Kim Charm, Park Jin Hwan, Choi Yeon Jeong, Jun Hyung Oh, Chung Jin Kwon, Park Tae Kwann, Yoon Jin Sook, Yang Jae Wook, Jang Sun Young
Department of Ophthalmology, Soonchunhyang University Cheonan Hospital, Soonchunhyang University College of Medicine, Cheonan, Republic of Korea.
Department of Ophthalmology, Soonchunhyang University Bucheon Hospital, Soonchunhyang University College of Medicine, Bucheon, Republic of Korea.
Front Endocrinol (Lausanne). 2024 Aug 30;15:1420024. doi: 10.3389/fendo.2024.1420024. eCollection 2024.
Bruton's tyrosine kinase (BTK) and interleukin (IL)-2 Inducible T-cell Kinase (ITK) inhibitors have anti-inflammatory properties. We investigated the therapeutic effect of ibrutinib, an orally bioavailable BTK/ITK inhibitor, in a mouse model of Graves' orbitopathy (GO).
Genetic immunization was performed through intramuscular administration of the recombinant plasmid, pCMV6-hTSHR cDNA, to 8-week-old female BALB/c mice. Serum levels of T3, T4, and thyroid-stimulating hormone receptor (TSHR) antibodies (TRAbs) were quantified using enzyme-linked immunosorbent assay. Histopathological changes in orbital tissues were examined using immunohistochemistry (IHC) staining for TSHR and various inflammatory markers. Following successful genetic immunization, ibrutinib was orally administered daily for 2 weeks in the GO model mice. After treatment, the mRNA and protein expression levels of BTK, ITK, IL-1β, and IL-6 in orbital tissues were evaluated using real-time PCR and Western blotting.
In total, 20 mice were sacrificed to confirm successful genetic immunization. The GO mouse group exhibited significantly increased serum T3, T4, and TRAb levels. IHC revealed increased expression of TSHR, IL-1β, IL-6, transforming growth factor-β1, interferon-γ, CD40, CD4, BTK, and ITK in the GO mouse model. The orbital inflammation was significantly attenuated in ibrutinib-treated mice. The mRNA and protein expression levels of BTK, ITK, IL-1β, and IL-6 in orbital tissue were lower in ibrutinib-treated GO mouse group compared to the phosphate-buffered saline-treated GO mouse group.
The GO mouse model demonstrated enhanced BTK and ITK expression. Ibrutinib, a BTK/ITK inhibitor, suppressed the inflammatory cytokine production. These findings highlight the potential involvement of BTK/ITK in the inflammatory pathogenesis of GO, suggesting its role as a novel therapeutic target.
布鲁顿酪氨酸激酶（BTK）和白细胞介素（IL）-2诱导型T细胞激酶（ITK）抑制剂具有抗炎特性。我们研究了口服生物利用度高的BTK/ITK抑制剂依鲁替尼在格雷夫斯眼病（GO）小鼠模型中的治疗效果。
通过肌肉注射重组质粒pCMV6-hTSHR cDNA对8周龄雌性BALB/c小鼠进行基因免疫。使用酶联免疫吸附测定法定量血清中T3、T4和促甲状腺激素受体（TSHR）抗体（TRAb）的水平。使用TSHR和各种炎症标志物的免疫组织化学（IHC）染色检查眼眶组织的组织病理学变化。在成功进行基因免疫后，对GO模型小鼠每日口服依鲁替尼，持续2周。治疗后，使用实时PCR和蛋白质免疫印迹法评估眼眶组织中BTK、ITK、IL-1β和IL-6的mRNA和蛋白质表达水平。
总共处死20只小鼠以确认基因免疫成功。GO小鼠组血清T3、T4和TRAb水平显著升高。免疫组化显示GO小鼠模型中TSHR、IL-1β、IL-6、转化生长因子-β1、干扰素-γ、CD40、CD4、BTK和ITK的表达增加。依鲁替尼治疗的小鼠眼眶炎症明显减轻。与磷酸盐缓冲盐水治疗的GO小鼠组相比，依鲁替尼治疗的GO小鼠组眼眶组织中BTK、ITK、IL-1β和IL-6的mRNA和蛋白质表达水平较低。
GO小鼠模型显示BTK和ITK表达增强。BTK/ITK抑制剂依鲁替尼抑制了炎性细胞因子的产生。这些发现突出了BTK/ITK在GO炎性发病机制中的潜在参与，表明其作为新型治疗靶点的作用。
