细胞信号转导抑制因子3影响青少年特发性脊柱侧凸畸形软骨内的成骨活性  

Effects of suppressor of cytokine signaling 3 on osteogenic activity in the cartilage of adolescent idiopathic scoliosis

作　　者：孙锦鹏 刘军[1] 白云峰 华峰 王浩然 郑宏瑞 吴涛[1] Sun Jinpeng;Liu Jun;Bai Yunfeng;Hua Feng;Wang Haoran;Zheng Hongrui;Wu Tao(Department of Orthopedics,the Second Affiliated Hospital of Nanjing Medical University,Nanjing 210011,Jiangsu Province,China)

机构地区：[1]南京医科大学第二附属医院骨科,江苏省南京市210011

出　　处：《中国组织工程研究》2022年第26期4160-4165,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金青年基金(81301523),项目负责人:吴涛;江苏省自然科学基金面上项目(BK20181499),项目负责人:吴涛。

摘　　要：背景:已有相关研究报道,瘦素与细胞信号转导抑制因子3(suppressor of cytokine signaling 3,SOCS3)表达异常在青少年特发性脊柱侧凸畸形发病中起重要作用。目的:探讨SOCS3是否通过瘦素信号传导通路调节软骨细胞内成骨活性。方法:①将人棘突软骨细胞分别以0(对照组),100μg/L瘦素刺激24 h,免疫组化染色观察Ⅱ型胶原表达。②将人棘突软骨细胞分9组培养,分别为空白对照组、转染pcDNA3.1-NC组、转染pcDNA3.1-SOCS3组、转染siRNA-NC组、转染siRNA-SOCS3组、瘦素+转染pcDNA3.1-NC组、瘦素+转染pcDNA3.1-SOCS3组、瘦素+转染siRNA-NC组、瘦素+转染siRNA-SOCS3组。置于培养箱中培养24 h后,实时荧光定量PCR法检测SOCS3 mRNA的表达量,MTT法检测细胞抑制率。结果与结论:①免疫组化染色显示,瘦素刺激组软骨细胞Ⅱ型胶原表达多于对照组。②实时荧光定量PCR检测显示,转染pcDNA3.1-SOCS3可显著提高软骨细胞SOCS3 mRNA表达量,转染siRNA-SOCS3可显著降低软骨细胞SOCS3 mRNA表达量,转染pcDNA3.1-NC、siRNA-NC对软骨细胞的SOCS3 mRNA表达量无影响;瘦素预刺激可提高转染pcDNA3.1-SOCS3、siRNA-SOCS3软骨细胞的SOCS3 mRNA表达量,对转染pcDNA3.1-NC、siRNA-NC软骨细胞的SOCS3 mRNA表达量无影响。MTT检测显示,转染pcDNA3.1-SOCS3可抑制软骨细胞内成骨活性,转染siRNA-SOCS3可增强软骨细胞内成骨活性;瘦素预刺激可提高转染pcDNA3.1-NC、pcDNA3.1-SOCS3、siRNA-NC、siRNA-SOCS3软骨细胞内成骨活性。③结果表明,瘦素刺激可提高软骨细胞内成骨活性,SOCS3表达上调可通过瘦素抵抗抑制软骨细胞内成骨活性。BACKGROUND:It has been reported that the abnormal expression of leptin and suppressor of cytokine signaling 3(SOCS3)plays an important role in the pathogenesis of adolescent idiopathic scoliosis.OBJECTIVE:To investigate whether SOCS3 regulates the osteogenic activity of chondrocytes through the leptin signaling pathway.METHODS:Human spinous chondrocytes were stimulated with 0(blank control group)and 100μg/L leptin for 24 hours,and immunohistochemical staining was used to detect the expression of type Ⅱ collagen.The human spinous chondrocytes were cultured in nine groups:a blank control group,a pcDNA3.1-NC transfected group,a pcDNA3.1-SOCS3 transfected group,a siRNA-NC transfected group,a siRNA-SOCS3 transfected group,a leptin+transfected pcDNA3.1-NC group,a leptin+transfected pcDNA3.1-SOCS3 group,a leptin+transfected siRNA-NC group,and a leptin+transfected siRNA-SOCS3 group.After being cultured in an incubator for 24 hours,real-time fluorescent quantitative PCR was used to detect the mRNA expression of SOCS3,and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to detect the cell inhibition rate.RESULTS AND CONCLUSION:Results of immunohistochemical staining showed that the expression of type Ⅱ collagen in chondrocytes s was higher in leptin-stimulated group than the blank control group.Results of real-time fluorescence quantitative PCR showed that transfection of pcDNA3.1-SOCS3 could significantly increase the mRNA expression of SOCS3 in chondrocytes,and transfection of siRNA-SOCS3 could significantly reduce the mRNA expression of SOCS3 in chondrocytes.In contrast,transfection of pcDNA3.1-NC and siRNA-NC had no effect on the mRNA expression of SOCS3 in chondrocytes.Leptin pre-stimulation could increase the mRNA expression of SOCS3 in chondrocytes transfected with pcDNA3.1-SOCS3 and siRNA-SOCS3,but it had no effect on the mRNA expression of SOCS3 in chondrocytes transfected with pcDNA3.1-NC and siRNA-NC.Results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay sho

关 键 词：瘦素 细胞信号转导抑制因子3 软骨细胞 青少年 脊柱侧凸畸形 质粒转染 

分 类 号：R446.8[医药卫生—诊断学] R322.71[医药卫生—临床医学]