Tohoku J. Exp. Med., 2002, 198 (1)

Establishment and Characterization of a Simian Virus 40-Immortalized Rat
Pancreatic Stellate Cell Line

MASAHIRO SATOH, ATSUSHI MASAMUNE, YOSHITAKA SAKAI, KAZUHIRO KIKUTA,
HIROFUMI HAMADA¹ and TOORU SHIMOSEGAWA

Division of Gastroenterology, Tohoku University Graduate School of Medicine, Sendai 980-8574, and ¹Department of Molecular Medicine, Sapporo Medical University School of Medicine, Sapporo 060-8556

Activated pancreatic stellate cells (PSCs) have recently been implicated in the pathogenesis of pancreatic fibrosis and inflammation. Primary PSCs can be subcultured only several times because of their limited growth potential. A continuous cell line would be valuable in studying molecular mechanisms of these pancreatic disorders. The aim of this study was to establish an immortalized cell line of rat PSCs. PSCs were isolated from the pancreas of male Wistar rats, and the simian virus 40 T antigen was introduced to PSCs by retrovirus-mediated gene transfer. This procedure yielded an actively growing cell line, designated as SAM-K. This cell line has been passaged repeatedly for almost 2 years, and is thus likely immortalized. SAM-K cells retained morphological characteristics of primary PSCs, and expressed a-smooth muscle actin, glial fibrillary acidic protein, type I collagen, fibronectin, and prolyl hydroxylases. The level of p53 expression was very high in SAM-K cells. Proliferation of SAM-K cells was stimulated by serum and platelet-derived growth factor-BB. Interleukin-1b (IL-1b) activated nuclear factor-kB, activator protein-1, and three classes of mitogen-activated protein (MAP) kinases: extracellular signal-regulated kinase1/2, c-Jun N-terminal kinase, and p38 MAP kinase. IL-1b induced expression of intercellular adhesion molecule-1 and monocyte chemoattractant protein-1, both of which were abolished in the presence of pyrrolidine dithiocarbamate, a specific inhibitor of nuclear factor-kB activation. IL-1b-induced monocyte chemoattractant protein-1 was partially inhibited by specific inhibitors of MAP kinase kinase (U0126) and of p38 MAP kinase (SB203580) whereas intercellular adhesion molecule-1 expression was not altered by the inhibitors. Thus, SAM-K would be useful for in vitro studies of cell biology and signal transduction of PSCs.

Keywords —— pancreatic stellate cells; chronic pancreatitis; pancreatic fibrosis; SV40 large T antigen; signal transduction

© 2002 Tohoku University Medical Press

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Tohoku J. Exp. Med., 2002, 198, 55-69

Address for reprints: Atsushi Masamune, M.D., Division of Gastroenterology, Tohoku University Graduate School of Medicine, 1-1 Seiryocho, Aoba-ku, Sendai 980-8574, Japan.

e-mail: amasamune@int3.med.tohoku.ac.jp

Abbreviations: AP-1, activator protein-1; ERK, extracellular signal-regulated kinase; FBS, fetal bovine serum; ICAM-1, intercellular adhesion molecule-1; Ik B-a , inhibitor of nuclear factor k B; IL, interleukin; JNK, c-Jun N-terminal kinase; MAP kinases, mitogen-activated protein kinases; MCP-1, monocyte chemoattractant protein-1; NF-k B, nuclear factor-k B; PSCs, pancreatic stellate cells; PDGF-BB, platelet-derived growth factor-BB isoform; PDTC, pyrrolidine dithiocarbamate; a -SMA, a -smooth muscle actin; SV40, simian virus 40; TNF-a , tumor necrosis factor-a