Downregulation of NUDCD1 Alleviates Development of Pulmonary Fibrosis via Upregulation of GPx4 and FSP1 Expression and Inhibition of Their DNA Methylation

Abstract

Background: NudC domain containing 1 (NUDCD1) plays a significant regulatory role in pulmonary fibrosis (PF). Similarly, the expression patterns of Glutathione Peroxidase 4 (GPx4) and Fibroblast-specific protein 1 (FSP1) are closely associated with the fibrotic process. However, it is not clear whether the regulatory mechanism of NUDCD1 in PF is related to GPx4 and FSP1. Therefore, this study aimed to investigate the role of NUDCD1 in PF and delve into the underlying mechanism of this disease progression. Methods: Initially, expression levels of NUDCD1 were assessed in PF using Quantitative Reverse Transcription Polymerase Chain Reaction (qRT-PCR), Western blot analysis, and Hematoxylin and Eosin (H&E) techniques. Furthermore, a silica (SiO₂)-induced pulmonary fibrosis cell model was developed. The cell models were subjected to various treatments, including Ferrostatin-1 (Fer-1) treatment, DNA methyltransferase inhibitor treatment, and transfection with siRNA and overexpression plasmid. Moreover, the effect of NUDCD1 knockdown was evaluated on iron death, apoptosis rates, and oxidative stress levels. Furthermore, the expression levels of NUDCD1, GPx4, and FSP1 were assessed in Type I alveolar epithelial (AECI) cells. Additionally, a bleomycin-induced PF mouse model was employed to assess the therapeutic effects of siRNA NUDCD1 transfection complex using various techniques, including qRT-PCR, Western blot analysis, H&E and Masson staining, and Enzyme-Linked Immunosorbent Assay (ELISA). Results: We observed that NUDCD1 was upregulated in PF. PF tissues exhibited severe collagen fiber deposition, significant cellular structural damage, and notable lipid oxidative damage (p < 0.01). Furthermore, Fer-1 and 5-Azacytidine (5-Aza) treatment significantly alleviated oxidative damage in PF and upregulated the expression of GPx4 and FSP1 (p < 0.05 and p < 0.01). Additionally, NUDCD1 knockdown significantly increased the expression of GPx4 and FSP1 (p < 0.01), while reducing the iron and Malondialdehyde (MDA) levels in the PF cell model (p < 0.05 and p < 0.01). Moreover, NUDCD1 knockdown reduced oxidative damage, mitigated cellular structural damage, and inhibited fibrosis in the PF animal model (p < 0.05 and p < 0.01). Additionally, reduced NUDCD1 level downregulated Alpha Smooth Muscle Actin (α-SMA) and Collagen I expression (p < 0.01), and upregulated GPx4 and FSP1 expression (p < 0.01). Conclusion: NUDCD1 was upregulated in PF. NUDCD1 mediated iron-dependent cell death by suppressing the expression of GPx4 and FSP1 genes through epigenetic mechanisms, thereby promoting the progression of PF.