Activation of the non-canonical NF-κB/p52 pathway in vascular endothelial cells by RANKL elicits pro-calcific signalling in co-cultured smooth muscle cells.
Activation of the non-canonical NF-κB/p52 pathway in vascular endothelial cells by RANKL elicits pro-calcific signalling in co-cultured smooth muscle cells.
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DOI：
10.1016/j.cellsig.2018.04.004
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发表时间：
2018-07
影响因子：
4.8
通讯作者：
Cummins PM
中科院分区：
文献类型：
--
作者：
Harper E;Rochfort KD;Forde H;Davenport C;Smith D;Cummins PM
已知内膜内皮调节血管壁的底层内侧平滑肌细胞 (SMC) 层，并且分别对核因子 kB 配体受体激活剂 (RANKL) 和肿瘤坏死因子相关凋亡诱导配体 (TRAIL)、促钙化剂和抗钙化剂高度敏感。在本文中，我们测试了以下假设：RANKL 诱导的内皮 NF-κB 信号传导激活对于底层 SMC 的促钙化激活至关重要。在这些研究中，人主动脉内皮细胞和平滑肌细胞单一培养物（HAEC、HASMC）用 RANKL（0–25 ng/ml ± 5 ng/ml TRAIL）处理 72 小时。还采用非接触式 Transwell HAEC:HASMC 共培养，其中用 RANKL (± 5 ng/ml TRAIL) 处理管腔 HAEC，然后分析底层管腔下 HASMC 中的促钙化标记。用 RANKL 处理 HAEC 或 HASMC 都会激活两种细胞类型中的非经典 NF-κB/p52 和经典 NF-κB/p65 通路。在 RANKL ± TRAIL 处理的 HAEC 中，我们的团队先前证明重组 TRAIL 可以强烈减弱 RANKL 的促钙化信号传导作用，但它可以特异性阻断 RANKL 介导的非经典 NF-κB/p52 的激活，清楚地表明这一特定途径与内皮细胞内 RANKL 功能的机制相关性。在最后一系列 HAEC:HASMC Transwell 共培养实验中，相对于乱序 siRNA 对照，对 NF-κB2 基因（NF-κB/p52 生成的分子先驱）进行基因沉默（通过 siRNA）的 HAEC 进行 RANKL 处理，表现出强烈减弱的潜在 HASMC 的促钙化激活。这些体外观察结果为 RANKL 如何通过激活替代 NF-κB 通路改变内皮旁分泌信号并引发底层血管平滑肌细胞内的促钙化反应而潜在地作用于内皮细胞提供了有价值的机制见解。
The intimal endothelium is known to condition the underlying medial smooth muscle cell (SMC) layer of the vessel wall, and is highly responsive to receptor-activator of nuclear factor-kB ligand (RANKL) and tumour necrosis factor-related apoptosis-inducing ligand (TRAIL), pro-calcific and anti-calcific agents, respectively. In this paper, we tested the hypothesis that RANKL-induced activation of endothelial NF-κB signalling is essential for pro-calcific activation of the underlying SMCs. For these studies, human aortic endothelial and smooth muscle cell mono-cultures (HAECs, HASMCs) were treated with RANKL (0–25 ng/ml ± 5 ng/ml TRAIL) for 72h. Non-contact transwell HAEC:HASMC cocultures were also employed in which the luminal HAECs were treated with RANKL (± 5 ng/ml TRAIL), followed by analysis of pro-calcific markers in the underlying subluminal HASMCs. Treatment of either HAECs or HASMCs with RANKL activated the non-canonical NF-κB/p52 and canonical NF-κB/p65 pathways in both cell types. In RANKL ± TRAIL-treated HAECs, recombinant TRAIL, previously demonstrated by our group to strongly attenuate the pro-calcific signalling effects of RANKL, was shown to specifically block the RANKL-mediated activation of non-canonical NF-κB/p52, clearly pointing to the mechanistic relevance of this specific pathway to RANKL function within endothelial cells. In a final series of HAEC:HASMC transwell co-culture experiments, RANKL treatment of HAECs that had been genetically silenced (via siRNA) for the NF-κB2 gene (the molecular forerunner to NF-κB/p52 generation) exhibited strongly attenuated pro-calcific activation of underlying HASMCs relative to scrambled siRNA controls. These in vitro observations provide valuable mechanistic insights into how RANKL may potentially act upon endothelial cells through activation of the alternative NF-κB pathway to alter endothelial paracrine signalling and elicit pro-calcific responses within underlying vascular smooth muscle cells.
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影响因子：
82.9
作者：
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通讯作者：
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影响因子：
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作者：
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影响因子：
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作者：
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影响因子：
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作者：
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