Ebenezer Angel Jemima, Prasad Kavya, Rajan Sanjana, Thangam Elden Berla
a Department of Biotechnology, School of Bioengineering , SRM University , Kattankulathur , India.
J Recept Signal Transduct Res. 2018 Jun;38(3):204-212. doi: 10.1080/10799893.2018.1468783. Epub 2018 Jun 4.
Mast cell (MC) activation through H4R releases various inflammatory mediators which are associated with allergic asthma.
To investigate the siRNA-mediated gene silencing effect of H4R on human mast cells (HMCs) functions and the activation of stress-activated protein kinases (SAPK)/jun amino-terminal kinases (JNK) signaling pathways for the release of ineterleukin-1β (IL-1β) in HMCs.
H4R expression was analyzed by RT-PCR and western blotting in human mast cell line-1 (HMC-1) cells and H4RsiRNA transfected cells. The effect of H4RsiRNA and H4R-antagonist on H4R mediated MC functions such as intracellular Ca release, degranulation, IL-6 and IL-1β release, and the activation SAPK/JNK signaling pathways were studied. HMC-1 cells were stimulated with 10 μM of histamine (His) and 4-methylhistamine (4-MH) and pretreated individually with H4R-antagonist JNJ7777120 (JNJ), histamine H1 receptor (H1R)-antagonist mepyramine, and signaling molecule inhibitors SP600125 (SP) and Bay117082.
We found that the HMC-1 cells expressed H4R and H4RsiRNA treatment down regulated the H4R expression in HMC-1 cells. Both His and 4-MH induced the intracellular Ca release and degranulation whereas; H4R siRNA and JNJ inhibited the effect. Furthermore, the activation of H4R caused the phosphorylation of SAPK/JNK pathways. H4R gene silencing and pretreatment with SP and JNJ decreased His and 4-MH induced phosphorylation of SAPK/JNK. We found that the activation of H4R caused the release of IL-1β (124.22 pg/ml) and IL-6 (122.50 pg/ml) on HMC-1 cells. Whereas, SAPK/JNK inhibitor (68.36 pg/ml) inhibited the H4R mediated IL-1β release.
Taken together, the silencing of H4R inhibited the H4R mediated MC functions and SAPK/JNK phosphorylation. Furthermore, the H4R activation utilized SAPK/JNK signaling pathway for IL-1β release in HMC-1 cells.
通过H4R激活肥大细胞（MC）会释放多种与过敏性哮喘相关的炎症介质。
研究小干扰RNA（siRNA）介导的H4R基因沉默对人肥大细胞（HMC）功能的影响，以及应激激活蛋白激酶（SAPK）/c-Jun氨基末端激酶（JNK）信号通路的激活对HMC中白细胞介素-1β（IL-1β）释放的影响。
通过逆转录聚合酶链反应（RT-PCR）和蛋白质免疫印迹法分析人肥大细胞系-1（HMC-1）细胞和转染H4R siRNA的细胞中H4R的表达。研究了H4R siRNA和H4R拮抗剂对H4R介导的MC功能的影响，如细胞内钙释放、脱颗粒、IL-6和IL-1β释放，以及SAPK/JNK信号通路的激活。用10μM组胺（His）和4-甲基组胺（4-MH）刺激HMC-1细胞，并分别用H4R拮抗剂JNJ7777120（JNJ）、组胺H1受体（H1R）拮抗剂美吡拉敏以及信号分子抑制剂SP600125（SP）和Bay117082进行预处理。
我们发现HMC-1细胞表达H4R，H4R siRNA处理下调了HMC-1细胞中H4R的表达。His和4-MH均诱导细胞内钙释放和脱颗粒，而H4R siRNA和JNJ抑制了这种作用。此外，H4R的激活导致SAPK/JNK通路的磷酸化。H4R基因沉默以及用SP和JNJ预处理可降低His和4-MH诱导的SAPK/JNK磷酸化。我们发现H4R的激活导致HMC-1细胞释放IL-1β（124.22 pg/ml）和IL-6（122.50 pg/ml）。而SAPK/JNK抑制剂（68.36 pg/ml）抑制了H4R介导的IL-1β释放。
综上所述，H4R的沉默抑制了H4R介导的MC功能和SAPK/JNK磷酸化。此外，H4R激活利用SAPK/JNK信号通路促进HMC-1细胞中IL-1β的释放。
