NBD多肽促进成骨细胞分化的实验研究

NBD peptide promotes osteoblast differentiation

[目的]探讨核因子κB必需分子(NF-κB essential modulator,NEMO)结合的小分子多肽(NEMO bind-ing domain,NBD)通过阻断肿瘤坏死因子-α信号通路影响成骨细胞分化的作用及其分子机制。[方法]应用BMP-2体外诱导鼠肌源细胞C2C12向成骨细胞分化模型,外源添加TNF-α和/或BMP-2细胞因子培养,通过碱性磷酸酶(ALP)活性检测,瞬时转染和基因测定,研究NBD多肽对抗NF-κB活性和改善TNF-α抑制成骨细胞分化的过程。[结果]ALP染色显示NBD多肽能明显阻断TNF-α对C2C12向成骨细胞分化的抑制而促进其分化,荧光素酶活性测定显示TNF-α降低BMP-2活性从7.12倍到1.31倍,而NBD多肽使其恢复到6.7倍和mNBD肽恢复到1.4倍。[结论]TNF-α抑制成骨细胞分化的分子生物机制是通过激活NF-kB阻碍成骨细胞的分化。NBD多肽具有对抗NF-κB活性和改善TNF-α抑制成骨细胞分化过程的作用。

[ Objective ] To investigate the effect and molecular mechanism of NBD peptide on osteoblast differentiation. [ Method] The C2C12 ceils stimulated with BMP -2 were used as an in vitro model for studying osteoblast differentiation. Transi- ent transfection, lnciferase reporter activity assay and real-time RT-PCR were performed to study the effect of NBD peptide on os- teoblast differentiation. [ Result] TNF-α significantly reduced the ALP activities in a dose dependent manner in BMP2-induced C2C12 cells. TNF-α decreased the BMP-2 activity from 7.12 to 1.31 fold,and NBD peptide restored it back to 6.7 fold,while mNBD peptide left it at I. 4 fold. [ Conclusion] TNF-a inhibits osteoblast differentiation through the activation of NF-κB. Appli- cation of NBD peptide ameliorates this inhibitory effect. Key words：osteoblast differentiation, tumor necrosis factor-alpha, nuclear factor kappa B, NEMO Binding domain