曲美他嗪对大鼠骨关节炎炎症反应和软骨基质降解的作用研究

Effects of trimetazidine on inflammation and cartilage matrix degradation in rats with osteo⁃arthritis

目的:探讨曲美他嗪对大鼠骨关节炎(OA)炎症反应和软骨基质降解作用。方法:将软骨细胞分为对照组、模型组和实验组。对照组加入单纯培养基培养,模型组加入含有10μg/mL IL-1β培养基培养,实验组加入含有10μg/mL IL-1β+0.1 mg/mL曲美他嗪的培养基培养。通过细胞计数试剂(CCK-8)法和钙黄绿素/碘化丙啶(Calcein-AM/PI)染色法检测曲美他嗪的毒性作用;实时荧光定量PCR(RT-qPCR)检测软骨形成和炎症相关基因的表达;将大鼠随机为对照组、模型组和实验组,除对照组外,模型组和实验组采用前交叉韧带切断手术建立大鼠OA模型,实验组大鼠关节腔内注射100μL 0.1 mg/mL曲美他嗪,模型组和对照组分别注射等量的0.9%NaCl溶液。处理4周后,对各组关节进行组织学染色及评分。结果:CCK-8结果显示,与0 mg/mL比较,曲美他嗪浓度低于0.1 mg/mL时,对软骨细胞无明显毒性(P>0.05);Calcein-AM/PI染色结果显示,与模型组比较,实验组活细胞数量增加,死细胞数量减少,细胞活力提高(P<0.01);RT-qPCR结果显示,与模型组比较,实验组的性别决定区域Y相关的高迁移率族框9(SOX9)和蛋白聚糖(ACAN)相对表达量均升高(均P<0.05),白介素-6(IL-6)、基质金属蛋白酶-3(MMP-3)、基质金属蛋白酶-13(MMP-13)、环氧酶-2(COX-2)及肿瘤坏死因子-α(TNF-α)表达均降低(均P<0.05)。相较于模型组,实验组软骨层的损伤与变形明显缓解;与模型组相比,实验组的组织学评分降低(P<0.05)。结论:曲美他嗪具有减轻OA炎症反应和保护软骨的作用。

Objective:To investigate the effects of trimetazidine on inflammation and cartilage matrix degradation in rats with osteoarthritis(OA).Methods:Chondrocytes were divided into control group,model group and experimental group.The control group was cultured with simple medium,the model group was cultured with 10μg/mL IL-1βmedium,and the experimental group was cultured with 10μg/mL IL-1β+0.1 mg/mL trimetazidine.The toxicity of trimetazidine was detected by CCK-8 method and Calcein/propidium iodide(Calcein-AM/PI)staining,and the expression of genes related to chondrogenesis and inflammation was detected by RT-qPCR.Rats were randomly divided into control group,model group and experimental group.Except for control group,OA model was established by anterior cruciate ligament transection in model group and experimental group.100μL 0.1 mg/mL trimetazidine was injected into articular cavity of rats in experimental group,and the same amount of 0.9%NaCl solution was injected into model group and control group.After 4 weeks of treatment,the joints of each group were stained and scored.Results:The results of CCK-8 showed that trimetazidine had no obvious toxicity to chondrocytes when the concentration of trimetazidine was lower than 0.1 mg/mL(P>0.05).Compared with model group,Calcein-AM/PI staining showed that the number of living cells increased,the number of dead cells decreased and the cell viability increased in the experimental group(P<0.01).Compared with model group,the mRNA expressions of high mobility group frame 9(SOX9)and proteoglycan(ACAN)related to gender determination region Y in the experimental group were higher than those in the model group(P<0.05).The expressions of interleukin-6(IL-6),matrix metalloproteinase-3(MMP-3),matrix metalloproteinase-13(MMP-13),cyclooxygenase-2(COX-2)and tumor necrosis factor-α(TNF-α)were decreased(P<0.05).Compared with the model group,the injury and deformation of the cartilage layer in the experimental group were alleviated,and the histological score in the experimental group was lower(P<0.05).Conclusion:Trimetazidine can reduce the inflammatory reaction of OA and protect cartilage.