DUSP8过表达通过JNK/p38 MAPK通路来改善LPS诱导的巨噬细胞炎症反应

双特异性磷酸酶8（dual-specificity phosphatase 8, DUSP8）是双特异性蛋白磷酸酶家族的成员之一，被报道参与多个疾病发生过程。然而，DUSP8是否参与巨噬细胞等免疫细胞炎性应答过程，目前仍未有研究证实。本研究旨在检测DUSP8在脂多糖(LPS)诱导的巨噬细胞炎症反应中的表达，并探讨过表达DUSP8在巨噬细胞炎症反应的作用。利用100 ng/mL LPS刺激野生型C57BL/6小鼠骨髓来源巨噬细胞（bone marrow derived macrophage，BMDM），分别在不同时间点收取细胞，实时PCR和Western 印迹检测发现LPS处理后，BMDM中DUSP8的表达水平明显降低（P<0.05），且在12 h达到最低值；随后，分别转染DUSP8过表达载体（DUSP8-EGFP）和对照载体（EGFP）于BMDM，Western 印迹检测发现DUSP8-EGFP转染能够显著上调DUSP8的表达水平（P<0.05）；进一步用流式细胞术(flow cytometry, FCM)检测发现DUSP8过表达使巨噬细胞表面分子CD80和CD86的表达显著下调（P<0.05）；同时，中性红吞噬实验结果显示，DUSP8过表达后巨噬细胞的吞噬能力明显降低（P<0.05）；此外，ELISA (enzyme linked immunosorbent assay)检测结果显示，过表达DUSP8显著降低IL-1β，IL-6的表达水平（P<0.05）；最后，Western 印迹结果显示，JNK和p38 MAPK的磷酸化水平在DUSP8过表达组中明显降低（P<0.05）。以上表明，DUSP8过表达可显著改善LPS诱导的巨噬细胞炎症反应，其机制主要通过抑制JNK和p38 MAPK的活化。

DUSP8 Overexpression Alleviates LPS-induced Macrophage Inflammatory Response Through JNK/p38 MAPK Pathways

Dual-specificity phosphatase 8 (DUSP8) is a member of the dual-specificity phosphatase family, which has been reported to participate in the development of many diseases. However, it is unclear whether DUSP8 plays an important role in the inflammatory response of macrophages and related immune cells. This study aims to detect the expression of DUSP8 in lipopolysaccharide (LPS)-induced macrophage inflammatory responses and to observe the effect of DUSP8 overexpression on macrophage inflammatory responses. 100 ng/ml LPS was used to treat bone marrow-derived macrophages (BMDMs) from wild-type C57BL/6 mice at different time points. Real-time PCR and Western blotting results showed that the expression of DUSP8 was significantly reduced in BMDMs (P<0.05) and reached the peak at 12 hours. Next, DUSP8 overexpression vectors (DUSP-EGFP) and control vectors (EGFP) were transfected into BMDM. Data showed that DUSP-EGFP transfection could significantly enhance the expression of DUSP8 in BMDMs (P<0.05). Moreover, FCM data showed that the expressions of CD80 and CD86 markedly decreased in BMDMs with DUSP8 overexpression (P<0.05). Meanwhile, the neutral red uptake assay data showed that the uptake in DUSP8 overexpression group was lower than that in the control group. Furthermore, ELISA results showed DUSP8 overexpression could significantly reduce the production of IL-1β and IL-6 (P<0.05). Besides, Western blotting results showed that the phosphorylation levels of p38 MAPK and JNK decreased in BMDMs after DUSP8 overexpression (P<0.05). All together, DUSP8 overexpression could significantly ameliorate LPS-induced macrophage inflammation, which was mainly related to the reduced expression of phosphorylated p38 MAPK and JNK.