Mucin 3A Hypomethylation Facilitated Gastric Cancer Cell Growth and Progression via Regulating Programmed Cell Death-Ligand 1 and AKT Phosphorylation

Abstract

Background: Methylation-induced alterations of mucin 3A (MUC3A) expression have been observed in various malignancies, yet few studies have investigated its role in gastric cancer (GC). Herein, we elucidate the role of MUC3A in GC progression and its associated mechanisms. Methods: GC cells were treated with 5-Aza-2′-Deoxycytidine (5-Aza) and the phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway inhibitor LY294002, or transfected with short hairpin RNA targeting MUC3A (shRNA-MUC3A), MUC3A overexpression, and DNA methyltransferase 2 (DNMT2) overexpression constructs. Quantification of MUC3A was conducted using quantitative reverse transcription-PCR (qRT-PCR), while the methylation level of MUC3A was evaluated through methylation-specific PCR. Cell proliferation, migration, invasion, and apoptosis were assessed using 5-Ethynyl-2′-Deoxyuridine (EdU) incorporation assay, wound healing assay, Transwell assay, and flow cytometry, respectively. Additionally, protein expressions of programmed cell death-ligand 1 (PD-L1), phospho-AKT (p-AKT), and total AKT were determined through Western blot analysis. Results: Upregulation of MUC3A was observed in GC tissues (p < 0.001), while methylation of MUC3A was nearly absent in tumor tissues. Treatment with 5-Aza promoted proliferation, migration, invasion, and PD-L1 expression while inhibiting apoptosis in GC cells (p < 0.05). Conversely, shRNA-MUC3A demonstrated opposite effects (p < 0.05). Moreover, the effects induced by 5-Aza on GC cells could be reversed by MUC3A knockdown (p < 0.05). Additionally, overexpression of MUC3A enhanced the malignant behaviors of GC cells (p < 0.01), but this effect was reversed by pre-treatment with LY294002, a PI3K/AKT pathway inhibitor (p < 0.05). Furthermore, DNMT2 was found to bind to the MUC3A CpG island site, and overexpression of DNMT2 weakened the impact of MUC3A overexpression on accelerating the progression of GC (p < 0.05). Conclusion: Hypomethylation of MUC3A can promote the malignant behaviors of GC cells through the regulation of PD-L1 and the PI3K/AKT pathway, suggesting its potential as a biomarker in gastric carcinogenesis.