基质金属蛋白酶9对骨肉瘤增殖、侵袭及迁移能力的影响及其临床意义

Effect of down-regulating matrix metalloproteinase 9 expression on the progression of osteosarcoma and its clinical significance

摘要:

目的：通过研究骨肉瘤组织中基质金属蛋白酶9(MMP-9)的表达情况，及沉默MMP-9后对骨肉瘤细胞增殖、侵袭及迁移能力的影响，阐明MMP-9对骨肉瘤进展的影响及其临床意义。 方法：通过定量PCR分别检测5例原发性骨肉瘤及5例正常骨组织标本中MMP家族的表达情况；通过慢病毒载体介导shRNA-MMP-9转染至骨肉瘤细胞Saos2中，将实验分为：空白对照组(磷酸盐缓冲液，PBS)、阴性对照组(转染shRNA-NC序列)及转染组(shRNA-MMP-9序列)，检测MMP-9沉默后Saos2细胞增殖、侵袭及迁移能力的变化；根据MMP-9在患者骨肉瘤标本中的表达水平，对64例随访病例分组，进行生存分析。 结果：筛查MMP家族基因，检测发现骨肉瘤组织中MMP-9的表达水平显著高于正常骨组织(t=12.040, P=0.000)。骨肉瘤细胞Saos2转染shRNA-MMP-9后，转染组细胞MMP-9 mRNA及蛋白表达水平均显著下降，提示在Saos2细胞中成功沉默MMP-9。在细胞增殖、侵袭及划痕实验中，空白对照组与阴性对照组均无显著差异，但转染组的细胞相较于阴性对照组，其增殖、侵袭及迁移能力均显著减弱(t=4.954, P=0.008; t=6.360, P=0.003; t=3.965, P=0.017)。此外，对64例随访病例进行生存分析发现，MMP-9高表达组患者的总生存率显著低于MMP-9低表达组( χ2=4.169, P=0.041)。结论：骨肉瘤组织中高表达MMP-9，其与骨肉瘤的增殖、侵袭、转移密切相关，且对骨肉瘤患者的临床预后具有重要提示作用。

Abstract:

Objective: To investigate the expression of matrix metalloproteinases 9 (MMP-9) in osteosarcoma tissues and the effect of MMP-9 on the progression of osteosarcoma and its clinical significance. Methods: The expression of MMPs’ family in 5 osteosarcoma specimens and 5 normal bone tissues was detected by real-time PCR. shRNA was used to knockdown the expression of MMP-9 in Saos2 cells. The proliferation, invasion and migration of the cells after transfection were detected for the effects of MMP-9 down-regulation. According to the expression of MMP-9 in osteosarcoma, 64 follow-up cases were grouped and analyzed for survival rate. Results: Screening of MMPs’ family revealed that the expression of MMP-9 in osteosarcoma tissues was significantly higher than that in normal bone tissues (t=12.040, P=0.000). In cell proliferation, invasion and scratch test, there was no significant difference between blank control group and negative control group. But compared with negative control group, down-regulation of MMP-9 in Saos2 cells markedly repressed cell proliferation，decreased invasion and migration (t=4.954, P=0.008; t=6.360, P=0.003; t=3.965, P=0.017) . In addition, a survival analysis in 64 follow-up cases showed that the overall survival rate of the high-MMP-9 group was significantly lower than that of the low-MMP-9 group (χ2=4.169，P=0.041). Conclusion: The high expression of MMP-9 in osteosarcoma tissue is closely related to the proliferation, invasion and migration of osteosarcoma, which has certain implication for the clinical prognosis of patients.