| 师婕,张钰,许秀峰,胡丽芳,赵巧珍,陈琛,黄丹,李锦,陈焕,陈立早.探究早期兔膝骨关节炎中ERK信号通路对细胞自噬的作用[J].中国康复,2020,35(1):3-6 |
| 探究早期兔膝骨关节炎中ERK信号通路对细胞自噬的作用 |
| Effect of ERK inhibitor on autophagy at the early stage of osteoarthritis in rabbits |
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| DOI： |
| 中文关键词: 骨关节炎 自噬 ERK U0126 |
| 英文关键词: osteoarthritis autophagy extracellular regulated protein kinass U0126 |
| 基金项目:长沙市科技计划项目（kq1706012） |
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| 中文摘要: |
| 目的：探究细胞外调节蛋白激酶（ERK）抑制剂U0126对早期兔膝骨关节炎（OA）中细胞自噬的作用。方法：将24只雄性大白兔随机分为对照组、OA组和OA+U0126组各8例，对照组不接受任何干预措施，OA组和OA+U0126组均用木瓜蛋白酶行双膝骨关节炎造模3d，OA+U1026组于7d后将U0126注射到双膝关节腔，15d时处死兔。3组实验兔均进行大体观察，取双膝股骨内侧髁，通过甲苯胺蓝染色半定量分析糖胺聚糖（GAG）；根据苏木精-伊红（HE）和番红-固绿染色，采用国际骨关节炎研究学会（OARSI）总分评价OA严重程度；蛋白免疫印迹分析检测每组中磷酸化的EPK（P-ERK）、金属蛋白酶3（MMP3）、UNC-51样激酶1（ULK1）、自噬相关蛋白Beclin1和LC3II/LC3I的含量。结果：3组白兔实验前后的体重以及体重的变化各组两两比较差异无统计学意义。对照组软骨表面光滑较薄，色泽明亮，未见裂隙，触之较硬，关节滑膜未见增生和水肿等，关节液无渗出；OA+U0126组与对照组比较，软骨色泽稍暗，膝关节软骨表面稍增厚，关节内滑液明显增多；OA组软骨增厚变粗糙最明显，滑液最多。OA组明显可见软骨表面增厚出现裂隙，软骨细胞增生，细胞基质失染以及细胞排列紊乱和簇集等典型骨关节炎改变；OA+U0126组软骨增厚、细胞增生排列紊乱等改变比OA组轻。实验后，OA+U0126组和OA组GAG含量均低于对照组(均P＜0.05)；OA+U0126与OA组比较，GAG含量升高（P＜0.01）。OA+U0126组和OA组OARSI评分均明显高于对照组(均P＜0.05)；OA+U0126组与OA组比较，OARSI评分降低（P＜0.01）。OA+U0126组和OA组MMP3、P-ERK含量较对照组均升高（均P＜0.05），LC3II/LC3I、ULK1和Beclin1含量较对照组均降低(均P＜0.05)。OA+U0126组与OA组比较，P-ERK、MMP3含量均降低（均P＜0.05），LC3II/LC3I、ULK1和Beclin1含量均升高(均P＜0.05)。结论：U0126通过减少MMP3促进细胞自噬，延缓骨关节炎的进程。 |
| 英文摘要: |
| Objective: To explore the effect of extracellular signal regulated kinase (ERK) inhibitor U0126 on autophagy at the early stage of osteoarthritis in rabbits. Methods: We randomly and equally assigned 24 male rabbits into control group, OA group and OA+U0126 group. In control group rabbits were not given any treatments; in OA group and OA+U0126 group, rabbits were treated with intra articular injection of papain into the knees for 3 days. Then we conducted intra articular injection of U0126 into the rabbits in OA+U0126 group after 7 days. Fifteen days later, all rabbits were sacrificed for general observation of the knees. Medial condyle cartilages were excised for histological observation and Western blotting. Semiquantitative analysis of glycosaminoglycan (GAG) was performed according to toluidine blue staining. The OARSI scoring system was introduced to assess the severity and extent of OA by HE staining and Safranin O and Fast green staining. We also analyzed the levels of phosphorylated ERK (P-ERK), matrix metalloproteinase 3 (MMP3), UNC-51 like kinase 1 (ULK1), autophagy related protein Beclin1 and LC3II/I by Western blotting. Results: The average weights of rabbits in each group before the study and at the execution had no significant difference. Compared to the control group whose cartilages showed a thin, light, hardened and smooth surface without fissures, proliferation and edema in synovium or exudation of joint fluid, OA+ U0126 group exhibited a slight dark and thicken surface with an increase of joint fluid. Representative OA changes could be seen in OA group, which was characterized by thick and tough cartilage surface, cell disorientation and clustering tendency, as well as a loss of dying of matrix and proliferation of chondrocytes. However, these changes were attenuated in OA+U0126 group compared to OA group. Contents of GAG in OA group and OA+U0126 group were significantly lower than in the control group (P＜0.05), and GAG content in OA+U0126 group was significantly higher than in OA group (P<0.01). OARSI scores in OA group and OA+U0126 group were significantly higher than in the control group (P<0.05), and those in OA+U0126 group were significantly lower than in OA group (P<0.01). Compared to the control group, levels of MMP3 and P-ERK were increased, while LC3II/I, ULK1 and Beclin1 were decreased in OA group and OA+U0126 group (P<0.05). In contrast to OA group, OA+U0126 group had a lower level of MMP3 and P-ERK and a higher level of LC3II/I, ULK1 and Beclin1 (P<0.05). Conclusions: Our study demonstrated that U0126, an ERK inhibitor, could relieve the OA process by reducing MMP3 and promoting autophagy. |
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