摘要
目的建立体外兔椎间盘脊索细胞藻酸盐凝胶培养模型,观察脊索细胞形态及生物学特点。方法采用胶原酶消化法及Percoll不连续密度梯度沉淀法体外分离收集原代椎间盘脊索细胞,于1.2%藻酸盐凝胶(低密度)中培养。倒置相差显微镜下观察细胞形态,经Ⅱ型胶原免疫荧光染色对细胞表型初步鉴定,并分别以细胞增殖和细胞毒性试剂-8(CCK-8)检测细胞在藻酸盐凝胶中的存活和增殖能力。结果成功分离获得原代椎间盘脊索细胞,可稳定表达Ⅱ型胶原。原代脊索细胞在藻酸盐凝胶中生长良好,但增殖缓慢。结论初步了解兔椎间盘脊索细胞体外生物学特性,为椎间盘退变机制及组织工程学髓核种子细胞的研究提供一定的实验依据。
Objective To make the culture model in vitro of notochordal cells in rabbit intervertebral disc with alginate gel and observe the morphological and biological characteristics. Methods Primary notochordal cells were obtained by conagenase digestion and Percoll discontinuous density gradient centrifugation, and then embedded in low-viscosity alginate gels （1.2 %）. The cell morphology was observed under inverted phase contrast microscope, preliminary identification was analyzed by immunofluorescence staining of collagen type iT. The cells staining and cell counting kit-8 （CCK-8） were used to detect the survival and proliferation of notochordal cells in alginate gels, respectively. Results The primary notochordal cells were isolated Successfully, the stable expression of collagen type 11 was observed in it. And it grew well in alginate gels, but the proliferation was very slow. Condusions The biological characteristics of notochordal cells in vitro have a preliminary understanding, it would provide certain experimental evidence for the researches on the mechanism of disc degeneration and the seed cells for tissue engineering nucleus pulposus.
出处
《国际骨科学杂志》
2012年第2期126-128,共3页
International Journal of Orthopaedics
基金
国家自然科学基金(30872610
30700841)
关键词
脊索细胞
藻酸盐
椎间盘
Notochordal cell Alginate
Intervertebral disc
