TRIM35通过介导EMT促进骨肉瘤细胞侵袭和迁移的作用机制研究

The mechanism of TRIM35 promoting the invasion and migration of osteosarcoma cells by mediating EMT

目的研究含三元基序家族蛋白35(tripartite motif-containing protein 35,TRIM35)在骨肉瘤组织中的表达、对人骨肉瘤细胞系143B、Saos-2侵袭和迁移的作用及机制。方法收集30例正常人骨组织与64例骨肉瘤患者的肿瘤组织,应用免疫组化技术检测TRIM35的表达情况;通过转染TRIM35的siRNA与过表达质粒载体,结合qRT-PCR与Western blot技术检测转染效率;在划痕与Transwell实验中观察TRIM35对骨肉瘤细胞侵袭与迁移能力的影响;在骨肉瘤细胞中沉默或过表达TRIM35后,利用Western blot技术检测上皮-间质转化(Epithelial-mesenchymal transtion,EMT)进程标志蛋白。结果免疫组化实验中,TRIM35在骨肉瘤组织中高表达,正常组织低表达。划痕实验结果表明,沉默TRIM35能够抑制143B的迁移能力,过表达后可加快Saos-2细胞的迁移;Transwell实验显示过表达TRIM35能够促进Saos-2细胞的侵袭,沉默TRIM35能够抑制143B细胞的侵袭;Western blot结果显示沉默或过表达TRIM35能够引起EMT进程的标志性蛋白的变化。结论TRIM35在骨肉瘤组织中高表达,通过介导EMT促进骨肉瘤细胞的侵袭及迁移。

Objective To study the clinical significance of tripartite motif-containing protein 35(TRIM35)in osteosarcoma tissues and the mechanism of its effect on the invasion and metastasis of osteosarcoma cells 143B and Saos-2.Methods The expression of TRIM35 was detected by immunohistochemistry in 30 normal bone tissues and 64 osteosarcoma tissues.The transfection efficiency was detected by siRNA and over-expression plasmid vector transfected with TRIM35 combined with qRT-PCR and western blot.The effect of TRIM35 on the invasion and metastasis of osteosarcoma cells was observed by scratch and Transwell experiments.After silencing or overexpression of TRIM35 in osteosarcoma cells,the epithelial-mesenchymal transition(EMT)process marker protein was detected by western blot.Results In the immunohistochemical experiment,TRIM35 was highly expressed in osteosarcoma tissues and lowly expressed in normal tissues.The scratch test results showed that silencing TRIM35 could inhibit the migration ability of 143B and accelerate the migration of Saos-2 cells after overexpression.Transwell experiment showed that overexpression of TRIM35 could promote the invasion of Saos-2 cells,and silencing TRIM35 could inhibit the invasion of 143B cells.Western blot results showed that silencing or overexpression of TRIM35 could cause changes in the marker proteins of EMT process.Conclusion TRIM35 is highly expressed in osteosarcoma tissues and promotes the invasion and migration of osteosarcoma cells by mediating EMT.