Effect of neodymium oxide exposure on inflammatory factors and transforming growth factor-β/Smads pathway in mouse lung tissues

Background  Rare earth neodymium oxide (Nd₂O₃) can cause cell fibrous nodules in rat lung tissues, but the mechanism is not clear. The transforming growth factor-β (TGF-β)/Smads signaling pathway is a classic fibrosis pathway, but its role in lung injury caused by Nd₂O₃ is rarely studied.

Objective  This experiments explores the effects of Nd₂O₃ on lung tissue inflammatory factors, fibrosis factors, and cell signaling transduction molecules Smad2 and Smad3, and preliminary elucidates the mechanism of Nd₂O₃ induced lung injury in mice.

Methods  A total of 144 SPF-grade healthy adult male mice were randomly divided into one control group and three Nd₂O₃ exposed groups, with 36 mice in each group. Non-exposed tracheal injection was used to establish mouse lung injury models. The three exposed groups were perfused with 0.1 mL Nd₂O₃ suspension at 62.5, 125, and 250 mg·mL^-1, respectively. The control group was given equal volumes of normal saline. Twelve mice in each group were sacrificed 7, 14, and 28 days after exposure respectively. Lung tissue morphology was observed with hematoxylin-eosin staining (HE staining); hydroxyproline (HYP) was determined by alkaline hydrolysis method; tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), transforming growth factor-β1 (TGF-β1), and connective tissue growth factor (CTGF) in mouse lung tissues were detected by enzyme-linked immunosorbent assay; the relative expression levels of Smad2 and Smad3 mRNA in the lung tissues of mice were evaluated by real-time quantitative PCR; the relative expressions of Smad2, Smad3, p-Smad2, and p-Smad3 proteins in the lung tissues of the 250 mg·mL^-1 group were detected by Western blotting.

Results  The results of HE staining showed that after the mice were exposed to Nd₂O₃, compared with the control group, the exposed groups showed inflammatory exudation on the 7th day, fibrogenesis on the 14th day, and cell fibrous nodules on the 28th day. The HYP levels in the lung tissues of mice on the 7th, 14th, and 28th days after exposure were higher than those in the control group, and the 250 mg·mL^-1 group on the 28th day reached the highest level, (1.87±0.19) μg·mg^-1. Compared with the control group, the levels of IL-6, TNF-α, TGF-β1, and CTGF in the lung tissues in the exposed groups were higher than those in the control group on the 7th, 14th, and 28th days (P < 0.05), except the TGF-β1 level on the 14th day; the levels of IL-6 and TNF-α rose to the highest on the 14th day, and the levels in the 250 mg·mL^-1 Nd₂O₃ group were (5 502.96±76.53) and (2 484.58±71.82) pg·mg^-1, respectively; the levels of TGF-β1 and CTGF reached the highest on the 28th day, and the levels in the 250 mg·mL^-1 Nd₂O₃ group were (0.71±0.08) and (6.17±0.19) ng·mg^-1, respectively. The relative expression levels of Smad2 and Smad3 mRNA in the lung tissues of mice at each time point after exposure were higher than those in the control group (P < 0.05), except Smad3 mRNA on the 7th day; the mRNA levels of Smad2 and Smad3 reached the highest on the 28th day, and the levels in the 250 mg·mL^-1 Nd₂O₃ group were 10.04±0.61 and 11.87±0.88 respectively. The relative protein expression levels of Smad2, Smad3, p-Smad2, and p-Smad3 in the 250 mg·mL^-1 Nd₂O₃ group were higher than those in the control group (P < 0.05); the Smad2 level reached a maximum of 1.16±0.16 on the 28th day, and the levels of Smad3, p-Smad2, and p-Smad3 reached a maximum of 2.05±0.10, 1.37±0.05, and 2.63±0.09 respectively on the 14th day.

Conclusion  Nd₂O₃ exposure can cause inflammation and fibrosis in the lung tissues of mice, and the mechanism may be related to the activation of the TGF-β/Smads pathway.