Keloid is a fibroproliferative skin disorder characterized by fibroblast hyperproliferation and excessive extracellular matrix (ECM) deposition. Astragalin (AST) is a bioactive natural flavonoid with multiple pharmacological properties. This study aims to investigate the effect of AST on keloid formation in vitro. Primary keloid fibroblasts (KFs) and normal fibroblasts (NFs) were isolated from human keloid tissues and normal skin tissues, respectively, and treated with or without AST. MTT, colony formation, and Transwell assays were utilized to evaluate AST's effect on fibroblast proliferation, migration, and invasiveness. Western blotting was implemented for detecting protein levels of ECM components and mitogen-activated protein kinases (MAPKs). The results showed that AST treatment hindered the proliferative, migratory, and invasive capacities of KFs and NFs, and KFs were more sensitive to AST than NFs. AST restrained ECM deposition and inactivated the MAPK signaling pathway in KFs and NFs. In conclusion, AST suppresses the invasive growth of keloid fibroblasts probably by inactivating MAPK signaling.