Purpose <p>Type 1 diabetes mellitus (T1DM) results from autoimmune-mediated destruction of pancreatic β-cells, leading to absolute insulin deficiency. Current treatments rely on insulin replacement and do not prevent β-cell loss. 4-Methylumbelliferone (4-MU), an inhibitor of hyaluronan synthesis, has shown anti-inflammatory and cytoprotective effects, but its therapeutic potential and mechanisms in T1DM remain unclear.</p> Methods <p>A streptozotocin (STZ)-induced mouse model of T1DM was treated with 4-MU for three weeks. Blood glucose levels and glucose tolerance were evaluated. Pancreatic islet morphology and cell composition were assessed by immunofluorescence. In parallel, STZ -injured MIN6 and βTC6 β-cells were used to investigate the effects of 4-MU on cell viability, oxidative stress, intracellular Ca²⁺ homeostasis, and glucose-stimulated insulin secretion. Network pharmacology, molecular docking, qPCR, and Western blot analyses were conducted to explore the underlying mechanisms.</p> Results <p>4-MU significantly reduced hyperglycemia and improved glucose tolerance in T1DM mice, accompanied by preservation of β-cell mass, normalization of the β/α-cell ratio, and reduced islet inflammation. In vitro, 4-MU protected β-cells from STZ-induced injury by decreasing reactive oxygen species (ROS) accumulation, restoring intracellular Ca²⁺ balance, and improving insulin secretion. Network pharmacology identified 48 shared targets between 4-MU and T1DM, with KEGG pathway enrichment highlighting the PI3K/Akt signaling pathway. Molecular docking revealed stable binding of 4-MU to key regulators, including EGFR, Akt, ESR1, INSR, and IGF1R. Consistently, 4-MU enhanced the phosphorylation of EGFR, PI3K, and Akt in injured β-cells.</p> Conclusion <p>4-MU exerts protective effects in T1DM by preserving pancreatic β-cells survival and function, potentially through activation of the EGFR/PI3K/Akt signaling pathway.</p>