Roles of Nrf2/HO-1 and ICAM-1 in the Protective Effect of Nano-Curcumin against Copper-Induced Lung Injury

Abstract

Copper (Cu) is an essential trace element for maintaining normal homeostasis in living organisms. Yet, an elevated level of Cu beyond homeostatic capacity may lead to oxidative damage of cellular components in several organs, including the lungs. This work investigated the effects of curcumin (Curc) and nano-curcumin (nCurc) against Cu-induced lung injury, accenting the roles of oxidative stress, inflammation, and the nuclear factor erythroid 2-related factor/heme oxygenase-1 Nrf2/HO-1 pathway. Rats were challenged with 100 mg/kg of copper sulfate (CuSO₄) while being treated with Curc or nCurc for 7 days. Cu-triggered lung oxidative stress detected as dysregulation of oxidative/antioxidant markers, a downregulation of Nrf-2/HO-1 signaling, and an increase in the inflammatory markers interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and intracellular adhesion molecule-1 (ICAM-1). Additionally, it decreased the expression of lung-specific proteins, surfactant protein-C (SP-C), and mucin-1 (MUC-1), induced apoptosis, and caused changes in lung histology. Curc and nCurc alleviated CuSO₄-induced lung injury by suppressing oxidative damage and inflammation and activating Nrf-2/HO-1. They also prevented apoptosis and restored the normal expression of SP-C and MUC-1. We concluded that nCurc exhibited superior efficacy compared with Curc in mitigating CuSO₄-induced lung injury. This was associated with reduced oxidative stress, inflammation, and apoptotic responses and increased Nrf2/HO-1 signaling and expression of SP-C and MUC-1.

Keywords: ICAM-1; Nrf2/HO-1 pathway; copper sulfate; curcumin; inflammation; lung toxicity; oxidative stress.

Figure 1

Curc and nCurc mitigate oxidative stress after CuSO₄-induced lung injury. Treatment with nCurc decreased (A) MDA and increased (B) SOD activity, (C) GSH, and (D) GPX2. Curc increased GSH and GPX2 after CuSO₄ overexposure. Data are depicted as mean ± SEM (n = 7). * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, and **** p ≤ 0.0001.

Figure 2

Curc and nCurc improve tissue appearance and reverse CuSO₄-induced lung injury. Representative images of H&E-stained lung sections of control rats (A,B) showing normal lung histology. CuSO₄-challenged rats (C,D) showing inter-alveolar septum (black arrow), intra-alveolar hemorrhage and congestion (yellow arrow), bronchus (red arrow), ruptured alveolus (green arrow). Cu-challenged rats treated with DFO (E,F) showing alveoli with thin inter-alveolar septum (black arrow), mild infiltration of lymphocytes (red arrow), bronchus (yellow arrow), type I and type II pneumocytes are also seen (green arrow). CuSO₄-challenged rats treated with Curc (G,H) and nCurc (I,J) showing alveoli with normal inter-alveolar septum (black arrow), type I and type II pneumocytes are also seen (red arrow). (200×: (A,C,E,G,I), scale bar = 200 µm, 400×: (B,D,F,H,J), scale bar = 100 µm).

Figure 3

Curc and nCurc modulate pulmonary Keap-1/Nrf-2/HO-1 signaling after CuSO₄-induced lung injury (A–D). Exposure to CuSO₄ caused an increase in Keap-1 and a decrease in Nrf-2 and HO-1 protein expression. Treatment with DFO, Curc, and nCurc improved the expression of these proteins. Data are depicted as mean ± SEM (n = 7). **** p ≤ 0.0001.

Figure 4

Curc and nCurc ameliorate pulmonary inflammation after CuSO₄-induced lung injury (A–C). Cu exposure caused an increase in inflammatory marker levels. Treatment with DFO, Curc, and nCurc ameliorated the inflammation. Data are depicted as mean ± SEM (n = 7). * p ≤ 0.05, ** p ≤ 0.01, **** p ≤ 0.0001.

Figure 5

Curc and nCurc restore the pulmonary expression of SP-C and MUC-1 after CuSO₄-induced lung injury (A,B). Data are depicted as mean ± S.E.M. (n = 7). * p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001, **** p ≤ 0.0001.

Figure 6

Curc and nCurc prevent apoptosis by regulating BAX and Bcl-2 gene expression after CuSO₄-induced lung injury (A–C). CuSO₄ exposure caused an increase in BAX and a reduction in Bcl2 mRNA levels. Treatment with DFO, Curc, and nCurc improved the levels of apoptotic markers. Data are depicted as mean ± SEM (n = 7). ** p ≤ 0.01, **** p ≤ 0.0001.