Background <p>Aflatoxin B₁ (AFB₁) is a potent hepatotoxic mycotoxin and a major environmental risk factor for hepatocellular carcinoma (HCC). Hepatic fibrosis is a critical intermediate stage in this process, and METTL3-mediated m⁶A modification may represent an important post-transcriptional mechanism linking AFB₁-induced liver injury to fibrogenic progression.</p> Methods <p>AFB₁-induced hepatic fibrosis was evaluated using in vivo mouse models and in vitro cultured hepatic stellate cells (HSC). Global m⁶A methylation and methyltransferase-like 3 (METTL3) expression were assessed by liquid chromatography-mass spectrometry, Western blotting, single-nucleus RNA sequencing, and quantitative real-time PCR. METTL3 was inhibited using small interfering RNA or the selective inhibitor STM2457. Molecular docking was performed to identify potential METTL3-binding compounds, followed by functional validation.</p> Results <p>AFB₁ exposure promoted hepatic fibrosis and HSC activation, accompanied by global m⁶A hypermethylation and upregulation of METTL3. METTL3 increased the m⁶A modification of collagen-related transcripts, enhancing their stability and promoting extracellular matrix production in a <i>YTHDF1</i>-dependent manner. Inhibition of METTL3 suppressed HSC activation and fibrotic gene expression both in vitro and in vivo. Molecular docking identified saxagliptin as a potential METTL3-binding compound, which reduced AFB₁-induced HSC activation and extracellular matrix accumulation, consistent with the effects of STM2457.</p> Conclusions <p>These findings indicate that METTL3 functions as a post-transcriptional regulator in AFB₁-induced liver fibrosis via m⁶A modification. METTL3 inhibition, achieved via genetic knockdown or selective inhibitors, affects HSC activation and fibrotic gene expression, supporting its role as a therapeutic target in AFB₁-induced liver fibrosis.</p>