Department of Internal Medicine, Beijing Obstetrics and Gynecology Hospital, Capital Medical University, Beijing 100026, P.R. China.
Department of Emergency, Beijing Chaoyang Hospital (Jingxi Campus), Capital Medical University, Beijing 100043, P.R. China.
Int J Mol Med. 2018 Jan;41(1):409-420. doi: 10.3892/ijmm.2017.3252. Epub 2017 Nov 13.
The aim of the present study was to investigate the role of the angiotensin-converting enzyme (ACE)2-angiotensin‑(Ang)-(1-7)-Mas axis in the pathogenesis of pancreatitis and the association between this axis and the p38 mitogen-activated protein kinase (p38 MAPK)/nuclear factor (NF-κB) signaling pathway in pancreatic acinar cells. Mouse pancreatic acinar cancer (MPC-83) cells were stimulated with 10 nM caerulein (CAE) to create an in vitro model of acute pancreatitis, and collected for analysis at 2, 6, 12, 24 and 48 h post stimulation. In addition, cells were pretreated with different concentrations of Ang‑(1‑7), Ang‑(1‑7) antagonist A779, p38 MAPK inhibitor SB203580 or ACE2 inhibitor DX600 for 30 min, and then stimulated with CAE for 24 h. The ACE2, Mas receptor, p38 MAPK, phosphorylated (p)-p38 MAPK and NF-κB expression levels were evaluated using western blotting and immunofluorescence. p38 MAPK, NF-κB, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), IL-8 and IL-10 mRNA expression levels were assessed using reverse transcription-quantitative polymerase chain reaction. The results of the immunofluorescence assay demonstrated that ACE2 and p38 MAPK were present mainly in the cytoplasm, while the Mas receptor was located mainly in the cell membrane. ACE2, p38 MAPK and p-p38 MAPK protein levels were significantly increased (P<0.05) following stimulation with CAE compared with those in the control group and peaked at 24 h. Mas receptor protein levels were significantly upregulated (P<0.05) between 6 and 24 h, peaking at 12 h. Ang‑(1‑7) and SB203580 downregulated p-p38 MAPK and NF-κB expression and the mRNA levels of inflammatory factors IL-6, TNF-α and IL-8, but upregulated the mRNA level of inflammatory factor IL-10 compared with those treated with CAE alone. These results were supported by the opposite outcomes observed for cells treated with A779 or DX600. Therefore, it was concluded that the ACE2-Ang‑(1‑7)-Mas axis significantly inhibits pancreatitis by inhibition of the p38 MAPK/NF-κB signaling pathway.
本研究旨在探讨血管紧张素转换酶（ACE）2-血管紧张素（Ang）-（1-7）-Mas 轴在胰腺炎发病机制中的作用，以及该轴与胰腺腺泡细胞中 p38 丝裂原激活蛋白激酶（p38 MAPK）/核因子（NF-κB）信号通路之间的关联。用 10 nM 蛙皮素（CAE）刺激小鼠胰腺癌细胞（MPC-83）以建立急性胰腺炎的体外模型，并在刺激后 2、6、12、24 和 48 h 进行分析。此外，细胞用不同浓度的 Ang-（1-7）、Ang-（1-7）拮抗剂 A779、p38 MAPK 抑制剂 SB203580 或 ACE2 抑制剂 DX600 预处理 30 min，然后用 CAE 刺激 24 h。采用 Western blot 和免疫荧光法检测 ACE2、Mas 受体、p38 MAPK、磷酸化（p）-p38 MAPK 和 NF-κB 的表达水平。采用逆转录-定量聚合酶链反应法检测 p38 MAPK、NF-κB、肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）、白细胞介素-8（IL-8）和白细胞介素-10（IL-10）mRNA 的表达水平。免疫荧光试验结果表明，ACE2 和 p38 MAPK 主要位于细胞质中，而 Mas 受体主要位于细胞膜上。与对照组相比，CAE 刺激后 ACE2、p38 MAPK 和 p-p38 MAPK 蛋白水平显著升高（P<0.05），并在 24 h 时达到峰值。Mas 受体蛋白水平在 6-24 h 之间显著上调（P<0.05），在 12 h 时达到峰值。Ang-（1-7）和 SB203580 下调 p-p38 MAPK 和 NF-κB 的表达以及炎症因子 IL-6、TNF-α 和 IL-8 的 mRNA 水平，但上调炎症因子 IL-10 的 mRNA 水平与单独用 CAE 处理相比。用 A779 或 DX600 处理的细胞观察到相反的结果，支持了这些结果。因此，结论是 ACE2-Ang-（1-7）-Mas 轴通过抑制 p38 MAPK/NF-κB 信号通路显著抑制胰腺炎。
