Department of Ophthalmology, The Affiliated Hospital of Qingdao University, Qingdao, China.
Department of Ophthalmology, Qingdao Central Hospital, The Second Clinical Hospital of Qingdao University, Qingdao, China.
Int Immunopharmacol. 2020 Nov;88:107021. doi: 10.1016/j.intimp.2020.107021. Epub 2020 Sep 22.
Mammalian Ste20-like kinase 4 (MST4), a new member of the germinal-center kinase STE20 family, was recently demonstrated to be a negative regulator of inflammation. However, whether MST4 participates in the inflammatory response to fungal infection remains unknown. Our study investigated the role and molecular mechanisms of MST4 in mice cornea and corneal epithelial cells exposed to Aspergillus fumigatus (A. fumigatus). Protein level of MST4 was detected in mice corneas and human corneal epithelial cells (HCECs) by Western blot analysis. The MST4 protein level was significantly elevated in mice corneas infected with A. fumigatus and HCECs exposed to A. fumigatus. MST4 expression was also detected in mice corneas by immunofluorescence staining. Furthermore, we found recombinant MST4 inhibited proinflammatory cytokines expressions induced by A. fumigatus at both the mRNA and protein levels in mice corneas and HCECs. To further investigate the mechanism of MST4's anti-inflammatory effect in A. fumigatus keratitis, we verified recombinant MST4 can inhibit curdlan-mediated proinflammatory cytokines production in HCECs. Surprisingly, recombinant MST4 protein downregulated A. fumigatus-induced Dectin-1 expression in both mRNA and protein levels in mice corneas. Recombinant MST4 can inhibit the mRNA expression level of Dectin-1 which was induced by curdlan in HCECs. MST4 can also inhibit the expression of Dectin-1 in mRNA levels increased by Dectin-1 overexpression plasmid in HCECs. Moreover, A. fumigatus or curdlan significantly induced the phosphorylation of Syk, which was consequently suppressed by recombinant MST4. Finally, recombinant MST4 promotes HCECs proliferation, which contribute to cornea wound healing. Taken together, our results provide evidences that MST4 inhibits inflammatory signaling response in A. fumigatus keratitis by downregulating Dectin-1/p-Syk pathway and simultaneously promotes HCECs proliferation.
哺乳动物 Ste20 样激酶 4（MST4）是生发中心激酶 STE20 家族的新成员，最近被证明是炎症的负调节剂。然而，MST4 是否参与真菌感染的炎症反应尚不清楚。我们的研究调查了 MST4 在感染烟曲霉（A. fumigatus）的小鼠角膜和角膜上皮细胞中的作用和分子机制。通过 Western blot 分析检测小鼠角膜和人角膜上皮细胞（HCEC）中的 MST4 蛋白水平。在感染 A. fumigatus 的小鼠角膜和暴露于 A. fumigatus 的 HCEC 中，MST4 蛋白水平显著升高。通过免疫荧光染色也在小鼠角膜中检测到 MST4 表达。此外，我们发现重组 MST4 抑制了 A. fumigatus 在小鼠角膜和 HCEC 中诱导的促炎细胞因子的表达，无论是在 mRNA 还是蛋白水平上。为了进一步研究 MST4 在烟曲霉角膜炎中的抗炎作用机制，我们验证了重组 MST4 可以抑制 HCEC 中 curdlan 介导的促炎细胞因子产生。令人惊讶的是，重组 MST4 蛋白在小鼠角膜中下调了 curdlan 诱导的 Dectin-1 在 mRNA 和蛋白水平上的表达。重组 MST4 可以抑制 curdlan 在 HCEC 中诱导的 Dectin-1 的 mRNA 表达水平。MST4 还可以抑制 HCEC 中转染 Dectin-1 过表达质粒后 Dectin-1 在 mRNA 水平上的表达。此外，A. fumigatus 或 curdlan 显著诱导了 Syk 的磷酸化，而重组 MST4 则抑制了这一过程。最后，重组 MST4 促进了 HCEC 的增殖，这有助于角膜伤口愈合。综上所述，我们的研究结果表明，MST4 通过下调 Dectin-1/p-Syk 通路抑制烟曲霉角膜炎中的炎症信号反应，同时促进 HCEC 的增殖。
