Clara cell secretory protein 16 protects against PM2.5-induced ferroptosis in mouse lung epithelial cells in a...

Background Clara cell secretory protein 16 (CC16) has been reported to exert anti-inflammatory and anti-oxidant effects. However, its underlying mechanism remains unclear. This study aimed to investigate the protective effect and mechanism of CC16 using an in vitro model of PM2.5-induced mouse pulmonary epithelial cells (TC-1), with a specific focus on its concentration-dependent effects.

TC-1 cells were exposed to PM2.5 to induce inflammatory injury and ferroptosis, followed by treatment with CC16 at different concentrations (0.25, 0.5, and 1.0 µg/mL). TC-1 cells were divided into six groups: control, CC16, PM2.5, PM2.5 + CC16 (0.25 μg/mL), PM2.5 + CC16 (0.5 μg/mL), and PM2.5 + CC16 (1.0 μg/mL) groups. Cell viability was assessed using the Cell Counting Kit-8 assay. Levels of inflammatory cytokines (interleukin (IL)-5, IL-6, IL-13, IL-17A, and IL-1β) in the supernatant were measured by enzyme-linked immunosorbent assay. Protein expression levels of Nrf2, ACSL4, SLC7A11, and GPX4 were detected using western blotting. Intracellular reactive oxygen species (ROS) were detected with a fluorescent probe, and mitochondrial membrane potential (MMP) was measured by JC-1 staining. Glutathione (GSH), malondialdehyde (MDA), and Fe²⁺ content were measured using relevant kits. Mitochondrial ferroptosis features were observed by transmission electron microscopy (TEM).

PM2.5 exposure significantly reduced cell viability, aggravated inflammation, induced ferroptosis, and disrupted the integrity of mouse pulmonary epithelial cells. CC16 treatment reversed these effects in a concentration-dependent manner.

CC16 effectively mitigates PM2.5-induced cellular injury in mouse pulmonary epithelial cells through inhibition of ferroptosis, with its protective effect showing a clear concentration dependence. These findings suggest CC16 as a novel strategy for PM2.5-induced respiratory disease.