Abstract: Objective To evaluate the role of serotonin (5-HT) in the crosstalk between cholangiocytes (biliary epithelia cells, BECs) and portal fibroblasts (PFs). Methods Cells were cultured and divided into 6 groups. 1) BECs were cultured alone; 2) BECs + TGF-β₁: BECs were cultured alone and then were treated with 2 ng/ml of recombinant transforming growth factor β₁(TGF-β₁) for 24 h; 3) BECs + 5-HT: BECs were cultured alone and then were treated with 60 ng/ml of 5-HT for 48 h; 4) PFs were cultured alone; 5) PFs + 5-HT: PFs were cultured alone and then were treated with 60 ng/ml of 5-HT for 48 h; 6) BECs + PFs: BECs and PFs were co-cultured. After culturing for 72 h, the concentration of 5-HT or TGF-β₁in culture medium was detected by ELISA. Expressions of TPH1, TPH2, 5-HTR1A, 5-HTR1B mRNA in BECs were detected by real-time QRT-PCR. BrdU and α-SMA as the marker of BECs proliferation and myofibroblastic transdifferentiation of PFs were evaluated by immunocytochemistry. Results Mono-cultured BECs expressed TPH1, TPH2, 5-HTR1A, 5-HTR1B mRNA with hypersecretion of 5-HT and slight proliferation of BECs. After being treated by TGF-β₁or co-cultured with PFs, BECs reduced the expression of TPH1 by 80% and TPH2 by 87%. Moreover, 5-HTR1A and HTR1B mRNA expression in BECs also dropped by 75% and 85% respectively, companied by the increasment of BECs proliferation. Mono-cultured PFs produced TGF-β₁with some MFs showing positive staining. However, after being treated by 5-HT or co-cultured with BECs, TGF-β₁secretion and α-SMA-positive cells were significantly increased. Conclusion 5-HT derived from BECs and TGF-β₁produced by PFs can mediate the autocrine and paracrine effects between BECs and PFs, and maintain the proliferation of BECs and the conversion of PFs to MFs, which may be meaningful to the pathogenesis of cholangiopathies.