Regulation of pituitary gene expression by adrenalectomy

Overproduction of adrenal hormones causes obesity and hypertension, whereas adrenalectomy (ADX) leads to weight loss and hypotension. The pituitary gland is a major target organ for the adrenal hormones. To identify potential mediators for the effect of ADX on pituitary system regulating weight loss and hypotension, we investigated the effect of ADX on pituitary gene expression using serial analysis of gene expression (SAGE). The numbers of analyzed SAGE tag were 126688 tags for intact mice (n = 51) and 59482 tags for ADX mice (n = 12), which corresponded to 45151 and 21790 distinct tag species, respectively. Thirty-three pituitary genes were differentially expressed between intact and ADX mice. Three genes encoding for pro-opiomelanocortin have been upregulated by ADX. The ADX has induced the gene expression of growth hormone while downregulating an expressed sequence tag similar to (EST) growth hormone. The ADX reduced the expression of genes involved in mitochondrial oxidative phosphorylation, such as NADH dehydrogenase 3 and cytochrome c oxidase 3, whereas neuromedin B gene were dramatically induced. In addition, the ADX increased the expression levels of other three genes involved in hormone release (response to metastatic cancers 1), extracellular matrix (matrix gamma-carboxyglutamate gla protein), and cation transport (solute carrier family member 17). Moreover, one functionally-uncharacterized and 20 novel transcripts were significantly modulated by ADX. Thus the current study has revealed the alterations in the pituitary gene expression that may play key roles in the mechanisms of ADX-induced weight loss and hypotension. Keywords: adrenalectomy, serial analysis of gene expression Rodents and pituitary gland sampling. Male C57BL6 mice were obtained from Charles River Laboratories (St. Constant, QC, Canada), at 12-14 weeks of age. Mice were housed in an air-conditioned room (19-25℃) with controlled lighting from 07:15 to 19:15 h and were given free access to food (Lab Rodent Diet No. 5002) and water. One week prior to sampling of pituitary gland, adrenalectomy was performed in mice of ADX groups (n = 12). The ADX mice received sodium chloride (0.9g/dl) in their drinking water after the surgery. All mice were killed between 08:30 and 12:30 by decapitation under isoflurane anesthesia. Brain was removed from the skull and the pituitary gland was immediately dissected. The pituitary gland was immediately frozen in liquid nitrogen, pooled together for each group, and stored at -80℃ until RNA extraction.肾上腺激素过量合成可诱发肥胖与高血压，而肾上腺切除术（adrenalectomy, ADX）则会导致体重减轻与低血压。垂体是肾上腺激素的主要靶器官。为明确ADX通过调节垂体系统介导体重减轻和低血压效应的潜在介质，我们采用基因表达系列分析（serial analysis of gene expression, SAGE）技术，探究了ADX对小鼠垂体基因表达的影响。本研究分析的SAGE标签总数分别为：完整对照组小鼠（n=51）126688个标签，ADX组小鼠（n=12）59482个标签，对应分别得到45151和21790种独特标签序列。两组小鼠垂体组织中共鉴定出33个差异表达基因。其中，3个编码阿片黑素皮质素原（pro-opiomelanocortin）的基因在ADX后表达上调；生长激素基因的表达被ADX诱导增强，而一条与生长激素相似的表达序列标签（expressed sequence tag, EST）的表达则被下调。ADX可降低参与线粒体氧化磷酸化的基因表达，例如NADH脱氢酶3（NADH dehydrogenase 3）和细胞色素c氧化酶3（cytochrome c oxidase 3），但神经介素B（neuromedin B）基因的表达则被显著诱导升高。此外，ADX还上调了另外三类基因的表达：分别参与激素释放过程的转移癌应答基因1（response to metastatic cancers 1）、细胞外基质相关的γ-羧基谷氨酸Gla蛋白基因（matrix gamma-carboxyglutamate gla protein），以及阳离子转运相关的溶质载体家族17成员（solute carrier family member 17）。另有1个功能未注释的转录本以及20个全新转录本的表达水平被ADX显著调控。综上，本研究揭示了垂体基因表达的改变，这些改变可能在ADX诱导的体重减轻和低血压发生机制中发挥关键作用。 关键词：肾上腺切除术、基因表达系列分析 实验动物与垂体取样。雄性C57BL/6小鼠购自加拿大Charles River Laboratories公司（魁北克省圣康斯坦特），周龄为12~14周。小鼠饲养于控温（19~25℃）空调房间，光照周期设置为07:15至19:15，自由摄食"Lab Rodent Diet No.5002"啮齿类专用饲料并自由饮水。在垂体取样前1周，对ADX组小鼠（n=12）实施肾上腺切除术，术后小鼠的饮用水中添加0.9g/dl氯化钠。所有小鼠均在08:30至12:30期间，经异氟烷麻醉后脱颈椎处死。剥离颅骨取出脑组织后，迅速分离垂体组织，立即置于液氮中速冻，按组混合后保存于-80℃冰箱，直至RNA提取。