正常与退变髓核细胞形态学及Ⅱ型胶原蛋白的定量分析  

Morphology of normal and degenerative nucleus pulposus cells and quantitative analysis of collagen type II protein

作　　者：王祺[1] 任龙韬[1] 卫成刚 何君仁[1] 

机构地区：[1]山西医科大学附属太原市中心医院骨科,山西省太原市030009 [2]秦皇岛海港医院,河北省秦皇岛市066000

出　　处：《中国组织工程研究》2014年第15期2363-2368,共6页Chinese Journal of Tissue Engineering Research

摘　　要：背景:椎间盘退变导致椎间隙狭窄的主要变化是髓核细胞表达软骨特异性基因产物蛋白聚糖和Ⅱ型胶原的减少。目的:对成人的正常与退变椎间盘髓核细胞Ⅱ型胶原蛋白免疫荧光染色及番红O染色进行定量观察比较。方法:取成人脊柱侧弯患者与椎间盘突出症患者自愿者术中取出的废弃髓核组织各3例,经培养后每个患者各测量26个细胞,正常组和退变组分别测量78个细胞。对正常与退变椎间盘髓核细胞进行番红"O"染色并行灰度及测定细胞内Ⅱ型胶原蛋白行免疫荧光染色定量测定。结果与结论:免疫荧光染色显示:退变椎间盘髓核细胞仅轻度染色,染色模糊,其呈类圆形,纺锤形,梭形及不规则形,其内仅有极少量荧光颗粒;Ⅱ型胶原表达较正常髓核细胞降低显著(P<0.05)。番红O染色显示:退变的髓核细胞肿胀,细胞核肿胀染色略淡,细胞突起延长,胞浆染色不匀伴有空泡,部分细胞崩解,细胞分布零散,混乱,可见成片坏死区。与正常髓核细胞图像灰度值相比差异无显著性意义(P>0.05)。结果表明退变的椎间盘髓核细胞减少、部分凋亡,其细胞内Ⅱ型胶原蛋白含量较正常髓核细胞显著减少。BACKGROUND：The narrowing of intervertebral space induced by the intervertebral disc degeneration is mainly characterized by the expression of proteoglycan in nucleus pulposus cells and the reduction of col agen type Ⅱ. OBJECTIVE：To quantitatively observe col agen type Ⅱ protein in adult normal and degenerative intervertebral disc nucleus pulposus cells by immunofluorescence staining and safranin O staining. METHODS：The nucleus pulposus specimens were col ected from adult scoliosis patients and patients with intervertebral disc protrusion, who were al volunteers. After culture, 26 cells in each patient were measured. There were 78 cells in both normal group and degeneration group. The normal and degenerative intervertebral disc nucleus pulposus cells were subjected to safranin“O”staining, and gray values were determined;intracellular col agen type Ⅱ was detected by immunofluorescence staining. RESULTS AND CONCLUSION：Immunofluorescence staining revealed that, degenerative intervertebral disc nucleus pulposus cells were only mildly stained, with the fuzzy staining, the shape was round, spindle, fusiform and irregular. There were a very smal amount of fluorescent particles within cells. The expression of col agen type Ⅱ was decreased significantly compared with normal cells （P〈0.05）. Safranin O staining showed that, degenerative nucleus pulposus cells began to swel , the nuclei swel ed and were stained slightly, cellprocesses were prolonged, cytoplasmic dyeing was uneven accompanying with vacuole, celldisruption, scattered and chaotic distribution were visible, patches of necrosis were observed. The image gray value showed no significant difference compared with normal nucleus pulposus cells （P〉0.05）. The degenerative intervertebral disc nucleus pulposus cells have a smal quantity and partial y become apoptotic, the content of col egen type Ⅱ protein is decreased significantly compared with normal nucleus pulposus cells.

关 键 词：组织构建 骨组织工程 椎间盘 髓核细胞 Ⅱ型胶原 番红O染色 

分 类 号：R318[医药卫生—生物医学工程]