Hassim Z, Maronese S E, Kumar R K
Inflammation Research Unit, School of Pathology, University of New South Wales, Sydney, Australia.
Thorax. 1998 May;53(5):368-71. doi: 10.1136/thx.53.5.368.
Pollens are important triggers for asthma but the mechanism of sensitisation to their proteins remains poorly understood. The intrinsic protease activity of some allergens may contribute to sensitisation by disrupting the integrity of the airway epithelial barrier. Pollens release a variety of enzymes, including proteases, upon hydration. The hypothesis that such enzymes might be able to damage airway epithelial cells was therefore tested.
Diffusates from pollens of Lolium perenne (ryegrass), Poa pratensis (Kentucky bluegrass), Acacia longifolia (Sydney golden wattle), or Casuarina distyla (she-oak) were incubated with mouse tracheal epithelial cells in culture and cellular detachment was quantified using a methylene blue dye binding assay.
Diffusates prepared using 100 mg/ml of pollen caused detachment of 30-90% of airway epithelial cells in separate experiments. Within each experiment comparable detachment was observed with all diffusates tested, although total protein in the diffusates varied markedly between species. Viability of the cells recovered after exposure to Acacia diffusate was higher than after detachment by exposure to Lolium diffusate. Cellular detachment by all of the diffusates could be almost completely inhibited by addition of 10% serum. Aprotinin, an inhibitor of serine proteases, partially blocked activity in diffusates of Lolium pollen but not of Acacia pollen. In contrast, alpha 1-protease inhibitor and secretory leucocyte protease inhibitor (SLPI) were not able to block the activity of either diffusate at concentrations which inhibited cellular detachment by trypsin.
Proteases released by pollens are able to cause detachment of airway epithelial cells from their substratum in vitro and may not be effectively inhibited by endogenous antiproteases.
花粉是哮喘的重要触发因素，但其蛋白质致敏机制仍知之甚少。一些过敏原的内在蛋白酶活性可能通过破坏气道上皮屏障的完整性而导致致敏。花粉在水化后会释放多种酶，包括蛋白酶。因此，对这种酶可能损害气道上皮细胞的假说进行了测试。
将黑麦草、草地早熟禾、长叶相思树或异叶木麻黄的花粉浸出液与培养的小鼠气管上皮细胞一起孵育，并使用亚甲蓝染料结合试验对细胞脱离情况进行定量。
在单独的实验中，用100mg/ml花粉制备的浸出液导致30%-90%的气道上皮细胞脱离。在每个实验中，所有测试浸出液均观察到类似的细胞脱离情况，尽管不同物种浸出液中的总蛋白含量差异显著。暴露于相思树浸出液后细胞的活力恢复情况高于暴露于黑麦草浸出液导致细胞脱离后的恢复情况。添加10%血清可几乎完全抑制所有浸出液导致的细胞脱离。丝氨酸蛋白酶抑制剂抑肽酶部分阻断了黑麦草花粉浸出液的活性，但未阻断相思树花粉浸出液的活性。相比之下，α1-蛋白酶抑制剂和分泌型白细胞蛋白酶抑制剂（SLPI）在抑制胰蛋白酶导致细胞脱离的浓度下，均无法阻断任何一种浸出液的活性。
花粉释放的蛋白酶能够在体外导致气道上皮细胞从其基底脱离，且可能无法被内源性抗蛋白酶有效抑制。
