| 杨勇,梁伟,罗卓荆,张世磊,陶惠人.不同Pfirrmann分级椎间盘内髓核细胞生物学特性的比较[J].脊柱外科杂志,2011,9(4):244-248. |
不同Pfirrmann分级椎间盘内髓核细胞生物学特性的比较  点此下载全文 (Fulltext) |
| 杨勇 梁伟 罗卓荆 张世磊 陶惠人 |
710032 陕西, 第四军医大学附属西京医院骨科(杨勇现工作于解放军535医院骨科)
;710032 陕西, 第四军医大学附属西京医院骨科(杨勇现工作于解放军535医院骨科)
;710032 陕西, 第四军医大学附属西京医院骨科(杨勇现工作于解放军535医院骨科)
;710032 陕西, 第四军医大学附属西京医院骨科(杨勇现工作于解放军535医院骨科)
;710032 陕西, 第四军医大学附属西京医院骨科(杨勇现工作于解放军535医院骨科)
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| 基金项目：国家自然科学基金资助项目（2007BA104B07） |
| DOI：10.3969/j.issn.1672-2957.2011.04.014 |
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| 摘要： |
| 目的 比较不同Pfirrmann分级椎间盘内髓核细胞的生物学特性，验证腰椎间盘退变Pfirrmann分级能否反应髓核组织在细胞水平的退变程度。方法 取术中获得的患者的髓核组织，按腰椎椎间盘退变Pfirrmann分级标准进行分组。组织切片甲苯胺蓝观察比较各组髓核内细胞的密度。采用酶消化法分离培养髓核细胞。台盼蓝染色计算各组细胞活性比率，光镜观察细胞形态，透射电镜观察细胞的超微结构。MTT法绘制各组2代细胞的生长曲线。阿利辛蓝法检测各组细胞的蛋白聚糖含量，比较各组差异。结果 随退变级别升高，标本内胶冻样物质含量减少，透明度降低，纤维化组织增多，Ⅴ级标本完全纤维化，并伴有钙化，无法区分髓核组织。Ⅰ级和Ⅱ级髓核内细胞密度明显高于Ⅲ级，Ⅳ级又明显高于Ⅳ级。Ⅰ级组髓核细胞的活性比率为（93.5±3.7）%；Ⅱ级组细胞活性比率为（91.6±4.3）%；Ⅲ级组细胞活性比率为（83.5±6.7）%；Ⅳ级组细胞活性比率为（74.8±5.9）%。除Ⅰ级与Ⅱ级比较差异无统计学意义外，其余各组两两比较差异均具有统计学意义（P<0.05）。光镜下Ⅰ级和Ⅱ级组细胞呈短梭形或多角形轮廓清晰饱满，折光性强。Ⅲ级组胞突较长，Ⅳ级组胞突更长，细胞轮廓模糊。电镜下Ⅰ级和Ⅱ级组细胞相似，胞浆内含有大量粗面内质网和线粒体，Ⅲ级组细胞内的粗面内质网和线粒体减少，出现小的空泡，可见溶酶体出现。Ⅳ级组细胞溶酶体大量聚集，可见巨大的空泡样结构。Ⅰ级组和Ⅱ级组细胞生长速率快于Ⅲ级组（P<0.01））。Ⅲ级组细胞生长速率快于Ⅳ级组（P<0.05）。Ⅰ、Ⅱ、Ⅲ、Ⅳ组细胞GAG含量分别为（423.19±41.21） mg/L，（408.23±29.25） mg/L、（273.05±52.44） mg/L、（91.73±38.06） mg/L，Ⅰ级组和Ⅱ级组细胞高于于Ⅲ级组（P<0.05））。Ⅲ级组细胞高于Ⅳ级组（P<0.05）。结论 椎间盘退变的Pfirrmann分级，能很好的反映髓核组织在细胞水平的退变程度。是一种简单有效的退变分级系统。 |
| 关键词：腰椎 椎间盘 细胞培养技术 |
| Biological characteristics of nucleus pulposus cells in intervertebral discs at different Pfirrmann grades Fulltext |
| YANG Yong LIANG Wei LUO Zhuo-jing ZHANG Shi-lei TAO Hui-ren |
| Department of Orthopaedics, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi, China;Department of Orthopaedics, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi, China;Department of Orthopaedics, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi, China;Department of Orthopaedics, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi, China;Department of Orthopaedics, Xijing Hospital, Fourth Military Medical University, Xi'an 710032, Shaanxi, China |
| Fund Project: |
| Abstract: |
| Objective To compare the biological characteristics of nucleus pulposus cells in intervertebral discs at different grades of Pfirrmann grading system in order to determine whether the system can reflex cell degeneration.Methods Human nucleus pulposus cells were isolated by collagenase digestion and primarily cultured. Then the cells were grouped based on the Pfirrmann grading system. Tissue sections were dyed by Toluidine blue to observe the cell density. Cell vitality assay was done with Trypan blue staining. Morphologic changes of the cells were observed by inverted microscopy. Cell ultra-structure morphological changes were examined by using transmission electron microscopy. Cell growth curves were determined by MTT assay. Proteoglycans expressions were examined by Alican blue chromatometry.Results With the increase of Pfirrmann grade, there were lesser gel-like substance and more fibrosis in the cells with decreased transparency. Cells in Grade Ⅴ were totally fibrillated with calcification, and their nucleus pulposus was unclear. Cell densities in Grade Ⅰ and Ⅱ were higher than that in Grade Ⅲ, followed by that in Grade Ⅳ. Cell vitalities of cells in Grade Ⅰ, Ⅱ,Ⅲ and Ⅳ were (93.5±3.7)%, (91.6±4.3)%, (83.5±6.7)% and (74.8±5.9)%, respectively, with significant differences found between every 2 groups, except for between Grade Ⅰ and Grade Ⅱ (P<0.05). The cells in Grade Ⅰ and Ⅱ were short spindle or polygonal in shape, with clear outlines and rich cytoplasm, a lot of rough endoplasmic reticulum and mitochondria. While the cells in Grade Ⅲ had long processes, vague outline, and lesser rough endoplasmic reticulum and mitochondria, small vacuoles and lysosome in the cytoplasm. For the cells in Grade Ⅳ, their processes were even longer, large amount lysosomes were accumulated and large vacuoles were formed. Under the same cellular density and culture conditions, the growth rate of passage 2 cells were markedly faster in the cells of Grade Ⅰ and Ⅱ than in Grade Ⅲ (P<0.01), and that of Grade Ⅲ was faster than of Grade Ⅳ (P<0.05). The contents of GAG of grade Ⅰ, Ⅱ, Ⅲ and Ⅳ were (423.19±41.21) mg/L, (408.23±29.25) mg/L, (273.05±52.44) mg/L and (91.73±38.06) mg/L, respectively. Those of Grade Ⅰ and Ⅱ were significantly higher than that of Grade Ⅲ, and so was that of Grade Ⅲ than Grade Ⅳ(P<0.05). Conclusion Pfirrmann grading system of disc degeneration can well reflect the degenerative degree of nucleus pulposus cells. It is a simple and effective disc degeneration grading system. |
| Keywords:Lumbar vertebrae Intervertebral disk Cell culture techniques |
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