Busold Stefanie, Akkerdaas Jaap H, Zijlstra-Willems Esther M, van der Graaf Kees, Tas Sander W, de Jong Esther C, van Ree Ronald, Geijtenbeek Teunis B H
Amsterdam University Medical Centers, location AMC, Department of Experimental Immunology, Amsterdam, Netherlands.
Amsterdam Institute for Infection and Immunity, Inflammatory Diseases, Amsterdam, Netherlands.
Front Med (Lausanne). 2023 Aug 8;10:1105538. doi: 10.3389/fmed.2023.1105538. eCollection 2023.
House dust mite (HDM) is a major cause of respiratory allergic diseases. Dendritic cells (DCs) play a central role in orchestrating adaptive allergic immune responses. However, it remains unclear how DCs become activated by HDM. Biochemical functions of the major HDM allergens Der p 1 (cysteine protease) and Der p 2 (MD2-mimick) have been implicated to contribute to DC activation.
We investigated the immune activating potential of HDM extract and its major allergens Der p 1 and Der p 2 using monocyte-derived DCs (moDCs). Maturation and activation markers were monitored by flow cytometry and cytokine production by ELISA. Allergen depletion and proteinase K digestion were used to investigate the involvement of proteins, and in particular of the major allergens. Inhibitors of spleen tyrosine kinase (Syk), Toll-like receptor 4 (TLR4) and of C-type lectin receptors (CLRs) were used to identify the involved receptors. The contribution of endotoxins in moDC activation was assessed by their removal from HDM extract.
HDM extract induced DC maturation and cytokine responses in contrast to the natural purified major allergens Der p 1 and Der p 2. Proteinase K digestion and removal of Der p 1 or Der p 2 did not alter the immune stimulatory capacity of HDM extract. Antibodies against the CLRs Dectin-1, Dectin-2, and DC-SIGN did not affect cytokine responses. In contrast, Syk inhibition partially reduced IL-6, IL-12 and completely blocked IL-10. Blocking TLR4 signaling reduced the HDM-induced IL-10 and IL-12p70 induction, but not IL-6, while endotoxin removal potently abolished the induced cytokine response.
Our data strongly suggest that HDM-induced DC activation is neither dependent on Der p 1 nor Der p 2, but depend on Syk and TLR4 activation, which might suggest a crosstalk between Syk and TLR4 pathways. Our data highlight that endotoxins play a potent role in immune responses targeting HDM.
屋尘螨（HDM）是呼吸道过敏性疾病的主要病因。树突状细胞（DCs）在协调适应性过敏性免疫反应中起核心作用。然而，目前尚不清楚DCs如何被HDM激活。主要的HDM变应原Der p 1（半胱氨酸蛋白酶）和Der p 2（MD2模拟物）的生化功能被认为与DC激活有关。
我们使用单核细胞衍生的DCs（moDCs）研究了HDM提取物及其主要变应原Der p 1和Der p 2的免疫激活潜力。通过流式细胞术监测成熟和激活标志物，通过ELISA检测细胞因子产生。变应原去除和蛋白酶K消化用于研究蛋白质，特别是主要变应原的参与情况。使用脾酪氨酸激酶（Syk）、Toll样受体4（TLR4）和C型凝集素受体（CLRs）的抑制剂来鉴定相关受体。通过从HDM提取物中去除内毒素来评估内毒素在moDC激活中的作用。
与天然纯化的主要变应原Der p 1和Der p 2相比，HDM提取物诱导DC成熟和细胞因子反应。蛋白酶K消化以及去除Der p 1或Der p 2并未改变HDM提取物的免疫刺激能力。针对CLRs Dectin-1、Dectin-2和DC-SIGN的抗体不影响细胞因子反应。相比之下，抑制Syk可部分降低IL-6、IL-12，并完全阻断IL-10。阻断TLR-4信号传导可降低HDM诱导的IL-10和IL-12p70诱导，但不影响IL-6，而去除内毒素可有效消除诱导的细胞因子反应。
我们的数据强烈表明，HDM诱导的DC激活既不依赖于Der p 1也不依赖于Der p 2，而是依赖于Syk和TLR4激活，这可能表明Syk和TLR4途径之间存在串扰。我们的数据突出表明，内毒素在针对HDM的免疫反应中起重要作用。
