中国组织工程研究与临床康复2011，Vol.15Issue(28)：5136-5140,5.DOI:10.3969/j.issn.1673-8225.2011.28.002
多次胶原酶消化法培养小鼠椎间盘髓核细胞
In vitro culture of mouse nucleus pulposus cells of intervertebral disc by multiple enzymatic digestions
摘要
Abstract
BACKGROUND: In vitro culture of nucleus pulposus cells of intervertebral disc is an important method to study degeneration of intervertebral disc. However, it is difficult to culture nucleus pulposus cells in vitro because the intervertebral disc is an avascular organ and nucleus pulposus cells poorly differentiate and proliferate.OBJECTIVE: To establish the method for in vitro culture of nucleus pulposus cells of intervertebral disc, and provide a reliable tool for research of phenotype changes of nucleus pulposus cells in degenerative disc and disc cell transplantation for treatment of degenerative disc diseases.METHODS: Mouse nucleus pulposus tissue was collected and repeatedly digested using collagenase. Cells were cultured and subcultured. The secretion of collagen Ⅱ and aggrecan of passage 2 cells were detected by immunohistochemistry and RT -PCR,The results were compared with other types of cells.RESULTS AND CONCLUSION: Nucleus pulposus cells of intervertebral disc exhibit a chondrocyte-like shape after adhesion.Immunohistochemistry study showed that the cells were positive for collagen Ⅱ and aggrecan staining. RT -PCR study showed that secretions of collagen Ⅱ and aggrecan in cultured cells were equal to those of chondrocyes and had significant difference from those of osteoblasts and fibroblasts. Multiple enzymatic digestions of nucleus pulposus can release a large amount of pure nucleus pulposus cells which had stable phenotype and can settle basis for research and treatment of intervertebal disc diseases.关键词
椎间盘退变/椎间盘髓核细胞/体外培养/Ⅱ型胶原/聚合蛋白分类
基础医学引用本文复制引用
张兴凯,曹鹏,王君,梁裕,吴文坚..多次胶原酶消化法培养小鼠椎间盘髓核细胞[J].中国组织工程研究与临床康复,2011,15(28):5136-5140,5.基金项目
国家自然科学基金(81071502),课题名称:缺氧诱导因子-1α(HIF-1α)对脊柱椎间盘发育及退变调控机制的研究. （81071502）
