iNKT Cells Exacerbate Sepsis-Associated Acute Lung Injury through IFN-?-Mediated Promotion of Macrophage Apoptosis

iNKT cells are a unique subset of T lymphocytes that play important and distinct roles in sepsis due to their specialized characteristics. Unlike conventional T cells, which are restricted by polymorphic major histocompatibility complex (MHC) molecules and recognize peptide antigens, iNKT cells are restricted by the non-polymorphic CD1d molecule and respond to lipid antigens. Upon activation, they rapidly produce large quantities of cytokines such as IFN-? and IL-4. While the immunoregulatory function of iNKT cells in sepsis has been well established, their dual capacity to exert both pro-inflammatory and anti-inflammatory effects remains incompletely understood. In particular, their specific role during the early stages of sepsis remains elusive. This study aims to elucidate the mechanism by which iNKT cells contribute to immune dysregulation in sepsis through the iNKT-macrophage axis. Overall design: We elucidated the mechanism by which iNKT cells exacerbate early inflammatory responses and contribute to immune dysregulation in sepsis-associated acute respiratory distress syndrome (ARDS), which involves the modulation of macrophage cell death. Using genetically engineered mouse models in combination with in vitro co-culture experiments (DN32.D3 cells with bone marrow-derived macrophages [BMDMs]), a co-culture system was established between BMDMs and DN32.D3 cells under standardized stimulation conditions. Specifically, DN32.D3 cells were seeded at a macrophage-to-iNKT cell ratio of 3:1 in 6-well plates, with 2 mL of RPMI-1640 complete medium per well. The modeling group was treated with LPS and a-Galactosylceramide. After 6 hours, suspended cells were collected and washed three times with ice-cold PBS.