Department of Biology, Molecular Biology Section, Faculty of Science, Hacettepe University, Ankara, Turkey.
Department of Physics Engineering, Faculty of Engineering, Ankara University, Ankara, Turkey.
Exp Lung Res. 2021 Nov;47(9):436-450. doi: 10.1080/01902148.2021.1998734. Epub 2021 Nov 5.
: Many allergens have protease activities. Although the immunomodulatory effects of these antigens are well known, the effects attributed to their protease activities are not thoroughly investigated. We set out to determine the effects of house dust mite (HDM) allergens with varying protease activities on bronchial epithelial cell functions. BEAS-2B cells were maintained in ALI-culture and stimulated with Der p1 (cysteine protease), Der p6 (serine protease), and Der p2 (non-protease) with and without specific protease inhibitors or heat denaturation. Cell viability and epithelial permeability were measured with MTT and paracellular flux assay, respectively. The effect of heat denaturation on allergen structure was examined using models. Matrix metalloproteinases (MMPs) were investigated at the transcription (qPCR), protein (ELISA), and functional (zymography) levels. Epithelial permeability increased only after Der p6 but not after Der p1 or Der p2 stimulation. Der p2 increased both MMP-2 and MMP-9 expression, while Der p1 increased only MMP-9 expression. The heat-denatured form of Der p1 unexpectedly increased MMP-9 gene expression, which, through the use of models, was attributed to its ability to change receptor connections by the formation of new electrostatic and hydrogen bonds. IL-8 and GM-CSF production were increased after Der p1 and Der p2 but decreased after Der p6 stimulation. IL-6 decreased after Der p1 but increased following stimulation with Der p6 and heat-denatured Der p2. Allergens in house dust mites are capable of inducing various changes in the epithelial cell functions by virtue of their protease activities.
尘螨过敏原具有蛋白酶活性。尽管这些抗原的免疫调节作用众所周知，但它们的蛋白酶活性所产生的影响尚未得到深入研究。我们旨在确定具有不同蛋白酶活性的屋尘螨过敏原对支气管上皮细胞功能的影响。BEAS-2B 细胞在 ALI 培养中维持，并分别用 Der p1（半胱氨酸蛋白酶）、Der p6（丝氨酸蛋白酶）和 Der p2（非蛋白酶）以及特异性蛋白酶抑制剂或热变性进行刺激。使用 MTT 和细胞旁通量测定法分别测量细胞活力和上皮通透性。使用模型检查热变性对过敏原结构的影响。在转录（qPCR）、蛋白（ELISA）和功能（酶谱）水平研究基质金属蛋白酶（MMPs）。上皮通透性仅在 Der p6 刺激后增加，但在 Der p1 或 Der p2 刺激后没有增加。Der p2 增加了 MMP-2 和 MMP-9 的表达，而 Der p1 仅增加了 MMP-9 的表达。令人意外的是，热变性的 Der p1 增加了 MMP-9 基因表达，通过使用模型，这归因于其通过形成新的静电和氢键改变受体连接的能力。Der p1 和 Der p2 刺激后 IL-8 和 GM-CSF 的产生增加，但 Der p6 刺激后减少。Der p1 刺激后 IL-6 减少，但随后 Der p6 和热变性的 Der p2 刺激后增加。
