Hypoosmotic stress activates p38, ERK 1 and 2, and SAPK/JNK in rat hepatocytes

Background, Following hepatocyte injury, changes in the perihepatocyte milieu modulate cell volume and influence growth. Hypoosmotic stress activates nuclear factor-kappa B (NF-kappa B), a transcription factor believed to prime cell cycle progression in hepatocytes, In this study, we investigate the role of mitogen-activated protein kinases (MAPKs) in the activation of NF-kappa B. Materials and methods. Quiescent primary hepatocytes were exposed to hypoosmotic serum-free William's E (WE) medium (200 mOsm/liter), with or without a l-h pretreatment with either PD 98059 (15 mu M) or SB 202190 (3 mu M). Parallel experiments were conducted using hepatocyte growth factor (HGF) at 0.1 mg/ml and normoosmotic WE medium as positive and negative controls, respectively (n = 3), Relative densitometries of Western blots measured phosphorylated cytoplasmic p38, ERK 1 and 2, and SAPK/JNK. Electromobility shift assays examined nuclear NF-kappa B activation. Results. (i) Hypoosmolar WE medium phosphorylated p38, ERK 1 and 2, and SAPK/JNK by 5 min. (ii) Hypoosmolar WE medium activated NF-kappa B at 60 min. (iii) HGF phosphorylated all three MAPKs and activated NF-kappa B with profiles similar to those of hypoosmotic stress, (iv) Both PD 98059 and SE 202190 abrogated the activation of NF-kappa B in HGF-stimulated cells but not in hypoosmotically stressed cells, Conclusion, (i) Both hypoosmotic cell swelling and HGF phosphorylate p38, ERK 1 and 2, and SAPK/JNK, and (ii) HGF, but not hypoosmotic stress, activates NF-kappa B via p38 and ERK 1 and 2 phosphorylation. These data suggest that cell swelling activates NF-kappa B through a pathway separate from that of growth factors, (C) 2000 Academic Press.