Loss of the adhesion G-protein coupled receptor ADGRF5 in mice induces airway inflammation and the expression of CCL2 in lung endothelial cells
Loss of the adhesion G-protein coupled receptor ADGRF5 in mice induces airway inflammation and the expression of CCL2 in lung endothelial cells
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DOI：
10.1186/s12931-019-0973-6
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发表时间：
2019-01-17
影响因子：
5.8
通讯作者：
Nakamura, Nobuhiro
中科院分区：
文献类型：
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作者：
Kubo, Fumimasa;Ariestanti, Donna Maretta;Nakamura, Nobuhiro
黏附G蛋白偶联受体F5(ADGRF5)是肺泡II型细胞肺表面活性物质动态平衡的重要调节因子。我们之前的研究表明，除了表面活性物质的异常积聚，Adgrf5缺陷(Adgrf5(-/-))小鼠还表现出肺气肿样症状，这表明ADGRF5可能在免疫调节中发挥作用。在这里，我们扩展了Adgrf5(-/-)小鼠的表型分析，以帮助了解它在肺中的生物学作用，特别是在免疫调节中的作用。方法用阿尔辛蓝和马森三色染色评价肺的组织学特征。采用实时定量聚合酶链式反应(QPCR)和免疫印迹方法分析野生型和Adgrf5(-/-)小鼠肺组织中与呼吸道炎症相关的基因/蛋白的差异表达。通过进行血气测试和尿液pH测量来评估酸碱状态。用Giemsa染色的支气管肺泡灌洗液细胞进行炎症细胞计数。采用双抗体夹心酶联免疫吸附试验测定血清IgE浓度。采用定量聚合酶链式反应和/或免疫印迹法检测原代肺内皮细胞CCL2、S100A8、S100A9和Saa3的表达。结果Adgrf5(-/-)小鼠表现出慢性呼吸道炎症(粘液细胞化生、粘液高分泌、上皮下纤维化、呼吸性酸中毒、血清高IgE、肥大细胞聚集和中性粒细胞增多)的一些特征，同时与粘液细胞化生(Muc5ac、MUC5B、SLC26A4和CLCA1)、纤维化(Tgfb1、Col1a1、Fn1和TNC)和2型免疫反应(IL4、IL5、IL13、IL-25、IL-25)相关的基因表达增加和IL-33)，在12和/或30周龄。相反，CCL2、S100A8和S100A9在胚胎或新生Adgrf5(-/-)小鼠肺以及1周龄Adgrf5(-/-)小鼠肺内皮细胞中表达上调。RS504393可抑制Adgrf5(-/-)肺S100A8、S100A9、SLC26A4和IL5的表达上调。结论靶向干扰ADGRF5可导致气道炎症的发生，其机制可能与2型免疫反应和CCL2介导的炎症反应有关。ADGRF5还在肺内皮细胞CCL2编码基因的调控中发挥潜在作用，从而维持免疫平衡。
BackgroundAdhesion G-protein coupled receptor F5 (ADGRF5) was recently identified as an essential regulator of pulmonary surfactant homeostasis in alveolar type II cells. We previously showed that in addition to abnormal surfactant accumulation, Adgrf5-deficient (Adgrf5(-/-)) mice exhibit emphysema-like signs, suggesting a possible role for ADGRF5 in immune regulation. Here, we extended the phenotypic analysis of Adgrf5(-/-) mice to help understand its biological role in the lung, and especially in immune regulation.MethodsHistological features of lungs were evaluated by Alcian blue and Masson's trichrome staining. Quantitative real-time PCR (qPCR) and western blot analyses were performed to analyze the differential expression of genes/proteins related to airway inflammation in lungs between wildtype and Adgrf5(-/-) mice. Acid-base status was assessed by performing blood gas tests and urine pH measurements. Inflammatory cell counting was performed using Giemsa-stained bronchoalveolar lavage cells. Serum IgE concentrations were determined by enzyme-linked immunosorbent assay. The expression of Ccl2, S100a8, S100a9, and Saa3 in primary lung endothelial cells (ECs) was determined by qPCR and/or western blotting. Finally, the effect of administrating RS504393 to 2-week-old Adgrf5(-/-) mice on gene expression in the lungs was analyzed by qPCR.ResultsAdgrf5(-/-) mice exhibited several features of chronic airway inflammation (mucous cell metaplasia, mucus hyperproduction, subepithelial fibrosis, respiratory acidosis, high serum IgE, mast cell accumulation, and neutrophilia) in parallel with elevated expression of genes involved in mucous cell metaplasia (Muc5ac, Muc5b, Slc26a4, and Clca1), fibrosis (Tgfb1, Col1a1, Fn1, and Tnc), and type 2 immune response (Il4, Il5, Il13, IL-25, and IL-33) at 12 and/or 30weeks of age. In contrast, mRNA expression of Ccl2, S100a8, and S100a9 was upregulated in embryonic or neonatal Adgrf5(-/-) lungs as well as in lung ECs of Adgrf5(-/-) mice at 1week of age. RS504393 treatment suppressed the upregulation of S100a8, S100a9, Slc26a4, and Il5 in Adgrf5(-/-) lungs.ConclusionsTargeted disruption of ADGRF5 results in the development of airway inflammation, which is likely mediated by the type 2 immune response and possibly CCL2-mediated inflammation. ADGRF5 also has a potential role in the regulation of genes encoding CCL2 in lung ECs, thereby maintaining immune homeostasis.