中国组织工程研究Issue(39)：6861-6866,6.DOI:10.3969/j.issn.2095-4344.2013.39.001
Ⅱ型胶原酶消化结合组织块贴壁法分离培养兔髓核细胞
Isolation and culture of rabbit nucleus pulposus cells in vitro by type Ⅱ collagenase digestion methods plus explant culture method
摘要
Abstract
BACKGROUND:Intervertebral disc can bear load but lack vessels. Nucleus pulposus cel s have the problem of phenotype loss during in vitro culture that can lead to degenerative changes. The mechanism of intervertebral disc degeneration remains unclear. <br> OBJECTIVE:To explore the approaches of isolation, adherence culture, amplification and identification of the rabbit nucleus pulposus cel s in vitro, to observe the growth characteristics of nucleus pulposus cel s in different generations. <br> METHODS:Type Ⅱ col agenase digestion method plus explants culture method was used to isolate and purify nucleus pulposus cel s and then amplify in vitro. The morphology and growth of primary and passaged cel s was observed under the inverted microscope, the number of cel s was counted and the growth curve was draw. The morphology of the cel s was observed under light microscope after hematoxylin-eosin staining, and the expressions of col agen type Ⅱ and aggrecan were examined by immunocytochemistry. <br> RESULTS AND CONCLUSION:Nucleus pulposus cel s of rabbit were isolated, cultured and amplified in vitro successful y. Growth activity was observed, and found that the 1-3 generation nucleus pulposus cel s proliferated more rapidly and vigorously. The proliferation of nucleus pulposus cel s was decreased while the cel passaged more generations. These isolated and cultured nucleus pulposus cel s could positively express the col agen type Ⅱ and aggrecan. In vitro combination of type Ⅱ col agenase digestion method and explants culture method could obtain high purity nucleus pulposus cel s, and the cultured nucleus pulposus cel s were grew in round or polygonal. The 1-3 generation of cel s had strong activity.关键词
骨关节植入物/骨关节损伤基础实验/髓核细胞/Ⅱ型胶原酶消化法/组织块贴壁法/Ⅱ型胶原/聚集蛋白聚糖/体外培养/细胞增殖/省级基金分类
医药卫生引用本文复制引用
李树文,武海军,银和平,白明,杜志才..Ⅱ型胶原酶消化结合组织块贴壁法分离培养兔髓核细胞[J].中国组织工程研究,2013,(39):6861-6866,6.基金项目
内蒙古自治区教育厅高等学校科学研究基金项目资助(NJZY13423)@@@@Science Foundation+of Universities by Education Ministry of Inner Mongolia Autonomous Region, No. NJZY13423 （NJZY13423）
