The aim of the present study was to investigate whether the expression of tribbles pseudokinase 3 (TRB3) is involved in pulmonary interstitial fibrosis and to examine the possible mechanisms. The expression of TRB3 in murine alveolar type II epithelial cells (MLE‑12 cells) following transforming growth factor β1 (TGF‑β1) stimulation was assessed using various techniques, including western blot and reverse transcription‑quantitative polymerase chain reaction assays. TRB3 overexpression and downregulation models were used to evaluate the impact of TRB3 on the TGF‑β1‑induced epithelial‑mesenchymal transition (EMT) of MLE‑12 cells. The downregulation of TRB3 was induced by RNA interference. The expression of TRB3 was significantly increased in MLE‑12 cells following the activation of TGF‑β1 (P<0.05). The overexpression of TRB3 was found to promote activation of the TGF‑β1/Smad3 signaling pathway, EMT, and the upregulated expression of β‑catenin and EMT‑related genes and proteins (P<0.05), whereas the downregulation of TRB3 attenuated the promoting effect on EMT induced by TGF‑β1. In addition, the overexpression of TRB3 inhibited MLE‑12 cell proliferation by stimulating apoptosis, leading to the formation of pulmonary fibrosis (PF). The positive feedback loop demonstrated that TGF‑β1 induced the expression of TRB3, and TRB3, in turn, stimulated EMT and promoted the onset of PF through activation of the TGF‑β1/Smad3 signaling pathway. Therefore, TRB3 may promote the formation of PF through the TGF‑β1/Smad3 signaling pathway.