Gata4 and LMO3 balance angiocrine signaling and autocrine inflammatory activation by BMP2 in liver sinusoidal endothelial cells [HUVEC]

Liver sinusoidal endothelial cells (LSEC) represent a unique, organ-specific type of discontinuous endothelial cells. LSEC instruct the hepatic vascular niche by paracrine-acting angiocrine factors. Recently, we have shown that LSEC-specific transcriptional regulator GATA4 induces expression of BMP2 in cultured endothelial cells (EC) in vitro. Furthermore, angiocrine Bmp2 signaling in the liver in vivo was demonstrated to control iron homeostasis. Here, we investigated GATA4-dependent autocrine BMP2 signaling in endothelial cells by gene expression profiling. GATA4 induced a large cluster of inflammatory endothelial response genes in cultured EC, which is similar to previously identified virus-induced and interferon-associated responses. Treating the cells with the BMP2 inhibitor Noggin counter-regulated the GATA4-dependent inflammatory phenotype of EC, indicating that BMP2 is indeed the major driver. In contrast to continuous EC, LSEC were less prone to activation by BMP2. Notably, GATA4-dependent induction of the inflammatory EC response gene cluster was attenuated by over-expression of the LSEC-specific transcriptional modifier LMO3 while hepatocyte activation was fully preserved, indicating conserved BMP2 synthesis. In summary, our data suggest that transcriptional counter-regulation by GATA4 and LMO3 in LSEC prevents autocrine induction of an inflammatory phenotype, while maintaining angiocrine BMP2-mediated cell communication in the liver vascular niche. HUVEC were transduced with human GATA4 or empty vector and stimulated with or without Noggin or BMP2 for RNA extraction and hybridization on Affymetrix microarrays.

肝窦内皮细胞（Liver sinusoidal endothelial cells, LSEC）是一类独特的器官特异性不连续性内皮细胞。它们通过发挥旁分泌作用的血管分泌因子调控肝脏血管微环境。此前本课题组已证实，LSEC特异性转录调控因子GATA4可在体外培养的内皮细胞（endothelial cells, EC）中诱导骨形态发生蛋白2（BMP2）的表达。进一步研究表明，体内肝脏中的血管分泌Bmp2信号通路可调控铁稳态。
本研究通过基因表达谱分析，探究了内皮细胞中GATA4依赖的自分泌BMP2信号通路。GATA4可在培养的内皮细胞中诱导大量炎症性内皮应答基因簇的表达，这与此前已报道的病毒诱导及干扰素相关应答特征相似。使用BMP2抑制剂Noggin处理细胞后，可抵消GATA4介导的内皮细胞炎症表型，证实BMP2确实是该过程的核心驱动因子。
相较于连续性内皮细胞，LSEC对BMP2介导的激活敏感性更低。值得注意的是，过表达LSEC特异性转录修饰因子LMO3可削弱GATA4依赖的炎症性内皮应答基因簇诱导效应，而肝细胞激活过程则完全不受影响，这提示BMP2的合成过程具有保守性。
综上，本研究数据表明，LSEC中GATA4与LMO3介导的转录反向调控，可阻止自分泌途径诱导的炎症表型产生，同时维持肝脏血管微环境中血管分泌BMP2介导的细胞通讯。本实验将人GATA4或空载体转导至人脐静脉内皮细胞（Human Umbilical Vein Endothelial Cells, HUVEC），并分别施以或不施以Noggin或BMP2刺激，随后提取RNA并在Affymetrix基因芯片上进行杂交检测。