BACKGROUND Expression of hepcidin, a hormone produced by hepatocytes which negatively regulates the circulating iron levels, is known to be positively regulated by BMP6, a member of transforming growth factor (TGF)-β family. Previous studies have shown that iron status is sensed by sinusoidal endothelial cells of hepatic lamina, leading to the modulation of BMP6 expression. METHODS ISOS-1, HUVEC, F-2, and SK-HEP1 endothelial cells were treated with either iron or 2,2'-dipyridyl (2DP), a cell-permeable iron-chelator, and expression level of Bmp6 was examined. To identify factors affecting Bmp6 transcription, stimulus screening for regulator of transcription (SSRT) was developed. RESULTS Treatment with iron slightly increased the expression levels of Bmp6, while 2DP unexpectedly increased Bmp6 expression in a dose-dependent manner. 2DP-induced Bmp6 expression was resistant to co-treatment with iron. 2DP-induced Bmp6 expression was also detected in HUVEC, F-2 cells, and SK-HEP1 cells. Luciferase-based reporter assays indicated that forced expression of JunB increased the transcription of Bmp6. 2DP induced phosphorylation of JunB; co-treatment with SP600125 blocked the 2DP-induced Bmp6 expression partially. JunB-induced Bmp6 transcription was not affected by mutations of putative JunB-responsive elements. Some endoplasmic reticulum stress inducers increased the expression of Bmp6. SSRT revealed pathways regulating Bmp6 transcription positively and negatively. Hepa1-6 liver cells and C2C12 myogenic cells were prone to 2DP induced Bmp6 expression. CONCLUSIONS The present study reveals non‑iron-regulated Bmp6 expression in endothelial cells. GENERAL SIGNIFICANCE Regulatory expression of Bmp6 may be important as a key step for fine tuning of BMP activity.

中文翻译：

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2,2'-联吡啶对骨形态发生蛋白6的调控表达。

背景技术已知铁调素是由肝细胞产生的一种负调节循环铁水平的激素，其表达受转化生长因子（TGF）-β家族成员BMP6的正调节。先前的研究表明，肝薄层的正弦内皮细胞可检测到铁的状态，从而导致BMP6表达的调节。方法用铁或2,2'-联吡啶（2DP），可渗透细胞的铁螯合剂处理ISOS-1，HUVEC，F-2和SK-HEP1内皮细胞，并检测Bmp6的表达水平。为了确定影响Bmp6转录的因素，开发了针对转录调节子（SSRT）的刺激筛选。结果用铁处理稍微增加了Bmp6的表达水平，而2DP却以剂量依赖的方式意外地增加了Bmp6的表达。2DP诱导的Bmp6表达对与铁共处理具有抗性。在HUVEC，F-2细胞和SK-HEP1细胞中也检测到2DP诱导的Bmp6表达。基于萤光素酶的报告基因检测表明，JunB的强制表达增加了Bmp6的转录。2DP诱导JunB的磷酸化；与SP600125共同治疗可部分阻断2DP诱导的Bmp6表达。JunB诱导的Bmp6转录不受假定的JunB反应元件突变的影响。一些内质网应激诱导剂增加了Bmp6的表达。SSRT揭示了正向和负向调节Bmp6转录的途径。Hepa1-6肝细胞和C2C12肌原细胞易于2DP诱导的Bmp6表达。结论本研究揭示了内皮细胞中非铁调节的Bmp6表达。