Rationale: Respiratory syncytial virus (RSV) bronchiolitis causes significant infant mortality. Bronchiolitis is characterized by airway epithelial cell (AEC) death; however, the mode of death remains unknown.Objectives: To determine whether necroptosis contributes to RSV bronchiolitis pathogenesis via HMGB1 (high mobility group box 1) release.Methods: Nasopharyngeal samples were collected from children presenting to the hospital with acute respiratory infection. Primary human AECs and neonatal mice were inoculated with RSV and murine Pneumovirus, respectively. Necroptosis was determined via viability assays and immunohistochemistry for RIPK1 (receptor-interacting protein kinase-1), MLKL (mixed lineage kinase domain-like pseudokinase) protein, and caspase-3. Necroptosis was blocked using pharmacological inhibitors and RIPK1 kinase-dead knockin mice.Measurements and Main Results: HMGB1 levels were elevated in nasopharyngeal samples of children with acute RSV infection. RSV-induced epithelial cell death was associated with increased phosphorylated RIPK1 and phosphorylated MLKL but not active caspase-3 expression. Inhibition of RIPK1 or MLKL attenuated RSV-induced HMGB1 translocation and release, and lowered viral load. MLKL inhibition increased active caspase-3 expression in a caspase-8/9-dependent manner. In susceptible mice, Pneumovirus infection upregulated RIPK1 and MLKL expression in the airway epithelium at 8 to 10 days after infection, coinciding with AEC sloughing, HMGB1 release, and neutrophilic inflammation. Genetic or pharmacological inhibition of RIPK1 or MLKL attenuated these pathologies, lowered viral load, and prevented type 2 inflammation and airway remodeling. Necroptosis inhibition in early life ameliorated asthma progression induced by viral or allergen challenge in later life.Conclusions: Pneumovirus infection induces AEC necroptosis. Inhibition of necroptosis may be a viable strategy to limit the severity of viral bronchiolitis and break its nexus with asthma.

中文翻译：

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呼吸道合胞病毒感染可促进气管上皮细胞坏死和HMGB1释放。

理由：呼吸道合胞病毒（RSV）毛细支气管炎可导致大量婴儿死亡。毛细支气管炎的特征是气道上皮细胞（AEC）死亡。目的：确定坏死病是否通过HMGB1（高迁移率第1组）的释放而促成RSV细支气管炎的发病。方法：从就诊于急性呼吸道感染医院的儿童中收集鼻咽样本。将原代人AEC和新生小鼠分别接种RSV和鼠肺炎病毒。通过活力测定和免疫组化确定RIPK1（受体相互作用蛋白激酶1），MLKL（混合谱系激酶域样假激酶）蛋白和caspase-3的坏死病。使用药理抑制剂和RIPK1激酶致死敲除小鼠阻断了坏死性坏死。测量和主要结果：急性RSV感染儿童的鼻咽样品中HMGB1水平升高。RSV诱导的上皮细胞死亡与磷酸化的RIPK1和磷酸化的MLKL增加有关，但与caspase-3活性无关。RIPK1或MLKL的抑制作用减弱了RSV诱导的HMGB1的转运和释放，并降低了病毒载量。MLKL抑制以caspase-8 / 9依赖性方式增加了活性caspase-3的表达。在易感小鼠中，肺炎病毒感染在感染后8到10天会上调RIPK1和MLKL在气道上皮中的表达，这与AEC脱落，HMGB1释放和嗜中性粒细胞炎症相吻合。RIPK1或MLKL的遗传或药理抑制作用减弱了这些病理，降低病毒载量，并防止2型炎症和气道重塑。早期抑制坏死病改善了病毒或过敏原激发的哮喘病发作。结论：肺炎病毒感染可诱发AEC坏死。抑制坏死病可能是限制病毒性毛细支气管炎的严重性并打破其与哮喘的联系的可行策略。