Inhibition of Tumor Progression Locus 2 Protein Kinase Decreases Lipopolysaccharide-Induced Tumor Necrosis Factor α Production Due to the Inhibition of the Tip-Associated Protein Induction in RAW264.7 Cells
Inhibition of Tumor Progression Locus 2 Protein Kinase Decreases Lipopolysaccharide-Induced Tumor Necrosis Factor α Production Due to the Inhibition of the Tip-Associated Protein Induction in RAW264.7 Cells
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DOI：
10.1248/bpb.33.1233
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发表时间：
2010-07-01
影响因子：
2
通讯作者：
Sugita, Takahisa
中科院分区：
文献类型：
--
作者：
Hirata, Kazuya;Miyashiro, Masahiko;Sugita, Takahisa
丝裂原活化蛋白激酶（MAPKs）的激活通过介导多种炎性细胞因子的产生而在炎症事件中起关键作用。Tp12（肿瘤进展位点2）-MEK（MAPK/ERK激酶）-ERK（细胞外信号调节激酶）信号通路在脂多糖（LPS）刺激的巨噬细胞中肿瘤坏死因子α（TNFα）的产生中起重要作用。在此，我们利用一种有效的Tp12激酶抑制剂1,7 - 萘啶 - 3 - 腈以及LPS刺激的RAW264.7细胞研究了Tp12介导的TNFα产生的分子机制。该抑制剂可有效抑制体外Tp12激酶活性，并通过特异性抑制MEK和ERK的磷酸化（而非p38和c - Jun N末端激酶（JNK）的磷酸化）显著降低TNFα的产生。一种p38抑制剂SB203580也呈剂量依赖性地抑制TNFα的产生。尽管两种抑制剂均未改变TNFα mRNA水平，但Tp12抑制剂增加了细胞核内TNFα mRNA水平，同时降低了细胞质中的水平。TNFα mRNA核质转运的关键分子Tip相关蛋白（TAP）受LPS上调，但这种增加被Tp12抑制剂削弱。在所有情况下，SB203580在LPS存在时均无作用。这些结果表明，LPS通过Tp12 - MEK - ERK信号通路诱导的TNFα产生是通过在核质转运水平改变TAP水平来调控的。这些结果增进了对TNFα调控机制的理解，并可能为炎症性疾病提供一种新的治疗策略。
The activation of mitogen-activated protein kinases (MAPKs) is critically involved in inflammatory events through mediation of the production of various inflammatory cytokines. The Tp12 (tumor progression locus 2)-MEK (MAPK/ERK kinase)-ERK (extracellular signal-regulated kinase) signaling pathway plays an essential role in the production of tumor necrosis factor alpha (TNF alpha) in macrophages stimulated with lipopolysaccharide (LPS). Here, we studied the molecular mechanisms of Tp12-mediated TNFa production using a potent Tp12 kinase inhibitor, 1,7-naphtyridine-3-carbonitrile, and LPS-stimulated RAW264.7 cells. This inhibitor was effective in suppressing the in vitro Tp12 kinase activity, and caused a significant reduction in TNF alpha production via specific suppression of the phosphorylation of MEK and ERK but not that of p38 and c-Jun N-terminal kinase (JNK). A p38 inhibitor, SB203580, also inhibited the TNFa production dose-dependently. Although the TNF alpha mRNA level was not altered by either inhibitor, the Tp12 inhibitor increased the nuclear TNFa mRNA level, while decreasing that in the cytoplasm. Tip-associated protein (TAP), a key molecule in the nucleocytoplasmic transport of TNF alpha mRNA, was up-regulated by LPS, but this increase was impaired by the Tp12 inhibitor. In all cases, SB203580 was without effect in the presence of LPS. These results suggest that the LPS-induced TNF alpha production via the Tp12-MEK-ERK signaling pathway is regulated by changing the TAP level at the nucleocytoplasmic transport level. These results improve understanding of TNF alpha regulatory mechanisms and might provide a new therapeutic strategy against inflammatory diseases.