BACKGROUND Exposure to fungi, especially Aspergillus fumigatus (A.f.), can elicit potent allergic inflammation that triggers and worsens asthmatic disease. Dendritic cells (DCs), initiate allergic inflammatory responses to allergic stimuli. However, it is unclear if A.f. spores during isotropic growth (early spore swelling) can activate DCs to initiate allergic responses or if germination is required. This lack of basic understanding of how A.f. causes disease is a barrier to the development of new treatments. OBJECTIVE To show that a precise A.f. morphotype stage during spore swelling can trigger DCs to mediate allergic inflammatory responses and ascertain if antifungal therapeutics can be effective at suppressing this process. METHODS We employed an A.f. strain deficient in pyrimidine biosynthesis (ΔpyrG) to generate populations of A.f. spores arrested at different stages of isotropic growth (swelling) via temporal removal of uracil and uridine from growth media. These arrested spore stages were cultured with bone marrow derived DCs (BMDCs), and their activation measured via flow cytometry and ELISA to interrogate which growth stage was able to activate BMDCs. These BMDCs were then adoptively transferred into the airways, to assess if they were able to mediate allergic inflammation in naive recipient mice. Allergic airway inflammation in vivo was determined via flow cytometry, ELISA and qPCR. This system was also used to determine if antifungal drug (itraconazole) treatment could alter early stages of spore swelling and therefore BMDC activation and in vivo allergic inflammation upon adoptive transfer. RESULTS We found that A.f. isotropic growth is essential to trigger BMDC activation and mediate allergic airway inflammation. Furthermore, using time arrested A.f. stages, we found that least 3h in growth media enabled spores to swell sufficiently to activate BMDCs to elicit allergic airway inflammation in vivo. Incubation of germinating A.f. with itraconazole reduced spore swelling and partially reduced their ability to activate BMDCs to elicit in vivo allergic airway inflammation. CONCLUSION In summary, our results have pinpointed the precise stage of A.f. development when germinating spores are able to activate DCs to mediate downstream allergic airway inflammation. Furthermore, we have identified that antifungal therapeutics partially reduced the potential of A.f. spores to stimulate allergic responses, highlighting a potential mechanism by which antifungal treatment might help to prevent the development of fungal allergy.

中文翻译：

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曲霉菌介导的过敏性气道炎症是由树突状细胞识别确定的孢子形态型引发的。


背景 接触真菌，尤其是烟曲霉 （A.f.），会引起强烈的过敏性炎症，从而引发并加重哮喘疾病。树突状细胞 （DC） 会引发对过敏刺激的过敏性炎症反应。然而，目前尚不清楚各向同性生长（早期孢子膨胀）期间的 A.f. 孢子是否可以激活 DC 以启动过敏反应，或者是否需要发芽。对 A.f. 如何导致疾病缺乏基本了解是开发新疗法的障碍。目的 表明孢子肿胀过程中精确的 AF 形态型阶段可以触发 DC 介导过敏性炎症反应，并确定抗真菌疗法是否可以有效抑制这一过程。方法 我们采用嘧啶生物合成缺陷的 A.f. 菌株 （ΔpyrG） 通过从生长培养基中暂时去除尿嘧啶和尿苷来产生停滞在各向同性生长（肿胀）不同阶段的 A.f. 孢子种群。这些停滞的孢子阶段用骨髓来源的 DC （BMDC） 培养，并通过流式细胞术和 ELISA 测量它们的激活，以询问哪个生长阶段能够激活 BMDC。然后将这些 BMDC 过继转移到气道中，以评估它们是否能够介导幼稚受体小鼠的过敏性炎症。通过流式细胞术、 ELISA 和 qPCR 确定体内过敏性气道炎症。该系统还用于确定抗真菌药物 （伊曲康唑） 治疗是否可以改变孢子肿胀的早期阶段，从而改变 BMDC 激活和过继转移时的体内过敏炎症。结果我们发现 AF 各向同性生长对于触发 BMDC 激活和介导过敏性气道炎症至关重要。此外，使用时间停止 A.f. 阶段，我们发现在生长培养基中至少 3 小时使孢子能够充分膨胀以激活 BMDC 以在体内引发过敏性气道炎症。发芽的 A.f. 与伊曲康唑一起孵育减少了孢子肿胀，并部分降低了它们激活 BMDC 以引发体内过敏性气道炎症的能力。结论 总之，我们的结果确定了 A.f. 发育的确切阶段，此时发芽的孢子能够激活 DC 以介导下游过敏性气道炎症。此外，我们已经确定抗真菌疗法部分降低了 AF 孢子刺激过敏反应的潜力，突出了抗真菌治疗可能有助于防止真菌过敏发展的潜在机制。