TRPV4抑制剂HC067047对小鼠膝骨关节炎软骨组织的影响

Effect of TRPV4 inhibitor HC067047 on cartilage tissue of knee osteoarthritis

目的探讨瞬时受体电位香草素受体4型通道蛋白(TRPV4)抑制剂HC067047对小鼠膝骨关节炎软骨组织的影响。方法将30只小鼠均分为假手术组、模型组、HC067047组。假手术组仅切开右侧膝关节髌韧带内侧皮肤,不予处理膝关节囊内结构;模型组、HC067047组手术切除小鼠板股韧带及内侧半月板前角以构建膝骨关节炎,分别用生理盐水、HC067047每天10 mg/kg灌胃。qRT-PCR检测各组小鼠软骨组织中TRPV4 mRNA表达水平。番红固绿染色评估关节软骨受损程度,HE染色分析膝关节软骨下骨的骨量变化。Western blotting检测各组软骨组织中细胞焦亡标志蛋白TRPV4、Caspase-1、核苷酸结合寡聚化结构域样受体家族热蛋白结构域蛋白3(NLRP3)、凋亡相关斑点样蛋白(ASC)、GSDMD-N蛋白水平;ELISA、Western blotting分别检测血清、软骨组织中炎症因子水平。结果与假手术组比较,模型组TRPV4、NLRP3、ASC、Caspase-1、GSDMD-N、IL-1β、IL-18水平显著升高(P<0.05)。与模型组比较,HC067047组NLRP3、ASC、Caspase-1、GSDMD-N、IL-1β、IL-18水平显著降低(P<0.05)。HC067047能明显改善关节软骨形态及降低骨关节炎受损程度,并能维持软骨下骨的骨量。结论TRPV4抑制剂HC067047能抑制软骨组织细胞焦亡,改善膝骨关节炎小鼠关节软骨的退变。

Aim To explore the effect of transient receptor potential vanilloid receptor 4(TRPV4)inhibitor HC067047 on osteoarthritis in Mice.Methods 30 mice were randomly divided into the sham group,the model group and HC067047 group.The sham group was only given an incision on the inner skin of the right knee joint patellar ligament without any treatment to the intra-articular structures.In contrast,the model group and the HC067047 group had their anterior cruciate ligaments and medial menisci anterior horns surgically removed to establish a knee osteoarthritis model.They were then administered either physiological saline or HC067047 at a daily dose of 10 mg/kg by oral gavage.qRT-PCR was used to measure the mRNA expression level of TRPV4 in the mice cartilage tissue from each group.Safranin solid green staining was employed to assess the extent of joint cartilage damage,while HE staining was used to analyze changes in subchondral bone mass in the knee joint.Western blotting was used to detect the protein levels of cell pyroptosis marker protein TRPV4,Caspase-1,nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3),apoptosis-associated speck-like protein(ASC),and GSDMD-N in the cartilage tissue of each group.ELISA and Western blotting were utilized to measure the levels of inflammatory factors in serum and cartilage tissue.Results Compared to the Sham group,the model group exhibited a significant increase in levels of TRPV4,NLRP3,ASC,Caspase-1,GSDMD-N,interleukin(IL)-1β,and IL-18(P<0.05).In comparison to the model group,the HC067047 group showed significant reductions in levels of NLRP3,ASC,Caspase-1,GSDMD-N,IL-1β,and IL-18(P<0.05).HC067047 group showed significantly improved joint cartilage morphology,reduced the extent of osteoarthritis damage,and maintained subchondral bone mass.Conclusion HC067047 can inhibit chondrocytes pyrolysis and ameliorate the articular cartilage degeneration in osteoarthritis mice.