芳香烃受体AhR通过调控IgE介导的肥大细胞活化在嗜酸粒细胞性CRSwNP 发病机制中的作用

The pathogenesis of eosinophilic chronic rhinosinusitis with nasal polyps (Eos CRSwNP) is not entirely clear now. We and other researchers found that IgE-mediated mast cell activation might play an important role in the pathogenesis of Eos CRSwNP. Recent discoveries have illustrated that aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor, has immunomodulatory functions. Our previous work found that AhR is mainly expressed in mast cells of nasal polyps and AhR pathway is activated significantly in Eos CRSwNP in human, and can enhance IgE-mediated mast cell activation in vitro. These findings lead to the hypothesis that AhR might contribute to the pathogenesis of Eos CRSwNP through regulating IgE-mediated mast cell activation. This project is designed to: ①establish Eos CRSwNP mouse model with mast cell deficient mice by adoptive transferred AhR knockout mast cells and wildtype mast cells, respectively; observe mucosal inflammatory reaction in Eos CRSwNP by molecular biological, immunological and histopathological techniques; screen the down-stream target molecules of AhR in the pathogenesis of Eos CRSwNP by gene array technology; ② separate and culture human mast cells from nasal polyps and peripheral blood mononuclear cells, intervene the expression of AhR in human mast cells: AhR over-expression by plasmid transfection, AhR knockout by siRNA techniques. Observe the influence of AhR pathway in IgE-mediated mast cell activation by inhibition some of the screened down-stream targets. Coculture mast cells with other immune cells (eosinophils, naive T lymphocytes)and tissue cells (nasal epithelial cells, fibroblasts) and observe the related functional change of immune cells and tissue cells regulated by mast cells through AhR pathway. This project seeks to clarify the precise role of AhR in regulation of IgE-mediated mast cell activation and mechanisms in the pathogenesis of Eos CRSwNP, in-depth understanding of AhR function, and might be helpful for discovering novel therapeutic targets in this disease.

嗜酸粒细胞性伴鼻息肉的慢性鼻-鼻窦炎（Eos CRSwNP）的发病机制尚不清楚。IgE介导的肥大细胞活化可能在Eos CRSwNP的发病中起重要作用。芳香烃受体（AhR）是新近发现的免疫调控分子。前期发现AhR主要表达于鼻黏膜肥大细胞，Eos CRSwNP中AhR激活增多，体外实验AhR能增强IgE介导的肥大细胞活化。据此推测AhR通过调控IgE介导的肥大细胞活化参与Eos CRSwNP的发病。拟①体内实验：AhR敲除和未敲除的肥大细胞分别过继转移至肥大细胞缺陷小鼠建立Eos CRSwNP 模型，证实AhR在Eos CRSwNP 发病中的作用，基因芯片筛选下游靶基因；②体外实验：分离培养肥大细胞，干预AhR的表达，揭示AhR调控IgE介导的肥大细胞活化的细胞内机制；与免疫细胞或组织结构细胞共培养，明确AhR通过调控IgE介导的肥大细胞活化参与Eos CRSwNP发病的精确分子机制。

芳香烃受体（aryl hydrocarbon receptor，AhR）是一种配体激活的核转录因子，近来发现其在慢性炎症及免疫过程中发挥重要作用。我们发现AhR在慢性鼻窦炎伴鼻息肉（CRSwNP）鼻粘膜上皮细胞（HNECs）和肥大细胞（MCs）中表达增高，提示其可能在CRSwNP的炎症过程起重要作。我们通过气液交界面（ALI）培养HNECs构建体外模型，使用含有AhR配体的空气污染物颗粒PM10刺激鼻黏膜上皮细胞，发现AhR通路的激活可以损伤上皮细胞紧密连接的功能，增加上皮细胞接触外界致病原，参与CRSwNP的发病过程；进一步通过单细胞测序发现上皮基底细胞和分泌细胞的分化明显影响，Suprabasal细胞的功能延迟，导致鼻黏膜上皮的纤毛细胞发育出现较晚，对各个细胞群之间进行KEGG通路分析，发现分泌细胞和鳞状细胞出现了抗病毒能力的减弱，BMP/TGF-β和WNT信号通路在AhR介导的上皮细胞分化中起到重要作用。另外还发现，CRSwNP患者鼻黏膜上皮细胞的葡萄糖摄取量、糖酵解和三羧酸循环代谢物、代谢酶以及细胞外酸化率和氧耗率均有所增加，且以糖酵解为主。AhR-葡萄糖转运蛋白(GLUT)通路在鼻黏膜上皮中的糖代谢过程发挥一定作用，AhR可以调节GLUT的表达和活性，从而影响细胞对葡萄糖的摄取和糖酵解。CRSwNP患者鼻分泌物中葡萄糖浓度增高，导致上皮细胞糖酵解增强，且引起其基因表达谱的改变，其中IL-36G显著上调。同时，我们发现乳酸转移酶（P300）在CRSwNP中表达增高，这可导致组蛋白乳酰化水平增高，提示鼻上皮细胞增强的糖酵解产生的乳酸可能通过组蛋白乳酰化修饰上调IL-36G的表达，从而调控CRSwNP的黏膜炎症。通过我们的研究，可以为CRSwNP发病机制和治疗提供新的靶点。