Department of Bone Traumatology, Affiliated Hospital of Hunan Academy of Chinese Medicine, Changsha, Hunan410006, China.
Cell Mol Biol (Noisy-le-grand). 2022 Feb 27;67(6):236-241. doi: 10.14715/cmb/2021.67.6.31.
This study aimed to investigate the regulatory mechanism of MDM2 gene expression on cartilage cell proliferation in Osteoarthritis (OA) rats. For this purpose, 22 SD rats were randomly divided into normal control (10 cases) and treated (12 cases) groups. Treated group was used for OA modelling with the modified Hulth method. After a week, RT-PCR was used to detect MDM2 in cartilage tissue of rats, Wnt 1, Wnt 3 a, Wnt 10 b and β-catenin genes mRNA expression. Rat chondrocytes were isolated and cultured, and the recombinant eukaryotic expression vector pcDNA3.1 myc-siRNA-MDM2-β-catenin and co-expression plasmid pcDNA3.1 myc-siRNA-MDM2-β-catenin was used to transfect chondrocytes and the proliferation and related gene expression levels of the transfected chondrocytes were detected by MTT method and RT-PCR. The results showed that compared with the control group, MDM2, Wnt 1, Wnt 3 a, Wnt 10b and β-catenin genes in OA rat cartilage constructed by Hulth method were increased (p<0.05). The pcDNA3.1 myc-beta-catenin transfection slowed down the proliferation of OA chondrocytes, different from the non-transfected OA group (p<0.001), and increased Wnt 1, Wnt 3a, Wnt 10b and β-catenin genes expression compared with the Control group (p<0.05), but did not affect the expression of MDM2. The transfection of siRNA-MDM2 was opposite to pcDNA3.1 myc-β-catenin. The co-expression plasmid pcDNA3.1 myc-siRNA-MDM2-beta-catenin transfection did not affect the proliferation of OA chondrocytes. In general, the high expression of MDM2 in OA rats restricts the proliferation of chondrocytes, which may be related to the main pathogenesis of the occurrence and development of OA in vivo, and the regulation of MDM2 on the proliferation of chondrocytes may be achieved through the Wnt/ β-catenin pathway.
本研究旨在探讨 MDM2 基因表达对骨关节炎(OA)大鼠软骨细胞增殖的调控机制。为此,将 22 只 SD 大鼠随机分为正常对照组(10 例)和治疗组(12 例)。治疗组采用改良 Hulth 法建立 OA 模型。1 周后,采用 RT-PCR 检测大鼠软骨组织中 MDM2、Wnt1、Wnt3a、Wnt10b 和β-catenin 基因 mRNA 表达。分离培养大鼠软骨细胞,采用重组真核表达载体 pcDNA3.1 myc-siRNA-MDM2-β-catenin 及其共表达质粒 pcDNA3.1 myc-siRNA-MDM2-β-catenin 转染软骨细胞,MTT 法和 RT-PCR 检测转染软骨细胞的增殖及相关基因表达水平。结果表明,与对照组比较,Hulth 法构建的 OA 大鼠软骨中 MDM2、Wnt1、Wnt3a、Wnt10b 和β-catenin 基因表达均升高(P<0.05)。pcDNA3.1 myc-β-catenin 转染后,OA 软骨细胞增殖速度较未转染 OA 组减慢(P<0.001),与对照组比较,Wnt1、Wnt3a、Wnt10b 和β-catenin 基因表达均升高(P<0.05),但对 MDM2 基因表达无影响。siRNA-MDM2 转染则与 pcDNA3.1 myc-β-catenin 相反。pcDNA3.1 myc-siRNA-MDM2-β-catenin 共表达质粒转染对 OA 软骨细胞增殖无影响。综上所述,OA 大鼠 MDM2 高表达抑制软骨细胞增殖,可能与体内 OA 发生发展的主要发病机制有关,MDM2 对软骨细胞增殖的调控可能通过 Wnt/β-catenin 通路实现。