Benign prostatic hyperplasia (BPH) is a common urinary disease among the elderly, characterized by abnormal prostatic cell proliferation. Neferine is a dibenzyl isoquinoline alkaloid extracted from Nelumbo nucifera and has antioxidant, anti-inflammatory and anti-prostate cancer effects. The beneficial therapeutic effects and mechanism of action of neferine in BPH remain unclear. A mouse model of BPH was generated by subcutaneous injection of 7.5 mg/kg testosterone propionate (TP) and 2 or 5 mg/kg neferine was given orally for 14 or 28 days. Pathological and morphological characteristics were evaluated. Prostate weight, prostate index (prostate/body weight ratio), expression of type â ?5alpha-reductase, androgen receptor (AR) and prostate specific antigen were all decreased in prostate tissue of BPH mice after administration of neferine. Neferine also downregulated the expression of pro-caspase-3, uncleaved PARP, TGF-beta1, TGF-beta receptor â ?(TGFBR2), p-Smad2/3, N-cadherin and vimentin. Expression of E-cadherin, cleaved PARP and cleaved caspase-3 was increased by neferine treatment. 1-100 muM neferine with 1 muM testosterone or 10 nM TGF-beta1 were added to the culture medium of the normal human prostate stroma cell line, WPMY-1, for 24 h or 48 h. Neferine inhibited cell growth and production of reactive oxygen species (ROS) in testosterone-treated WPMY-1 cells and regulated the expression of androgen signaling pathway proteins and those related to epithelial-mesenchymal transition (EMT). Moreover, TGF-beta1, TGFBR2 and p-Smad2/3, N-cadherin and vimentin expression were increased but E-cadherin was decreased after 24 h TGF-beta1 treatment in WPMY-1 cells. Neferine reversed the effects of TGF-beta1 treatment in WPMY-1 cells. Neferine appeared to suppress prostate growth by regulating the EMT, AR and TGF-beta/Smad signaling pathways in the prostate and is suggested as a potential agent for BPH treatment. CI - (c) 2023 The Author(s).