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Background <p>Ventilator-induced lung injury (VILI) is characterized by acute lung damage and NF-κB–driven cytokine production during mechanical ventilation. However, the upstream signals that initiate and exacerbate NF-κB-driven inflammation remain poorly understood. Although endoplasmic reticulum (ER) stress is implicated in VILI, how ER stress pathways interact with innate immune sensors to promote inflammation is incompletely defined. Here, we investigate the roles of NOD1 and the ER stress–responsive PERK/eIF2α pathway in a VILI mice model. </p> Methods <p>C57BL/6 mice were subjected to high-tidal volume (HTV) ventilation for 4&#xa0;h to establish an in vivo model of VILI. To investigate the role of NOD1 in VILI, Nod1-knockout (Nod1-KO) mice were generated. To further determine the involvement of the PERK pathway, wild-type and Nod1-KO mice were treated with PERK inhibitor GSK2606414 prior to HTV ventilation. Lung injury was evaluated using H&amp;E staining, wet/dry ratio, and analysis of bronchoalveolar lavage fluid (BALF) for total protein concentration and inflammatory cell counts. Inflammatory cytokine levels were quantified by ELISA. Relative proteins expression was analyzed by immunoblotting and immunofluorescence.</p> Results <p>HTV ventilation induced lung histopathological damage, alveolar-capillary barrier dysfunction, and increased levels of IL-1β, IL-6, and TNF-α, along with upregulation of NOD1 and the ER stress marker GRP78. Genetic deletion of Nod1 attenuated lung injury and inflammatory cytokine release, associated with reduced GRP78 abundance and decreased NF-κB activation. In parallel, HTV increased phosphorylation of PERK and eIF2α, and pharmacological inhibition of PERK with GSK2606414 ameliorated lung injury and inflammation while suppressing NF-κB activation, without altering NOD1 protein abundance. Notably, combining NOD1 deficiency with PERK inhibition produced more pronounced protection against HTV-induced lung injury than either intervention alone, whereas the reduction of NF-κB readouts did not further deepen under the combined condition.</p> Conclusions <p>HTV-induced inflammatory lung injury is associated with activation of both NOD1 and PERK/eIF2α signaling. Mechanistically, these pathways contribute to NF-κB-associated inflammatory responses and lung injury in VILI. Notably, the greater protection observed under combined NOD1 deficiency and PERK inhibition, despite a plateau in NF-κB readouts, suggests that additional injury mechanisms beyond NF-κB may also be involved.</p>

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Mechanical ventilation activates PERK/eIF2α and NOD1 pathways that contribute to NF-κB–driven inflammation in VILI

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年6月24日

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  • Maoyao Ling,
  • Yalan Jiang,
  • Umut Cakir,
  • Kejian Lu,
  • Xiaoting Liao,
  • Guiting Yang,
  • Liu Ji,
  • Tingting Meng,
  • Cuiyuan Huang,
  • Liu Ye,
  • Linghui Pan

摘要

Background

Ventilator-induced lung injury (VILI) is characterized by acute lung damage and NF-κB–driven cytokine production during mechanical ventilation. However, the upstream signals that initiate and exacerbate NF-κB-driven inflammation remain poorly understood. Although endoplasmic reticulum (ER) stress is implicated in VILI, how ER stress pathways interact with innate immune sensors to promote inflammation is incompletely defined. Here, we investigate the roles of NOD1 and the ER stress–responsive PERK/eIF2α pathway in a VILI mice model.

Methods

C57BL/6 mice were subjected to high-tidal volume (HTV) ventilation for 4 h to establish an in vivo model of VILI. To investigate the role of NOD1 in VILI, Nod1-knockout (Nod1-KO) mice were generated. To further determine the involvement of the PERK pathway, wild-type and Nod1-KO mice were treated with PERK inhibitor GSK2606414 prior to HTV ventilation. Lung injury was evaluated using H&E staining, wet/dry ratio, and analysis of bronchoalveolar lavage fluid (BALF) for total protein concentration and inflammatory cell counts. Inflammatory cytokine levels were quantified by ELISA. Relative proteins expression was analyzed by immunoblotting and immunofluorescence.

Results

HTV ventilation induced lung histopathological damage, alveolar-capillary barrier dysfunction, and increased levels of IL-1β, IL-6, and TNF-α, along with upregulation of NOD1 and the ER stress marker GRP78. Genetic deletion of Nod1 attenuated lung injury and inflammatory cytokine release, associated with reduced GRP78 abundance and decreased NF-κB activation. In parallel, HTV increased phosphorylation of PERK and eIF2α, and pharmacological inhibition of PERK with GSK2606414 ameliorated lung injury and inflammation while suppressing NF-κB activation, without altering NOD1 protein abundance. Notably, combining NOD1 deficiency with PERK inhibition produced more pronounced protection against HTV-induced lung injury than either intervention alone, whereas the reduction of NF-κB readouts did not further deepen under the combined condition.

Conclusions

HTV-induced inflammatory lung injury is associated with activation of both NOD1 and PERK/eIF2α signaling. Mechanistically, these pathways contribute to NF-κB-associated inflammatory responses and lung injury in VILI. Notably, the greater protection observed under combined NOD1 deficiency and PERK inhibition, despite a plateau in NF-κB readouts, suggests that additional injury mechanisms beyond NF-κB may also be involved.