Serotonin-induced DNA synthesis and proliferation are mediated by autocrine secretion of transforming growth factor-α in primary cultures of adult rat hepatocytes

• Published in: Proceedings for Annual Meeting of The Japanese Pharmacological Society Vol. WCP2018; p. PO2-6-3
• Main Authors: Naito, Kota, Kurihara, Kazuki, Moteki, Hajime, Kimura, Mitsutoshi, Ogihara, Masahiko
• Format: Journal Article
• Language: English
• Published: Japanese Pharmacological Society 2018
• Subjects: hepatocytes; serotonin; TGF-α
• ISSN: 2435-4953; 2435-4953
• DOI: 10.1254/jpssuppl.WCP2018.0_PO2-6-3

Previously, we demonstrated that serotonin (5-hydroxytryotamine; 5-HT)-induced hepatocyte DNA synthesis and proliferation are mediated through 5-HT2B receptor. Moreover, 5-HT2B receptor-mediated hepatocyte mitogenesis involves activation of the Gq/phospholipase C (PLC) pathway and the epidermal growth factor (EGF)/transforming growth factor (TGF)- α receptor tyrosine kinase (RTK)/phosphatidylinositol 3-kinase (PI3K)/extracellular signal-regulated kinase (ERK) 2/mammalian target of rapamycin (mTOR) pathway. However, how these two signaling pathways are associated with each other remains unknown. Thus we hypothesized that 5-HT2B receptor stimulated by 5-HT induces secretion of a putative primary mitogen (e.g., TGF-α, insulin-like growth factor (IGF)-I) via the Gq/PLC pathway in primary cultured hepatocytes, followed by induction of DNA synthesis and proliferation that is mediated by the EGF/TGF-α RTK/PI3K/ERK2/mTOR pathway. 　Hepatocytes were isolated from normal livers by the two-step in situ collagenase perfusion to facilitate disaggregation of the adult rat liver. Isolated hepatocytes were plated onto collagen-coated plastic culture dishes and incubated for 3-h in Williams' medium E supplemented with 5% newborn calf serum, 0.1 nM dexamethasone in 5% CO2 in air at 37ºC to allow attachment. The medium was changed to Williams' medium E without serum and dexamethasone, and 5-HT with or without test substances was added.　We investigated effects of monoclonal antibody against TGF-α (1-100 ng/ml) or IGF-I (1-100 ng/ml) on 5-HT-induced hepatocytes DNA synthesis and proliferation. The TGF-α monoclonal antibody, but not IGF-I monoclonal antibody inhibited hepatocytes DNA synthesis and proliferation induced by 5-HT in a dose-dependent manner. Next, we tested to determine autocrine secretion of TGF-α from cultured hepatocytes. 5-HT (10-6 M) significantly increased TGF-α levels in the culture medium. The maximum TGF-α concentration (30 pg/ml) peaked 10 min after addition of 5-HT. The TGF-α secretion induced by 5-HT was completely blocked by a 5-HT2B receptor antagonist (LY272015). These results suggest that the proliferative mechanism of action of 5-HT is mediated by 5-HT2B receptor, and stimulation of 5-HT2B receptor increases autocrine TGF-α secretion from primary cultured hepatocytes.　In conclusion, our data indicate that 5-HT stimulates DNA synthesis and proliferation in primary cultures of adult rat hepatocytes by acting via autocrine secretion of TGF-α induced by the serotonin 5-HT2B receptor/Gq/PLC pathway.