Wang Li, Luo Jianmin, Fu Yiling, He Shaoheng
Allergy and Inflammation Research Institute, The Key Immunopathology Laboratory of Guangdong Province, The Shantou University Medical College, Shantou 515031, China.
Int J Biochem Cell Biol. 2006;38(9):1571-83. doi: 10.1016/j.biocel.2006.03.016. Epub 2006 Apr 18.
It was reported that thrombin could induce IL-8 secretion from human dermal fibroblasts (HDFs) through activation of proteinase activated receptor (PAR)-1. However, little is known of intracellular signaling pathways involved in the event. In the present study, expression of PARs in primarily cultured HDFs was determined by flow cytometry analysis and reverse transcription polymerase chain reaction (RT-PCR), levels of IL-8 were determined by using ELISA and signaling pathways were examined by using Western blot. It was found that HDFs express PAR-1 and PAR-3, and thrombin induces approximately 7.4-fold increase in IL-8 secretion from HDFs. Hirudin and a PAR-1 blocking antibody completely abolish the action of thrombin. It was also found that PD98059, a mitogen-activated protein kinase (MAPK) pathway inhibitor and U0126, an inhibitor of extracellular signal-regulated kinase (ERK) blocks thrombin-induced phosphorylation of ERK1/2 and IL-8 secretion, indicating the involvement of MAPK/ERK signaling pathway in thrombin-induced IL-8 secretion. p38 MAPK pathway appears also being involved as SB203580, a selective inhibitor of p38 MAPK inhibit phosphorylation of p38 MAPK and thrombin-induced IL-8 secretion. Furthermore, Janus kinase (JAK)/signal transducer and activator of transcription 3 (STAT3) pathway, but not phosphotidylinositol 3-kinase (PI3K)/Akt signaling pathway may also be activated by thrombin. In conclusion, thrombin potently induce IL-8 release via PAR-1 from HDFs. Thrombin elicited IL-8 release is predominantly conducted through MAPK/ERK and p38 MAPK signaling pathways. Discovery of the signaling pathways of thrombin in HDFs may help to understand the role of thrombin in inflammation and tissue remodeling.
据报道，凝血酶可通过激活蛋白酶激活受体（PAR）-1诱导人皮肤成纤维细胞（HDFs）分泌白细胞介素-8（IL-8）。然而，对于该事件所涉及的细胞内信号通路却知之甚少。在本研究中，通过流式细胞术分析和逆转录聚合酶链反应（RT-PCR）测定原代培养的HDFs中PARs的表达，使用酶联免疫吸附测定（ELISA）测定IL-8水平，并通过蛋白质印迹法检测信号通路。结果发现，HDFs表达PAR-1和PAR-3，凝血酶可诱导HDFs分泌的IL-8增加约7.4倍。水蛭素和一种PAR-1阻断抗体可完全消除凝血酶的作用。还发现，丝裂原活化蛋白激酶（MAPK）途径抑制剂PD98059和细胞外信号调节激酶（ERK）抑制剂U0126可阻断凝血酶诱导的ERK1/2磷酸化和IL-8分泌，表明MAPK/ERK信号通路参与了凝血酶诱导的IL-8分泌。p38 MAPK途径似乎也参与其中，因为p38 MAPK的选择性抑制剂SB203580可抑制p38 MAPK的磷酸化以及凝血酶诱导的IL-8分泌。此外，Janus激酶（JAK）/信号转导和转录激活因子3（STAT3）途径可能也被凝血酶激活，但磷脂酰肌醇3-激酶（PI3K）/蛋白激酶B（Akt）信号通路未被激活。总之，凝血酶可通过PAR-1从HDFs中有效诱导IL-８释放。凝血酶引发的IL-８释放主要通过MAPK/ERK和p38 MAPK信号通路进行。发现凝血酶在HDFs中的信号通路可能有助于理解凝血酶在炎症和组织重塑中的作用。
