Abstract Rationale Candida albicans, commonly isolated from the lower respiratory tract of critically ill patients, has been linked with prolonged mechanical ventilation and adverse clinical outcomes. C. albicans causes lung epithelial injury and barrier disruption, yet the underlying mechanisms are poorly understood. In oral epithelial cells, the tyrosine kinase receptor, EphA2, has been shown to bind C. albicans and induce a cytotoxic response. Therefore, we hypothesize that C. albicans interacts with lung epithelial cells via EphA2, mediating injury caused by regulated cell death pathways. Methods The reference research strain of C. albicans, SC 5314, was grown in YPD broth in hyphal phase at 30 °C for 16 hours then centrifuged, washed, and diluted to reach the desired multiplicity of infection (MOI). MLE12 murine lung epithelial cells (MLEC) were pre-incubated with ferroptosis inhibitor Ferrostatin-1 (Fer1) at 0.1µM, the EphA2 inhibitor ALW at 1µM, or a combination of the two. Separately, MLEC were pre-incubated for one hour with either Z-VAD-FMK, an apoptosis inhibitor, or Necrostatin-1s, a necroptosis inhibitor, at concentrations of 1µM, 10µM, and 50µM. After pre-incubation, MLEC were then co-cultured with C. albicans at 37 °C for 16 hours. Cell death was measured using a LDH detection assay. Fold change in cytotoxicity was calculated relative to C. albicans control group that was pre-incubated with vehicle. In a separate experiment, MLEC were treated with C. albicans or heat-killed C. albicans (HKCA) at MOI 1 and 10 with sequential fluorescence measurements to assess cell membrane permeability every 30 minutes for 6 hours. Results Live C. albicans induced regulated cell death in MLEC as seen by elevated LDH levels and a dose-dependent increase in cell membrane disruption over time. When compared to controls, there was no significant change in cell death when inhibiting apoptosis, necroptosis, or ferroptosis. However, when cells were pre-incubated with ALW, an inhibitor of the tyrosine kinase receptor EphA2, there was a significant reduction in median cell death by 23.4% (p 0.001). Conclusions These in vitro findings suggest that C. albicans can induce regulated cell death pathways in murine lung epithelial cells. Interestingly, EphA2 inhibition leads to a decrease in murine lung epithelial cell death whereas inhibitors of apoptosis, necroptosis, and ferroptosis alone do not significantly limit cell death during co-culture with C. albicans. Future research will investigate how inhibition of the EphA2 receptor tyrosine kinase interrupts lung epithelial cell death. This abstract is funded by: None
Coulter et al. (2026) studied this question.