Key Points PLY induces PMN migration across pulmonary epithelial cells. PLY-dependent PMN migration requires pore formation and epithelial 12-LOX activity. Diverse pore-forming toxins induce PMN migration. Streptococcus pneumoniae is a major cause of pneumonia, wherein infection of respiratory mucosa drives a robust influx of neutrophils. We have previously shown that S. pneumoniae infection of the respiratory epithelium induces the production of the 12-lipoxygenase (12-LOX)–dependent lipid inflammatory mediator hepoxilin A3, which promotes recruitment of neutrophils into the airways, tissue damage, and lethal septicemia. Pneumolysin (PLY), a member of the cholesterol-dependent cytolysin (CDC) family, is a major S. pneumoniae virulence factor that generates ∼25-nm diameter pores in eukaryotic membranes and promotes acute inflammation, tissue damage, and bacteremia. We show that a PLY-deficient S. pneumoniae mutant was impaired in triggering human neutrophil transepithelial migration in vitro. Ectopic production of PLY endowed the nonpathogenic Bacillus subtilis with the ability to trigger neutrophil recruitment across human-cultured monolayers. Purified PLY, several other CDC family members, and the α-toxin of Clostridium septicum, which generates pores with cross-sectional areas nearly 300 times smaller than CDCs, reproduced this robust neutrophil transmigration. PLY non–pore-forming point mutants that are trapped at various stages of pore assembly did not recruit neutrophils. PLY triggered neutrophil recruitment in a 12-LOX–dependent manner in vitro. Instillation of wild-type PLY but not inactive derivatives into the lungs of mice induced robust 12-LOX–dependent neutrophil migration into the airways, although residual inflammation induced by PLY in 12-LOX–deficient mice indicates that 12-LOX–independent pathways also contribute to PLY-triggered pulmonary inflammation. These data indicate that PLY is an important factor in promoting hepoxilin A3–dependent neutrophil recruitment across pulmonary epithelium in a pore-dependent fashion.

中文翻译：

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肺炎链球菌感染期间肺炎球菌溶血素诱导依赖 12-脂氧合酶的中性粒细胞迁移

关键点 PLY 诱导 PMN 跨肺上皮细胞迁移。PLY 依赖性 PMN 迁移需要孔形成和上皮 12-LOX 活性。不同的成孔毒素诱导 PMN 迁移。肺炎链球菌是肺炎的主要原因，其中呼吸道粘膜感染会导致中性粒细胞大量流入。我们之前已经表明，呼吸道上皮的肺炎链球菌感染会诱导 12-脂氧合酶 (12-LOX) 依赖性脂质炎症介质 hepoxilin A3 的产生，从而促进中性粒细胞募集到气道、组织损伤和致命性败血症。Pneumolysin (PLY) 是胆固醇依赖性溶细胞素 (CDC) 家族的成员，是一种主要的肺炎链球菌毒力因子，可在真核细胞膜中产生直径约 25 nm 的孔并促进急性炎症，组织损伤和菌血症。我们表明，缺乏 PLY 的肺炎链球菌突变体在体外触发人类中性粒细胞跨上皮迁移方面受损。PLY 的异位生产赋予了非致病性枯草芽孢杆菌在人类培养的单层中触发中性粒细胞募集的能力。纯化的 PLY、其他几个 CDC 家族成员以及 Clostridium septicum 的 α-毒素（其产生的孔的横截面积比 CDC 小近 300 倍）再现了这种强大的中性粒细胞迁移。在孔组装的各个阶段被困的 PLY 非孔形成点突变体不会招募中性粒细胞。PLY 在体外以 12-LOX 依赖性方式触发中性粒细胞募集。尽管 PLY 在 12-LOX 缺陷小鼠中引起的残留炎症表明 12-LOX 非依赖性通路也存在，但将野生型 PLY 而非非活性衍生物滴注到小鼠肺中会诱导强烈的 12-LOX 依赖性中性粒细胞迁移到气道中导致 PLY 引发的肺部炎症。这些数据表明，PLY 是促进以孔依赖性方式跨肺上皮募集依赖于 hepoxilin A3 的中性粒细胞的重要因素。