敲降胰高血糖素基因对胰岛素瘤形成的影响  

The effect of knockdown of glucagon gene on insulinoma formation

作　　者：朱国玲[1,2] 成兰云 张恒 田浩 门秀丽 ZHU Guo-ling;CHENG Lan-yun;ZHANG Heng;TIAN Hao;MEN Xiu-li(Department of Gastroenterology, Kailuan General Hospital, Tangshan 063000, China;Tangshan Key Laboratory forPreclinical and Basic Research on Chronic Diseases, School of Basic Medical Sciences,North China University of Science and Technology)

机构地区：[1]开滦总医院消化内科,063000 [2]华北理工大学基础医学院,唐山市慢性病临床基础研究重点实验室

出　　处：《天津医药》2019年第10期1035-1039,I0001,共6页Tianjin Medical Journal

基　　金：河北省卫计委专项科研基金(20171437);国家自然科学基金资助项目(81370918)

摘　　要：目的通过体外培养的胰岛素瘤细胞和胰岛素瘤动物模型,探讨敲降胰高血糖素基因对胰岛素瘤形成的影响。方法通过免疫荧光染色,激光共聚焦显微镜观察INS-1细胞胰岛素和胰高血糖素蛋白的表达,应用慢病毒pLVTHM-glucagon敲降INS-1细胞的胰高血糖素基因,建立胰高血糖素低表达的INS-1细胞系,将细胞分成空白对照组(不进行感染)、单纯转染慢病毒组(感染pLVTHM)和敲降胰高血糖素组(感染pLVTHM-shglucagon)。 MTT法比较敲降胰高血糖素后INS-1细胞增殖能力、ELISA法和放射免疫分析法观察胰岛素和胰高血糖素的释放能力。将3组细胞分别移植到胰岛素瘤模型裸鼠左肾包膜下,观察细胞成瘤能力,并通过免疫组织化学染色法观察肿瘤组织中胰岛素和胰高血糖素的表达。结果与空白对照组和单纯转染慢病毒组相比,敲降胰高血糖素组的INS-1细胞增殖能力减弱,葡萄糖刺激的胰岛素分泌功能无明显变化,但胰高血糖素分泌明显减少;与单纯转染慢病毒组相比,敲降胰高血糖素组细胞移植到裸鼠肾包膜下时,细胞成瘤速度明显降低;免疫组织化学染色结果显示此胰岛素瘤组织胰高血糖素表达明显下调。结论敲降胰高血糖素基因可抑制胰岛素瘤细胞的增殖和胰高血糖素的释放,从而抑制胰岛素瘤的形成,提示胰高血糖素基因表达可能是胰岛素瘤发生的一个重要机制。Objective To discuss the effect of knockdown of glucagon gene on insulinoma formation by using cells in vitro and animal model of insulinoma. Methods The expressions of insulin and glucagon proteins in INS-1 cells were observed by immunofluorescence staining and laser scanning confocal microscopy. Then INS-1 cells were infected with lentivirus pLVTHM-glucagon to knock down the glucagon gene. Therefore, an INS-1 cell line with low glucagon expression was established. Cells were divided into blank control group, lentivirus transfection group and glucagon knockdown group. The cell proliferation was detected by MTT assay. The ability to release insulin and glucagon was detected using ELISA and radioimmunity. And in vivo tumorigenicity of INS-1 cells was evaluated after glucagon gene knockdown. Expression levels of insulin and glucagon of tumor tissue were further detected by immunohistochemistry staining. Results Knockdown of glucagon gene impaired proliferation of INS-1 cells. The release of glucagon, but not insulin was decreased after glucagon knockdown. Subrenal capsul assay showed significantly reduced speed of neoplasia of INS-1 cells, with markedly decreased glucagon gene expression in this insulinoma tissue, which was displayed by immunohistochemical staining. Conclusion The knockdown of glucagon gene can inhibit proliferation of insulinoma cells, and release of glucagon, further restrain insulinoma formation. The result indicates that glucagon gene expression may be an important factor of the insulinoma formation.

关 键 词：胰岛素瘤 胰岛素 葡萄糖 胰高血糖素 INS-1 

分 类 号：R735.9[医药卫生—肿瘤]