IGF-1信号通路相关蛋白在大鼠髁突软骨细胞增殖分化中的表达  

Proliferation and Differentiation of Mandibular Condylar Chondrocytes Require IGF-1 Signal Pathway Related-proteins

作　　者：江莉婷[1] 魏立[2] 谢银银[3] 周琦[2] 朱雅萍[2] 高益鸣[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院口腔科,上海200025 [2]上海市伤骨科研究所,上海200025 [3]上海交通大学医学院附属瑞金医院上海血液学研究所,医学基因组学国家重点实验室,上海200025

出　　处：《口腔颌面外科杂志》2013年第3期171-177,共7页Journal of Oral and Maxillofacial Surgery

基　　金：上海市科学技术委员会医学引导项目(114119a3500)

摘　　要：目的:研究胰岛素样生长因子-1(IGF-1)信号转导通路在大鼠髁突软骨细胞增殖分化调控过程中相关基因的表达。方法:体外单层培养并鉴定出生后1、7、14、28 d共4组SD大鼠髁突软骨细胞。免疫组织化学方法检测细胞内IGF-1表达情况,Real-time PCR及Western印迹法检测细胞内IGF-1R、Bcl-2、Bax mRNA及蛋白表达。饥饿培养24 h后,实验组加入质量浓度为100 ng/mL的重组大鼠IGF-1细胞因子(rrIGF-1)继续培养24、48 h,CCK-8检测细胞增殖情况,Real-time PCR技术检测各组髁突软骨细胞内IGF-1R、IGF-2R、Raf1、GSK-3、IGFBP3、NF-κB、Bcl-2、Bax、integrin、TGF-βmRNA表达情况;对照组培养液不加rrIGF-1。结果:各鼠龄组SD大鼠髁突软骨细胞内IGF-1表达均阳性,IGF-1R mRNA及蛋白表达相对平稳,Bcl-2、Bax mRNA及蛋白表达逐渐增强,在出生14 d后表达下降(P<0.05)。加入100 ng/mL rrIGF-1后,髁突软骨细胞增殖速度显著增加,细胞凋亡减少,Real-time PCR结果显示实验组IGF-1R、GSK-3、Bax、integrin mRNA表达整体呈下降趋势;IGF-2R、Raf1、IGFBP3、NF-κB、Bcl-2、TGF-βmRNA表达水平总体呈上调趋势,差异有统计学意义。结论 :IGF-1介导的信号转导途径参与了髁突软骨细胞的增殖、分化以及软骨形成早期的调控,并可能与integrin、TGF-β等其他蛋白相互作用,共同参与髁突发育过程。Objective： To investigate the insulin-like growth factor-1 （IGF-1） signaling pathway related gene expressions on regulating proliferation and differentiation of rat condylar chondrocytes. Methods： The 1, 7, 14 and 28 day-old SD rat condylar chondrocytes were cultured and identified in vitro, and IGF-1 expression of chondroeytes in each group were evaluated by immunohistochemistry. The gene and protein expression of IGF-1R,Bcl-2 ,Bax were investigated by Real-time PCR and Western blot analysis. After being starved for 24 hours, chondrocytes were then incubated with 100ng/mL recombinant rat IGF-1 （rrIGF-1） in serum-free medium for 24 and 48 hours,cell proliferation was measured by Cell Counting Kit-8 （CCK-8） method. Real-time PCR was performed by using IGF-1R, IGF-2R, Rafl, GSK-3, IGFBP3, NF-κB, Bcl-2, Bax, integrin, TGF-β specific primers to detect their gene expressions. Otherwise, the chondrocytes of negativecontrol groups were incubated without rrIGF-1. Results： The immunohistochemical staining of IGF-1 was positive in 1-,7-, 14-,28-day-old SD rat condylar chondrocytes. IGF-1R gene and protein expression levels were well maintained, while Bcl-2, Bax gene and protein expression levels progressively increased,and then decreased after 14 days （P〈O.05）. The condylar chondrocytes, incubated with 100 ng/mL rrIGF-1, showed high speed proliferation and increasing cell numbers. The result of Real-time PCR showed that IGF-IR, GSK-3, Bax, integrin gene expressions were down-regulated, while other factors, IGF-2R, Rafl,IGFBP3, NF-KB, Bcl-2 and TGF-13 gene expression levels tended to up-reguhed（P〈0.05）. Conclusions： IGF-1 signal pathway,which may interact with other proteins, such as integrin,TGF-13, regulates proliferation and differentiation of rat condylar chondrocytes and cartilage forming.

关 键 词：胰岛素样生长因子-1 髁突 软骨细胞 信号通路 鼠 

分 类 号：R329.28[医药卫生—人体解剖和组织胚胎学]