目的:骨肉瘤转移的分子机制研究仍是骨肉瘤(OS)防治的研究热点。本研究探讨了含valosin蛋白(VCP)通过自噬和上皮间质转化(EMT)促进OS转移的机制。
方法:本研究采用不同的骨肉瘤细胞系(143B和MG63)。骨肉瘤细胞VCP表达水平改变,观察自噬水平和上皮间质转化(EMT)的进展。然后人工改变OS细胞的自噬和EMT,并观察到增殖和迁移能力。
结果:LC3II/I的表达减少,但VCP抑制组和自噬抑制剂治疗组P62蛋白表达增加。同时,在VCP抑制组中,E-cadherin蛋白表达增加,而N-cadherin蛋白表达减少,但在TGF-β1治疗组中增加。此外,抑制VCP可以导致转化生长因子β1(TGF-β1)的减少,smad2,smad3,磷酸化smad2(p-smad2),和磷酸化的smad3(p-smad3)。自噬抑制剂和激动剂对OS细胞的迁移和侵袭没有明显影响,但可以显着影响细胞抵抗失巢凋亡的能力。EMT抑制剂和激动剂对OS细胞的迁移和侵袭具有成比例的作用。
结论:VCP可能通过诱导EMT促进OS细胞的迁移和侵袭,可能通过TGF-β1/smad2/3信号通路。在这个过程中,VCP介导的自噬可能间接促进肿瘤细胞的远处转移。
OBJECTIVE: Study of the molecular mechanisms of metastasis is still the research focus for osteosarcoma (OS) prevention. This study investigates the mechanism of valosin-containing protein (VCP) promoting OS metastasis in vitro through autophagy and epithelial-mesenchymal transition (EMT).
METHODS: Different cell lines of osteosarcoma (143B and MG63) were adopted in this study. The level of VCP expression in osteosarcoma cells was changed, and the level of autophagy and the progression of the epithelial-mesenchymal transition (EMT) were observed. Then autophagy and EMT in OS cells were changed artificially, and proliferation and migration ability were observed.
RESULTS: The expression of LC3II/I was decreased, but the insolubilized P62 protein expression was increased in the VCP inhibiting group and the autophagy inhibitor treatment group. Simultaneously, E-cadherin protein expression increased while N-cadherin protein expression decreased in the VCP inhibiting group but increased in the TGF-β1 treatment group. In addition, suppressing VCP can cause a decrease in Transforming Growth Factor β1 (TGF-β1), smad2, smad3, phosphorylated smad2 (p-smad2), and phosphorylated smad3 (p-smad3). Autophagy inhibitors and agonists have no significant effect on the migration and invasion of OS cells but can significantly affect the ability of cells to resist anoikis. EMT inhibitors and agonists have a proportional effect on the migration and invasion of OS cells.
CONCLUSIONS: VCP is likely to promote the migration and invasion of OS cells by inducing EMT, possibly via TGF-β1/smad2/3 signaling pathway. In this process, VCP-mediated autophagy may contribute to successful distant metastasis of tumor cells indirectly.