Renal mesangial cell proliferation is a major clinical feature of diabetic nephropathy (DN) and includes glomerulosclerosis and renal fibrosis. Samchuleum (SCE) is a traditional herbal mixture that is recorded in the ancient Korean medical book, Donguibogam. The present study attempted to determine whether SCE treatment was able to improve high glucose (HG)‑induced mesangial cell fibrosis and glomerulosclerosis in primary cultured human mesangial cells. Thymidine incorporation under HG induction was increased, but was decreased by SCE in a dose dependent manner. Pretreatment with SCE led to a downregulation in the expression of cyclins and cyclin‑dependent kinases (CDKs), and an upregulation of the CDK inhibitors, p21waf1/cip1 and p27kip1. In addition, SCE exposure markedly suppressed the reactive oxygen species signaling pathway under HG conditions. HG enhanced the expression levels of fibrosis‑promoting mediators, including collagen IV and connective tissue growth factor, which were markedly attenuated by SCE. SCE treatment inhibited HG‑induced fibronectin mRNA expression and decreased the expression of transforming growth factor (TGF)‑β1, Smad‑2 and Smad‑4, whereas Smad‑7 expression increased under HG. SCE treatment induced the degradation of the extracellular matrix by blocking TGF‑β1/Smad signaling. Therefore, the present study suggested that the inhibitory effect of SCE on mesangial proliferation and renal fibrosis may be an effective therapy in the treatment of renal dysfunction leading to DN.