Sonic Hedgehog通过上调Timp-3促进小鼠退行性膝关节炎的修复

刺猬因子（Hedgehog, Hh）信号通路广泛参与脊椎动物的胚胎发育和组织稳态的调控。已有报道认为，Hh参与骨性关节炎的发生和发展，但是其机制尚未详细阐述。本研究通过小鼠右后肢膝关节前交叉韧带切断手术，建立退行性膝骨关节炎小鼠模型，探索sonic hedgehog（Shh）因子在退行性膝关节炎中的作用及其机制。小鼠成功建模4周后，在模型组和假手术组小鼠膝关节腔中注射小鼠Shh-N蛋白因子，通过ELISA检测关节液中炎性因子IL-1β、TNF-α和解聚素金属4（a disintegrin and metalloprotease with thrombospondin motifs4，ADAMTS-4）蛋白酶的水平，以及通过qRT-PCR检测软骨组织中金属蛋白酶组织抑制剂-3（metalloproteinase inhibitor 3，Timp-3）基因的表达。结果显示，与假手术组相比，退行性膝骨关节炎模型小鼠骨关节腔内，IL-1β和TNF-α水平显著增高（24.5±5 vs 251.0±17.5 pg/mL，P < 0.001；16.5±3.4 vs 197.0±15.6 pg/mL, P < 0.001）；与PBS对照组相比， Shh-N组小鼠关节液中炎性因子的IL-1β和TNF-α水平明显降低（116.0±8.7 vs 251.0±17.5 pg/mL，P < 0.001； 89.0±7.8 vs 197.0±15.6 pg/mL, P < 0.001）。其中，小鼠的攀爬时间（1 700.0±185.6 vs 116±8.7 s，P < 0.001）和长距离运动能力均有显著改善（174.0±162.3 vs 132.0±34.5 m，P < 0.001）；qRT-PCR结果显示，软骨组织中Timp-3基因表达明显增高（5.0±0.3 vs 2.4±0.6 fold，P < 0.001）。同时，伴随着ADAMTS-4的水平显著降低（P < 0.001）。本研究提示，Shh因子显著改善小鼠退行性膝关节炎的恢复，其机制可能是通过上调Timp-3基因的表达，降解内源性聚蛋白多糖酶而实现的。

Sonic Hedgehog Promotes Recovery from Degenerative Knee Arthritis via Up-regulation of Timp-3 in Mice

Hedgehog (Hh) signaling pathway plays extensive and critical roles in embryonic development and tissue homeostasis in vertebrates. It has been reported that Hh is involved in the initiation and progress of osteoarthritis, however, the underlying mechanisms are still unclear. The purpose of our study is to investigate the role and mechanism of sonic hedgehog (Shh) factor in the model of degenerative knee arthritis induced by anterior cruciate ligament excision in mice. Four weeks after the degenerative knee osteoarthritis mouse model was established, Shh-N protein factor was injected into the knee joint cavity of the model group and sham-operated group. The levels of IL-1β, TNF-α and ADAMTS-4 in the synovial fluid were detected by ELISA, and the expression of Timp-3 in cartilage tissue was detected by qRT-PCR. The results indicated that compared with the sham operated group, the levels of IL-1β and TNF-α in the osteoarthritis lumen of degenerative knee osteoarthritis model mice were significantly increased (24.5 ± 5 vs 251.0 ± 17.5 pg/mL, P<0.001; 16.5 ± 3.4 vs 197.0 ± 15.6 pg/mL, P<0.001); as compared with the PBS control group, the symptoms of osteoarthritis in Shh-N group were significantly alleviated, and the levels of inflammatory factors IL-1β and TNF-α in the synovial fluid were significantly lower than those of the PBS control group (116.0 ± 8.7 vs 251.0 ± 17.5 pg/mL, P<0.001; 89.0 ± 7.8 vs 197.0±15.6 pg/mL, P<0.001), the climbing time and long-distance movement ability of mice were significantly improved (1 700.0 ± 185.6 vs 116.0 ± 8.7 s, P<0.001; 174.0 ± 162.3 vs 132.0 ± 34.5 m, P<0.001, respectively); the expression of metalloproteinase inhibitor 3 (Timp-3) gene in cartilage tissue was significantly increased by qRT-PCR analysis (5.0 ± 0.3 vs 2.4 ± 0.6 fold, P<0.001), and the A Disintegrin and Metalloprotease with Thrombospondin motifs 4 (ADAMTS-4) level decreased significantly (P < 0.001). The study indicated that Shh factor can significantly improve the recovery from degenerative knee arthritis in mice by up-regulation of Timp-3 gene expression and degradation of endogenous polyproteoglycan enzymes.