Aloxistatin attenuates TNF-α-induced inflammatory responses in human bronchial epithelial cells: Implications for ARDS therapy

<bold>Introduction:</bold> Acute respiratory distress syndrome (ARDS) is a severe pulmonary condition that is influenced by elevated TNF-α. TNF-α triggers the activation of cathepsins, enzymes known for worsening lung damage by degrading the extracellular matrix. Aloxistatin, a targeted cysteine protease inhibitor, shows promise in mitigating TNF-α and cathepsin activity, offering hope for ARDS treatment. This study investigates the impact of Aloxistatin on TNF-α-induced proinflammatory cytokines and matrix metalloproteinases (MMPs) in human bronchial epithelial cells (A549), shedding light on its therapeutic potential in ARDS. <bold>Methods:</bold> A549 cells were pretreated with Aloxistatin for 1 hour prior to stimulation with TNF-α. Cell samples and culture medium were collected at various time intervals, and gene and protein levels of IL-6, IL-8, MMP1, and MMP2 were quantified using quantitative PCR and ELISA. The impact of Aloxistatin on cell viability was assessed using the CCK8 assay. <bold>Results:</bold> Aloxistatin (1-1000μM) was not toxic in A549 cells. Following TNF-α stimulation, a significant increase in gene and protein expression of IL-6, IL-8, MMP1 and MMP2 was observed, indicating an inflammatory response. Pretreatment with Aloxistatin significantly reduced TNF-α-induced upregulation of IL-6, IL-8, MMP1 and MMP2 in a dose-dependent manner, both in gene and protein expression at 12 and 24 hours. <bold>Conclusions:</bold> Aloxistatin effectively dampens the TNF-α-induced inflammatory response in bronchial epithelial cells by attenuating key proinflammatory cytokines and MMPs. Therefore, Aloxistatin may serve as a promising therapeutic candidate for ARDS.