Interleukin-1β up-regulation in human of Smad7 via NF-κB activation chondrocytes
Interleukin-1β up-regulation in human of Smad7 via NF-κB activation chondrocytes
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DOI：
10.1002/art.23154
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发表时间：
2008-01-01
影响因子：
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通讯作者：
Boumediene, K.
中科院分区：
文献类型：
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作者：
Bauge, C.;Attia, J.;Boumediene, K.
目标。我们先前已经证明，白介素1β(IL-1β)通过下调转化生长因子β受体II(TGFβRII)和上调Smad7而损害转化生长因子β(TGFβ)信号转导。这一机制可以解释骨关节炎软骨细胞对转化生长因子β的反应性降低，以及与这种疾病有关的软骨破坏。本研究旨在探讨IL-1β刺激人关节软骨细胞Smad7的分子机制。人关节软骨细胞在含有转化生长因子β1、吡咯烷二硫代氨基甲酸酯(一种核因子-kappa B途径的阻滞剂)或放线菌亚胺的情况下，用IL-1β处理。用实时逆转录聚合酶链式反应和免疫细胞学方法检测细胞内的稳态信使RNA和蛋白水平。此外，瞬时转染p65表达载体或p65靶向的短发夹状RNA以确定核因子-kappa B对Smad7表达的影响。人关节软骨细胞过度表达转化生长因子βRII可恢复转化生长因子β反应。然而，这种影响是短暂的，这意味着长期暴露于IL-1β的转化生长因子β反应的改变是由次级机制引起的。此外，IL-1β导致抑制Smad7的迟发诱导。这种效果是直接的，因为它不需要从头合成。此外，通过功能得失的实验，我们证实了IL-1β上调Smad7是通过核因子-kappaB途径，尤其是p65亚单位介导的。这些发现阐明了IL-1β对Smad7表达的调节过程。了解IL-1β诱导Smad7和降低软骨细胞对转化生长因子β的反应性的分子基础有助于深入了解骨关节炎的分子机制，并可能有助于确定其治疗的新方法。
Objective. We have previously shown that interleukin-1 beta (IL-1 beta) impairs transforming growth factor beta (TGF beta) signaling through TGF beta receptor type II (TGF beta RII) down-regulation and Smad7 up-regulation. This mechanism could account for the reduced responsiveness of osteoarthritic chondrocytes to TGF beta and the cartilage breakdown linked to this disease. The aim of this study was to investigate the molecular mechanism underlying the IL-1 beta-induced stimulation of Smad7 in human articular chondrocytes.Methods. Human articular chondrocytes were treated with IL-1 beta in the presence of TGF beta 1, pyrrolidine dithiocarbamate (a repressor of the NF-kappa B pathway), or cycloheximide. Then, steady-state messenger RNA and protein levels were estimated by real-time reverse transcription-polymerase chain reaction and immunocytology. In addition, transient transfections of p65 expression vector or p65-targeted short hairpin RNA were performed to define the effect of NF-kappa B on Smad7 expression.Results. TGF beta RII overexpression restored the TGF beta response of human articular chondrocytes. However, this effect was transient, implying that a secondary mechanism was responsible for the alteration of the TGF beta response with long-term exposure to IL-1 beta. Moreover, IL-1 beta caused a late induction of the inhibitory Smad7. This effect was direct, since it did not require de novo synthesis. In addition, we established, by experiments with gain/loss of function, that the up-regulation of Smad7 by IL-1 beta is mediated through the NF-kappa B pathway, especially the p65 subunit.Conclusion. These findings clarify the regulatory process of IL-1 beta on Smad7 expression. Understanding the molecular basis of IL-1 beta induction of Smad7 and the reduction of chondrocyte responsiveness to TGF beta provides new insights into the molecular mechanisms of osteoarthritis and may facilitate the identification of novel approaches for its treatment.