阿米洛利抑制载脂蛋白A1诱导小鼠巨噬细胞ABCA1的降解  

Amiloride Inhibits the Degradation of Apolipoprotein A1-induced ABCA1 in Mice Macrophage

作　　者：莫显刚[1] 王兰[1] 洪伟[2] 张莉[1] 代陆军[3] 蒋金[3] 

机构地区：[1]贵州医科大学附属医院老年病科,贵州省贵阳市550004 [2]贵州医科大学分子生物学重点实验室,贵州省贵阳市550004 [3]贵州医科大学附属医院病理科,贵州省贵阳市550004

出　　处：《中国动脉硬化杂志》2016年第3期234-238,共5页Chinese Journal of Arteriosclerosis

基　　金：国家自然科学基金(31260250)

摘　　要：目的探讨钠氢交换体1抑制剂阿米洛利对载脂蛋白A1(Apo A1)诱导小鼠三磷酸腺苷结合盒转运体A1(ABCA1)降解的影响。方法给小鼠静脉注射Apo A1后不同时间点收集腹腔巨噬细胞,定量实时聚合酶链反应及Western blot检测巨噬细胞ABCA1 mRNA及蛋白水平变化。进而,Apo A1干预8 h小鼠给予腹腔注射阿米洛利或钙蛋白酶抑制剂ALLN,实验分为4组:对照组、阿米洛利组、ALLN组及阿米洛利+ALLN组,Western blot检测巨噬细胞ABCA1蛋白水平,荧光法检测钙蛋白酶活性。结果 Apo A1干预小鼠后ABCA1 mRNA无明显改变,而ABCA1蛋白水平迅速升高,8 h达高峰后逐渐降低。阿米洛利组、ALLN组及阿米洛利+ALLN组ABCA1蛋白水平均比对照组高;在0、4、8 h时间点,阿米洛利组、ALLN组及阿米洛利+ALLN组间ABCA1蛋白水平差异无统计学意义;而在12、16 h时间点,阿米洛利+ALLN组ABCA1蛋白水平较阿米洛利组、ALLN组升高。与对照组比较,阿米洛利组、ALLN组及阿米洛利+ALLN组钙蛋白酶活性均降低;阿米洛利组、ALLN组、阿米洛利+ALLN组3组组间比较,钙蛋白酶活性各时间点均无明显差异。结论阿米洛利在活体内抑制Apo A1诱导ABCA1蛋白降解及钙蛋白酶活性,提示钠氢交换体1可能至少部分通过改变钙蛋白酶活性参与ABCA1的降解。Aim To investigate the effect of Na＋/ H＋exchanger-1（ NHE1） inhibitor amiloride on the degradation of apolipoprotein A1（ Apo A1）-induced ATP binding cassette transporter A1（ ABCA1） in the mice macrophages.Methods The peritoneal macrophages were collected from the mice at various indicated time points after intravenous injection of Apo A1. ABCA1 mRNA and protein levels in the macrophages were determined using quantitative real-time polymerase chain reaction（ qRT-PCR） and Western blot. Furthermore,the mice intervened with Apo A1 for 8 h were intraperitoneally injected with amiloride or calpain inhibitor ALLN. The treated mice were divided into four groups： control group,amiloride group,ALLN group and amiloride ＋ ALLN group. ABCA1 protein was detected by Western blot and calpain activity was assessed by fluorescence method in macrophages. Results After mice intervened with Apo A1,ABCA1 mRNA had no obvious change,but the level of ABCA1 protein increased rapidly,and reached the peak at 8 h,then gradually declined. The ABCA1 protein levels were higher in amiloride group,ALLN group and amiloride ＋ ALLN group than that in control group. The differences of ABCA1 protein level were not statistically significant in amiloride group,ALLN group and amiloride ＋ ALLN group at 0,4 and 8 h. Whereas the ABCA1 protein level in amiloride ＋ ALLN group was higher than those in amiloride group and ALLN group at 12 and 16 h. Compared with the control group,calpain activities were decreased in amiloride group,ALLN group and amiloride ＋ ALLN group. There were no obvious differences in calpain activity among amiloride group,ALLN group and amiloride ＋ ALLN group at each time point. Conclusions Amiloride inhibits the degradation of Apo A1-induced ABCA1 and calpain activity in mice peritoneal macrophages in vivo. It indicates that NHE1 may be involved in ABCA1 degradation at least in part by changing the activity of calpain.

关 键 词：阿米洛利 钠氢交换体1 载脂蛋白A1 三磷酸腺苷结合盒转运体A1 钙蛋白酶 ALLN 

分 类 号：Q46[生物学—生理学]