Fan Wenjun, Liu Peiqiang, Tan Lu, Lv Hao, Zhou Huiqin, Tao Zezhang, Xu Yu
Department of Otolaryngology-Head and Neck Surgery, Renmin Hospital of Wuhan University, Wuhan, China.
Department of Otolaryngology-Head and Neck Surgery, Renmin Hospital of Wuhan University, Wuhan, China.
Int Immunopharmacol. 2024 Dec 25;143(Pt 3):113495. doi: 10.1016/j.intimp.2024.113495. Epub 2024 Oct 31.
Allergic rhinitis (AR) represents a hallmark of obvious hypersensitivity with an imbalance of immune responses, including abnormal macrophage activity in local tissues. It has been reported that alternatively activated macrophages (M2) may contribute to allergic pathogenesis. Ten-eleven translocation (Tet) enzymes can oxidize 5-methylcytosine (m5C) in mRNA, implying the epigenetic regulation of post-transcriptional RNA modification. Our previous study suggested that decreased Tet2 impairs the function of regulatory T cells, failing to exert a protective role in AR. However, the mechanism of Tet2 in macrophage polarization has been little discussed. In this paper, we investigate the regulatory role of Tet2 in macrophage polarization under allergic inflammation.
Macrophage immunofluorescence and eosinophil counts were used to confirm the inflammatory and polarized state in the nasal mucosa of AR patients. Additionally, we used Raw264.7 cells to explore the relationships among mRNA methylation, Tet2 expression, and the macrophage polarization process. Furthermore an Ovalbumin (OVA)-mediated AR mouse model was established with wild-type (WT) and Tet2 gene knockout (Tet2) mice to verify the role of Tet2 in AR severity and macrophage polarization. The final stage comprised RNA sequencing, methylated RNA immunoprecipitation with qPCR (MeRIP-qPCR) using bone marrow-derived macrophages (BMDMs) from WT and Tet2 mice to explore the effect of Tet2 deficiency on the mRNA methylation level of M2-related genes under OVA treatment. A two-tailed Student's t-test was used to compare two groups, and Spearman correlation analysis was applied for relationship analysis.
M2-macrophages were confirmed as the dominant subtype associated with eosinophil levels in AR nasal tissues. In vitro analyses demonstrated that mRNA methylation and Tet2 are linked to M2 macrophages. Additionally, we found that Tet2 influences local allergic severity and macrophage polarization. Specifically, Tet2 deficiency decreased the mRNA m5C demethylation levels of Klf4 and Rock1, contributing to M2 polarization in an allergic state.
The findings of this study demonstrate that Tet2 may play a protective role in AR by negatively regulating M2-related factors through mRNA m5C demethylation. These findings provide new insights into AR therapy, suggesting that intervening in macrophage polarization at the post-transcriptional level could be a novel therapeutic strategy.
过敏性鼻炎（AR）是明显超敏反应的标志，伴有免疫反应失衡，包括局部组织中异常的巨噬细胞活性。据报道，交替活化的巨噬细胞（M2）可能参与过敏性发病机制。10-11易位（Tet）酶可氧化mRNA中的5-甲基胞嘧啶（m5C），这意味着转录后RNA修饰的表观遗传调控。我们之前的研究表明，Tet2表达降低会损害调节性T细胞的功能，无法在AR中发挥保护作用。然而，Tet2在巨噬细胞极化中的机制鲜有讨论。在本文中，我们研究了Tet2在过敏性炎症下巨噬细胞极化中的调节作用。
采用巨噬细胞免疫荧光和嗜酸性粒细胞计数来确认AR患者鼻黏膜中的炎症和极化状态。此外，我们使用Raw264.7细胞来探索mRNA甲基化、Tet2表达与巨噬细胞极化过程之间的关系。此外，建立了卵清蛋白（OVA）介导的AR小鼠模型，使用野生型（WT）和Tet2基因敲除（Tet2-/-）小鼠来验证Tet2在AR严重程度和巨噬细胞极化中的作用。最后阶段包括RNA测序、使用来自WT和Tet2-/-小鼠的骨髓来源巨噬细胞（BMDM）进行甲基化RNA免疫沉淀-qPCR（MeRIP-qPCR），以探索Tet2缺陷对OVA处理下M2相关基因mRNA甲基化水平的影响。采用双侧Student t检验比较两组，并应用Spearman相关性分析进行关系分析。
M2巨噬细胞被确认为与AR鼻组织中嗜酸性粒细胞水平相关的主要亚型。体外分析表明，mRNA甲基化和Tet2与M2巨噬细胞有关。此外，我们发现Tet2影响局部过敏严重程度和巨噬细胞极化。具体而言，Tet2缺陷降低了Klf4和Rock1的mRNA m5C去甲基化水平，导致过敏状态下的M2极化。
本研究结果表明，Tet2可能通过mRNA m5C去甲基化负向调节M2相关因子，从而在AR中发挥保护作用。这些发现为AR治疗提供了新的见解，表明在转录后水平干预巨噬细胞极化可能是一种新的治疗策略。
