兔退变椎间盘纤维环及髓核组织中的PG、MMP-3、TIMP-2表达变化及意义

Expression changes and meaning of PG,MMP-3,TIMP-2 in rabbit degenerated intervertebral disc fibrous ring and nucleus pulposus tissues

目的：观察兔退变椎间盘纤维环和髓核组织中蛋白多糖（PG）、基质金属蛋白酶-3（MMP-3）、基质金属蛋白酶组织抑制因子-2（TIMP-2）的表达，并探讨其意义。方法新西兰大白兔42只，直立位下建立椎间盘退变模型，均分为实验组和对照组。两组分别于建模后4、8、12周各取7只动物，取 L4～5椎间盘组织采用高碘酸-希夫（PAS）染色检测髓核与纤维环组织中的 PG，以灰度值表示其相对表达量；免疫组化 SP法检测椎间盘纤维环和髓核组织中的 MMP-3及 TIMP-2，以平均光密度值（MOD）表示其相对表达量。结果实验组、对照组建模4、8、12周髓核组织中 PG灰度值均高于纤维环组织（P＜0．05）；两组建模12周时纤维环、髓核PG灰度值均高于8周时，且建模8周时髓核 PG灰度值高于4周时（P＜0．05）。随着建模时间的延长，实验组中MMP-3和TIMP-2的相对表达量均增加明显。髓核中，对照组及实验组同一时间点 MMP-3的平均 MOD值明显高于 TIMP-2。实验组纤维环中TIMP-2较 MMP-3表达变化明显。结论椎间盘退变过程中髓核中PG的含量变化早于纤维环，通过对 MMP-3/TIMP-2在椎间盘退变中所起作用的研究结合髓核 PG的变化可能会对临床诊断、基因治疗和组织工程技术的应用有一定帮助。而且对 MMPs和TIMPs的表达水平的调节来控制椎间盘基质代谢的平衡，可能会成为治疗椎间盘退变的新方向。

Objective To observe the expression of proteoglycan （PG）,MMP-3,TIMP-2 in the rabbit degenerative interverte-bral disc fibrous ring and nucleus pulposus tissues and to investigate their significance.Methods 42 New Zealand white rabbits were used for establishing the intervertebral disc degeneration model under the upright position and equally divided into the experi-mental group and the control group.The L4-5 intervertebral disc tissues were taken in 7 rabbits per group at 4,8,12 weeks after modeling.The periodate Schiff （PAS）staining was adopted to detect PG in the nucleus pulposus and fibrous ring tissues with the grayscale value for expressing the relative expression quantity;the immunohistochemical SP method was adopted to detect MMPs-3 and TIMP-2 in the disc fibrous ring and nucleus pulposus tissues,with the mean optical density（MOD）value for expressing the relative expression quantity.Results The PG gray values in the nucleus pulposus tissue after 4,8,12 weeks in the experimental group and the control group model were higher than those in the fiber ring tissue （P〈0.05）;the PG gray values in the nucleus pulposus and fiber ring tissue at 1 2 weeks were higher than those at 8 weeks,and at 8 weeks of modeling,the PG gray value in the nucleus pulposus was higher than that at 4 weeks （P〈0.05）.As the time extension of modeling,the relative expression amounts of MMP-3 and TIMP-2 in the experimental group were significantly increased.The average MOD value of MMP-3 in the nucleus pul-posus at the same time point in the control group and the experimental group was obviously higher than that of TIMP-2.The ex-pression of TIMP-2 in the fibrous ring in the experimental group had more change than the expression of MMP-3.Conclusion Dur-ing the intervertebral disc degeneration process,the change of PG content in the nucleus pulposus is earlier than that of the fibrous ring.The study of the MMP3/TIMP-2 role in the intervertebral disc degeneration in combination with the changes of nucleus pulpo-sus PG may conduce to clinical diagnosis,gene therapy and the application of tissue engineering technique.Moreover regulating the expression level of MMPs and TIMPs for controlling the balance of intervertebral disc matrix metabolism could become the new di-rection for treating the intervertebral disc degeneration.