利用GT1-7细胞系研究ghrelin对下丘脑刺鼠相关蛋白的表达效果  

A study of the effect of ghrelin on hypothalamic agouti-related protein expressing by using GT1-7 cell lines

作　　者：李培鑫[1] 饶志坚 黄虎 Li Peixin;Rao Zhijian;Huang Hu(Department of Comprehensive Surgery,Beijing Friendship Hospital,Capital Medical University,Beijing 100050,China;Department of Kinesiology,Physical Education College,Shanghai Normal University,Shanghai 200234,China;Human Performance Lab,Department of Kinesiology&Physiology,East Carolina University,East Carolina Diabetes and Obesity Institute,North Carolina 27834,USA)

机构地区：[1]首都医科大学附属北京友谊医院医疗保健中心综合外科,北京100050 [2]上海师范大学体育学院人体科学教研室,上海200234 [3]东卡罗来纳大学运动人体科学系、生理系、人体机能研究所东卡罗来纳糖尿病与肥胖研究所,美国北卡来纳州27834

出　　处：《首都医科大学学报》2022年第6期924-930,共7页Journal of Capital Medical University

基　　金：北京市保健课题(京19-10号)。

摘　　要：目的介绍一种可对ghrelin(胃促生长素,俗称饥饿素)刺激产生反应的细胞系,并观察ghrelin对刺鼠相关蛋白(agouti-related protein,AgRP)基因表达的影响。方法通过蛋白印迹(Western blotting)法检测证实GT1-7细胞是否具有ghrelin可以结合的生长激素促分泌素受体(growth hormone secretagogue receptor,GHSR);通过给予GT1-7细胞血清饥饿,以及在含血清状态下给予不同时间ghrelin刺激或不同浓度的ghrelin刺激,经实时定量聚合酶链式反应(real-time quantitative polymerase chain reaction,RT-qPCR)检验AgRP mRNA表达量差异。结果GT1-7细胞具有GHSR;血清饥饿显著增加了GT1-7细胞的AgRP mRNA表达(P<0.001);在含血清状态下,与空白对照组及ghrelin刺激1 h组比较,ghrelin刺激3 h组可使GT1-7细胞AgRP mRNA的表达显著增加(P=0.002,P=0.008);在含血清状态下,低浓度(10、50 nmol/L)ghrelin显著低于高浓度(100 nmol/L)ghrelin刺激的AgRP mRNA表达(P=0.021,P=0.016)。结论血清饥饿可以使GT1-7细胞增加AgRP mRNA的表达。Ghrelin即使在含血清状态下也可以刺激GT1-7细胞增加AgRP mRNA表达,并且为时间及浓度依赖。因此,GT1-7细胞可作为研究下丘脑神经元对AgRP调控,以及下丘脑神经元对其他代谢功能的调节,尤其是其背后分子机制研究的体外模型。Objective To establish a cell line that responds to ghrelin,and to observe the effect of ghrelin induced AgRP gene expression.Methods Western blotting was used to determine the growth hormone secretagogue receptor(GHSR)expression in GT1-7 cell line.And real-time quantitative polymerase chain reaction(RT-qPCR)was carried out to determine the AgRP gene expression upon serum starving or upon ghrelin treatment in dose and time dependent manner with serum.The measurement data were expressed as mean±standard error(x±SE)independent samples t-test was used for comparison between two groups,and ANOVA(least significant difference method)was used for two-way comparison if the overall variance of data among three or four groups is equal.Results Western blotting data revealed that GT1-7 cell line expressed GHSR protein.Furthermore,serum-starvation significantly increased AgRP mRNA(P<0.001).3 hours of ghrelin treatment increased the expression of AgRP mRNA in a non-serum-starvation experiment compared with control(P=0.002)and 1 hour of ghrelin treatment(P=0.008).Additionally,while 10 nmol/L and 50 nmol/L ghrelin increased AgPP mRNA expression in a similar attitude,the 100 nmol/L of ghrelin shown even further induction of AgPP mRNA expression in non-serum-starvation experiments in GT1-7 cell lines.Conclusion Taken together,our data identified that the hypothalamic GT1-7 cell line that responds to serum-starvation and ghrelin stimulation in a time-and dose-dependent manner in terms of AgRP mRNA expression.Thus,GT1-7 cell can be used as an in vitro model to study the regulation of AgRP by hypothalamic neurons,as well as the regulation of other metabolic functions by hypothalamic neurons,especially the molecular mechanisms behind them.

关 键 词：胃促生长素 胃促生长素受体 刺鼠相关蛋白质 下丘脑漏斗核 

分 类 号：R34[医药卫生—基础医学]