二肽酰肽酶-4抑制剂通过PKA/NF-κB信号通路调节LPS诱导的RAW264.7细胞炎症反应的实验研究  

DPP-4inhibitor modulates LPS-induced inflammation via PKA/NF-κB signaling pathway in RAW264.7 cells

作　　者：黄玉芳[1] 邓文[2] 朱涛[3] 

机构地区：[1]遂宁市中心医院呼吸中心,四川遂宁629000 [2]遂宁市中心医院骨科中心,四川遂宁629000 [3]四川大学华西医院呼吸病学研究室/呼吸内科,四川成都610041

出　　处：《西部医学》2016年第1期28-31,35,共5页Medical Journal of West China

基　　金：中国博士后科学基金(2014M552369);四川省医学会科研课题(171140342)

摘　　要：目的探讨二肽酰肽酶-4(DPP-4)竞争性抑制剂沙格列汀对LPS诱导的小鼠巨噬细胞(RAW264.7细胞)炎症反应的调节作用及相关的分子机制。方法将RAW264.7细胞分为4组,分别为对照组(Control组)、LPS组、LPS+沙格列汀(LPS+SAX组)和LPS+沙格列汀+PKA抑制剂H-89组(LPS+SAX+H-89组)。在干预6小时后使用qPCR法和western blot法对细胞ICAM-1mRNA和蛋白的表达水平进行检测;并采用western blot法对细胞PKA和NF-κB p65活化水平进行分析。结果与Control组相比较,在LPS干预6小时后RAW264.7细胞ICAM-1mRNA和蛋白的表达水平及NF-κB p65活化水平均明显上升,而PKA活化水平明显降低,差异均有统计学意义(P均<0.05);但LPS组较LPS+SAX组ICAM-1表达和NF-κB p65活化水平的增加更明显,而PKA活化下降更加明显,差异均有统计学意义(P均<0.05)。同时发现在LPS诱导状态下沙格列汀对于ICAM-1表达和NF-κB p65活化水平的抑制作用和PKA活化水平的促进作用可被PKA抑制剂H-89显著拮抗,差异均有统计学意义(P均<0.05)。结论本研究显示DPP-4抑制剂可以通过调控PKA/NF-κB信号通路下调LPS诱导的小鼠巨噬细胞系RAW264.7细胞的炎症反应,为DPP-4抑制剂应用于炎症性疾病的治疗奠定了初步的理论基础。Objective To explore the anti inflammatory property of DPP-4 （dipeptidyl peptidase-4） inhibitor saxa- gliptin in LPS-induced inflammation in RAW264.7 cells and its underlying mechanism. Methods RAW264.7 cells were separated into Control group, LPS group, LPS＋saxagliptin group （LPS＋SAX group） and LPS＋saxagliptin＋PKA in hibitor H-89 group （LPS＋ SAX＋ H-89 group）. After 6 hours interventions, qPCR and Western blot were used to detect the mRNA and protein expression of ICAM-1. Then, the phosphorylation of PKA and NF-κB p65 were also measured by western blot. Results After 6 hours LPS stimulation, the mRNA and protein expression of ICAM-1 was significantly up-regulated, the phosphorylation of NF-κB p65 was markedly enhanced and the phosphoryiation of PKA was largely re duced in RAW264.7 cells. Our data showed that LPS-induced expression of ICAM-1, LPS-enhanced phosphorylation of NF-κB p65 and LPS-reduced the phosphorylation of PKA were all substantially improved by saxagliptin. Meanwhile, we also found that these effects of saxagliptin were noticeably abrogated by PKA inhibitor H-89 in RAW264.7 cells. Conclu- sions DPP-4 inhibitor saxagliptin could attenuate LPS-induced inflammation via modulation of PKA/NF-κB signaling pathway in RAW264.7 cells.

关 键 词：沙格列汀 RAW264.7细胞 细胞内细胞黏附分-1(ICAM-1) 蛋白激酶A(PKA) 核转录因子-κB(NF-κB) 

分 类 号：R446.62[医药卫生—诊断学]