退变腰椎软骨终板细胞生物学特征实验研究

Biological characteristics of human degenerative vertebral endplate cells

目的通过对人腰椎软骨终板细胞的培养,观察细胞的形态学和生物学性状,探讨影响其生物学行为的因素。方法取腰椎退变和未退变终板软骨细胞,在加10%灭火胎牛血清的DMEM培养液中培养,建立体外终板软骨细胞培养模型,采取HE染色、绘制细胞生长曲线、Annexin-V/PI法、免疫组织化学染色、real-timePCR等方法,对细胞形态、活力、生长情况、凋亡、及软骨细胞基质合成进检测。结果软骨终板细胞可以在体外进行培养;终板软骨细胞的生长情况及细胞表型类似关节软骨,有II型胶原表达。退变软骨终板较未退变软骨终板活性及增殖能力降低,细胞凋亡增加,Ⅱ型胶原合成减少。结论体外成功培养了人腰椎终板软骨细胞,并证明了退变软骨终板细胞凋亡增加,而活性降低,基质合成减少。该研究也为进一步研究终板软骨的生物学性状奠定基础。

Objective To observe the morphology and phenotypes of cells extracted from the endplate in the intervertebral discs and identify the factors affecting their biological characteristic.Methods The intervertebral disc endplate were digested enzymatically,and the morphology of the obtained cells was examined under light microscope.Immunhistochemical analysis of collagen II and real-time PCR was carried out,and the morphologies,viability,cell growth,apoptosis and chondrocyte matrix production were compared between the cells isolated from the degenerative and normal vertebral endplates.Results The cells in primary culture presented with spherical and oval morphology,and the cytoplasm was stained blue with toluidine blue.The morphologies of the cartilage endplate cells and the articular cells were almost identical.All the freshly isolated cells expressed collagen II.The degenerative vertebral endplate cells showed decreased expression of collagen II with increased apoptotic cells as compared with normal vertebral endplate cells.Conclusion The intervertebral disc endplate cells,like articular cartilage cells,express cartilage-specific matrix proteins.Degenerative vertebral endplate cells show decreased cell vitality with increases cell apoptosis