Cardiovascular Division, King's College London BHF Centre of Research Excellence, London, United Kingdom.
PLoS One. 2011;6(8):e22457. doi: 10.1371/journal.pone.0022457. Epub 2011 Aug 11.
Small intestinal (SI) neuroendocrine tumors (NET) are increasing in incidence, however little is known about their biology. High throughput techniques such as inference of gene regulatory networks from microarray experiments can objectively define signaling machinery in this disease. Genome-wide co-expression analysis was used to infer gene relevance network in SI-NETs. The network was confirmed to be non-random, scale-free, and highly modular. Functional analysis of gene co-expression modules revealed processes including 'Nervous system development', 'Immune response', and 'Cell-cycle'. Importantly, gene network topology and differential expression analysis identified over-expression of the GPCR signaling regulators, the cAMP synthetase, ADCY2, and the protein kinase A, PRKAR1A. Seven CREB response element (CRE) transcripts associated with proliferation and secretion: BEX1, BICD1, CHGB, CPE, GABRB3, SCG2 and SCG3 as well as ADCY2 and PRKAR1A were measured in an independent SI dataset (n = 10 NETs; n = 8 normal preparations). All were up-regulated (p<0.035) with the exception of SCG3 which was not differently expressed. Forskolin (a direct cAMP activator, 10(-5) M) significantly stimulated transcription of pCREB and 3/7 CREB targets, isoproterenol (a selective ß-adrenergic receptor agonist and cAMP activator, 10(-5) M) stimulated pCREB and 4/7 targets while BIM-53061 (a dopamine D(2) and Serotonin [5-HT(2)] receptor agonist, 10(-6) M) stimulated 100% of targets as well as pCREB; CRE transcription correlated with the levels of cAMP accumulation and PKA activity; BIM-53061 stimulated the highest levels of cAMP and PKA (2.8-fold and 2.5-fold vs. 1.8-2-fold for isoproterenol and forskolin). Gene network inference and graph topology analysis in SI NETs suggests that SI NETs express neural GPCRs that activate different CRE targets associated with proliferation and secretion. In vitro studies, in a model NET cell system, confirmed that transcriptional effects are signaled through the cAMP/PKA/pCREB signaling pathway and that a SI NET cell line was most sensitive to a D(2) and 5-HT(2) receptor agonist BIM-53061.
小肠（SI）神经内分泌肿瘤（NET）的发病率正在增加，但对其生物学特性知之甚少。高通量技术，如从微阵列实验推断基因调控网络，可以客观地定义这种疾病中的信号机制。全基因组共表达分析用于推断 SI-NETs 的基因相关性网络。该网络被证实是非随机的、无标度的和高度模块化的。基因共表达模块的功能分析揭示了包括“神经系统发育”、“免疫反应”和“细胞周期”在内的过程。重要的是，基因网络拓扑结构和差异表达分析确定了 GPCR 信号调节剂、cAMP 合成酶 ADCY2 和蛋白激酶 A PRKAR1A 的过度表达。七种与增殖和分泌相关的 CREB 反应元件（CRE）转录物：BEX1、BICD1、CHGB、CPE、GABRB3、SCG2 和 SCG3 以及 ADCY2 和 PRKAR1A，在一个独立的 SI 数据集（n=10 NET；n=8 个正常制剂）中进行了测量。所有转录物均上调（p<0.035），除了 SCG3 没有差异表达。forskolin（一种直接的 cAMP 激活剂，10（-5）M）显著刺激 pCREB 和 3/7 CREB 靶基因的转录，异丙肾上腺素（一种选择性β-肾上腺素能受体激动剂和 cAMP 激活剂，10（-5）M）刺激 pCREB 和 4/7 靶基因，而 BIM-53061（一种多巴胺 D2 和血清素[5-HT2]受体激动剂，10（-6）M）刺激 100%的靶基因以及 pCREB；CRE 转录与 cAMP 积累和 PKA 活性水平相关；BIM-53061 刺激产生最高水平的 cAMP 和 PKA（2.8 倍和 2.5 倍，分别为异丙肾上腺素和 forskolin的 1.8-2 倍）。SI NETs 中的基因网络推断和图形拓扑分析表明，SI NETs 表达神经 GPCR，其激活与增殖和分泌相关的不同 CRE 靶基因。在体外研究中，在一个模型 NET 细胞系统中，证实转录效应是通过 cAMP/PKA/pCREB 信号通路发出的，并且 SI NET 细胞系对 D2 和 5-HT2 受体激动剂 BIM-53061 最敏感。
