Chen Chiachen, Breslin Mary B, Lan Michael S
Research Institute for Children, Children's Hospital, 200 Henry Clay Avenue, Research and Education Building, Room. 2211, New Orleans, LA, 70118, USA.
Departments of Pediatrics, Louisiana State University Health Sciences Center, New Orleans, LA, 70112, USA.
BMC Pulm Med. 2016 Apr 12;16:49. doi: 10.1186/s12890-016-0215-3.
Insulinoma associated-1 (INSM1) gene is expressed exclusively in early embryonic neuroendocrine tissues, but has been found highly re-activated in most of the neuroendocrine tumors including small cell lung carcinoma.
In order to elucidate the functional effects of INSM1 in normal lung development, we used a conditional lung-specific INSM1 transgenic mouse model. Transgenic (Tet-on system) CMV-INSM1 responder mice were bred with the lung-specific, club cell secretory protein (CCSP) promoter-rtTA activator mice to produce bi-transgenic progeny carrying both alleles, CCSP-rtTA and Tet-on-INSM1. Mice were fed with doxycycline containing food at the initial mating day to the postnatal day 21. Lung samples were collected at embryonic day 17.5, newborn, and postnatal day 21 for analyses.
Northern blot, RT-PCR, and immunohistochemical analyses revealed that doxycycline induced respiratory epithelium-specific INSM1 expression in bi-transgenic mice. Samples from postnatal day 21 mice revealed a larger lung size in the bi-transgenic mouse as compared to the single-transgenic or wild-type littermates. The histopathology results showed that the alveolar space in the bi-transgenic mice were 4 times larger than those in the single transgenic or wild-type littermates. In contrast, the size was not significantly different in the lungs collected at E17.5 or newborn among the bi-transgenic, single transgenic, or wild type mice. The respiratory epithelium with INSM1 ectopic expression suppressed cyclin D1 signal. Further in vitro studies revealed that the ectopic expression of INSM1 suppresses cyclin D1 expression and delays cell cycle progression.
The current study suggests that CCSP promoter-driven INSM1 ectopic expression impairs normal lung development especially in postnatal alveologenesis.
胰岛素瘤相关-1（INSM1）基因仅在胚胎早期神经内分泌组织中表达，但已发现在包括小细胞肺癌在内的大多数神经内分泌肿瘤中高度重新激活。
为了阐明INSM1在正常肺发育中的功能作用，我们使用了条件性肺特异性INSM1转基因小鼠模型。将转基因（Tet-on系统）CMV-INSM1反应小鼠与肺特异性的俱乐部细胞分泌蛋白（CCSP）启动子-rtTA激活小鼠杂交，以产生携带CCSP-rtTA和Tet-on-INSM1两个等位基因的双转基因后代。在初始交配日至出生后第21天，给小鼠喂食含强力霉素的食物。在胚胎第17.5天、新生期和出生后第21天收集肺样本进行分析。
Northern印迹、RT-PCR和免疫组织化学分析显示，强力霉素诱导双转基因小鼠中呼吸道上皮特异性INSM1表达。出生后第21天小鼠的样本显示，与单转基因或野生型同窝小鼠相比，双转基因小鼠的肺更大。组织病理学结果表明，双转基因小鼠的肺泡腔比单转基因或野生型同窝小鼠大4倍。相比之下，在胚胎第17.5天或新生期收集的双转基因、单转基因或野生型小鼠的肺大小没有显著差异。异位表达INSM1的呼吸道上皮抑制细胞周期蛋白D1信号。进一步的体外研究表明，INSM1的异位表达抑制细胞周期蛋白D1表达并延迟细胞周期进程。
当前研究表明，CCSP启动子驱动的INSM1异位表达损害正常肺发育，尤其是在出生后肺泡形成过程中。
