马钱子碱治疗早期小鼠膝骨关节炎的作用机制研究

Study on the Mechanism of Brucine in Treating Early Knee Osteoarthritis in Mice

目的研究马钱子碱治疗早期膝骨关节炎(KOA)模型小鼠的作用机制。方法随机将24只C57BL/6雄性小鼠分为4组:空白对照组(不采取任何干预措施)、模型组(构建KOA动物模型+腹腔注射等体积的生理盐水)、马钱子碱中剂量组(构建KOA动物模型+腹腔注射马钱子碱溶液,5 mg/kg)、马钱子碱高剂量组(构建KOA动物模型+腹腔注射马钱子碱溶液,10 mg/kg),药物干预4周后,取小鼠关节软骨。采用免疫荧光染色技术检测4组小鼠软骨细胞内Zn^(^(2+))表达,采用RT-qPCR和蛋白质印迹技术分别检测4组小鼠软骨ZIP8、ZNT9、MTF1、Mt1、Mt2 mRNA和蛋白表达。结果与空白对照组比较,模型组ZIP8、MTF1 mRNA和蛋白相对表达量明显增加(P<0.05)。与模型组相比较,马钱子碱高、中剂量组ZIP8、MTF1 mRNA和蛋白表达明显降低(P<0.05)。与模型组相比较,马钱子碱高、中剂量组ZNT9、Mt1、Mt2 mRNA和蛋白的相对表达量差异无统计学意义(P>0.05)。荧光染色结果显示,与空白对照组相比,模型组软骨细胞内Zn^(2+)表达明显增加,经马钱子碱药物干预后,马钱子碱高、中剂量组小鼠软骨细胞内Zn^(2+)表达明显降低,且马钱子碱高剂量组降低更明显(P<0.05)。结论马钱子碱通过降低KOA小鼠ZIP8、MTF1 mRNA和蛋白表达来调控软骨细胞内Zn^(2+)表达量,从而延缓早期KOA模型小鼠软骨退变。

Objective To study the mechanism of brucine in the treatment of early knee osteoarthritis(KOA)in mouse model.Methods A total of 24 male C57BL/6 mice were randomly divided into four groups blank control group(without any intervention),model group(KOA mouse model+intraperitoneal injection of equal volume of normal saline),brucine medium-dose group(KOA mouse model+intraperitoneal injection of brucine solution,5 mg/kg),brucine high-dose group(KOA mouse model+intraperitoneal injection of brucine solution,10 mg/kg).After 4 weeks of administration,the articular cartilages of knee joint of mice were collected.The expression of Zn^(2+)in the chondrocytes of the 4 groups of mice was detected by fluorescence immunoassay.And the mRNA and protein expressions of ZIP8,ZNT9,MTF1,Mt1 and Mt2 in the chondrocytes of the 4 groups of mice were detected by RT-qPCR(real-time quantitative polymerasechain reaction)and Western blot.Results Compared with blank control group,the relative expression levels of mRNA and protein of ZIP8 and MTF1 in model group significantly increased(P<0.05).Compared with model group,the relative expression levels of mRNA and protein of ZIP8 and MTF1 in brucine high-dose and medium-dose groups significantly decreased(P<0.05).Compared with model group,the relative expression levels of mRNA and protein of ZNT9,Mt1 and Mt2 in brucine high-dose and medium-dose groups showed no significant difference(P>0.05).Fluorescence staining results showed that,compared with blank control group,the expression of Zn^(2+)in chondrocytes of model group significantly increased,and that of brucine high-dose and medium-dose groups significantly decreased after brucine intervention,and the decrease was more obvious in brucine high-dose group(P<0.05).Conclusion Brucine can regulate the expression of Zn^(2+)in chondrocytes by reducing the expression levels of mRNA and protein of ZIP8 and MTF1 in KOA mouse model,so as to delay the cartilage degeneration in early KOA mouse model.