calcium nodules after mineralization induction .

Conclusion : The bone and ilium sources have different biological characteristics in vitro , suggesting that they may possess their unique biological characteristics and provide experimental basis and reference for further study of the mechanism of mandibular lesions and regenerative repair .

【学位授予单位】：南京医科大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R329

【引证文献】