兔髓核组织中BNIP3表达与椎间盘退变的关系

Preliminary research for correlation between BNIP3 expression and disc degeneration

中文关键词:  椎间盘退变  髓核  BNIP3  兔

英文关键词:Disc degeneration  Nucleus pulposus  BNIP3  Rabbit

中文摘要:

  【摘要】 目的：探讨兔椎间盘髓核组织中BNIP3表达量与椎间盘退变的相关性。方法：健康1月龄新西兰白兔24只，随机分为4组，每组6只。在每只动物的L4/5、L5/6和L6/7椎间盘进行纤维环穿刺术，建立椎间盘退变模型，作为实验椎间盘；穿刺椎间盘近端3个椎间盘（L1/2、L2/3、L3/4）作为正常对照椎间盘。分别于术后即刻、2、4、8周对椎间盘进行MRI及组织学评分，应用Real-time PCR定量检测髓核组织BNIP3 mRNA表达，免疫组织化学染色半定量髓核组织BNIP3表达，同时分析同一椎间盘BNIP3表达与MRI评分、组织学评分之间的相关性，并与正常椎间盘进行对照。结果：正常及手术后即刻实验椎间盘在MRI T2加权像上呈高密度，评分为1.0±0.0；手术后2周实验椎间盘信号呈不均一的高密度，评分为2.9±0.4；手术后4周实验椎间盘信号呈不均一中低密度，评分4.2±0.3；手术后8周实验椎间盘信号呈低密度，评分4.9±0.1，各时间点MRI评分比较有显著性差异（P<0.05）。组织学染色显示正常及手术后即刻椎间盘结构整齐，髓核与纤维环交界清晰，组织学评分4.0±0.0；手术后2周实验椎间盘纤维环出现小裂隙，髓核与纤维环交界轻度不清，组织学评分7.5±0.2；手术后4周实验椎间盘纤维环出现较大裂隙，髓核与纤维环交界中度不清，组织学评分10.0±1.0；手术后8周实验椎间盘正常结构丧失，髓核组织纤维化，组织学评分11.8±0.2，各时间点间组织学评分比较有显著性差异（P<0.05）。手术后即刻、2周、4周及8周实验椎间盘BNIP3 mRNA表达是正常组的1.0±0.3倍、2.0±0.1倍、2.8±0.3倍和4.2±0.2倍，与椎间盘MRI退变评分呈显著性正相关（r=0.92，P<0.01）。正常及手术后即刻实验椎间盘髓核组织BNIP3灰度值分别为55.3±6.2和60.7±4.4；而手术后2周、4周及8周实验椎间盘分别为150.4±13.4、176.0±35.1和173.6±7.9，其灰度值与椎间盘组织学评分呈显著性正相关（r=0.92，P<0.01）。结论：兔椎间盘髓核组织中BNIP3 mRNA表达量与椎间盘退变相关，BNIP3表达上调可能在椎间盘退变过程中发挥了重要作用。

英文摘要:

  【Abstract】 Objective：To study the correlation between BNIP3 expression and disc degeneration.Method：Twenty-four one-month white New Zealand rabbits were randomly and equally assigned into four groups：post-surgery for 0 week（n=6），post-surgery for 2 weeks（n=6），post-surgery for 4 weeks（n=6），and post-surgery for 8 weeks（n=6）.By puncturing the experimental disc annulus （L4/5，L5/6 and L6/7） with needle，we established a disc degeneration model in New Zealand white rabbit，the normal discs above the experimental discs（L1/2，L2/3 and L3/4） served as control.After the surgery for 0，2，4 and 8 weeks，the disc degeneration，BNIP3 expression and cell morphology were evaluated by using MRI，histochemistry staining，immunohistochemical staining and real-time-PCR，and the correlationship between the BNIP3 expression and the disc degeneration evaluated by MRI and histochemistry was also been analyzed.Result：The MRI showed that the normal discs and discs of post surgery for 0 week exhibited hyperintense in T2 weighting image with the average MRI grade of 1.0±0.0；the discs of post surgery for 2 weeks showed inhomogeneous hyperintense with the average MRI grade of 2.9±0.4；the discs of post surgery for 4 weeks showed intermediate to hypointense with the average MRI grade of 4.2±0.3；the discs of post surgery for 8 weeks showed a hypointense which the average MRI grade of 4.9±0.1.There were significant differences among normal discs and post surgery discs（P<0.05）.The histochemistry staining also showed the normal discs and discs of post surgery for 0 week with intact structure and clear border between the annulus fibrosus and nucleus pulposus，the average histological grade was 4.0±0.0；the discs of post surgery for 2 weeks showed slight rupture in annulus fibrosus and minimally interrupted between annulus fibrosus and nucleus pulposus，the average histological grade was 7.5±0.2；the discs of post surgery for 4 weeks showed larger rupture in annulus fibrosus and moderate rupture between annulus fibrosus and nucleus pulposus，the average histological grade was 10.0±1.0；the discs of post surgery for 8 weeks showed the collapse of disc structure and fibrosis of nucleus pulposus，the average histological grade was 11.8±0.2.There were significant differences among normal discs and post surgery discs（P<0.05）.The BNIP3 mRNA real-time-PCR of the nucleus pulposus region indicated increased expression of BNIP3 with 1.0±0.3-fold，2.0±0.1-fold，2.8±0.3-fold and 4.2±0.2-fold respectively at 0，2，4 and 8 weeks after surgery，and there was a high correlation between MRI grade and BNIP3 mRNA expression（r=0.92，P<0.01）.The BNIP3 immunohistochemical staining of the nucleus pulposus region suggested increased positive reaction with 150.4±13.4，176.0±35.1 and 173.6±7.9 of gray intensity at 2，4 and 8 weeks after surgery，which had a high correlation with histological grade（r=0.92，P<0.01）.Conclusion：The expression of BNIP3 mRNA has some relationship with the disc degeneration，and the increased expression of BNIP3 may play an important role in disc degeneration.