Biliary tract malignancies represent challenges because of the lack of effective therapy and poor prognosis, in part because of the paucity of information regarding the mechanisms regulating their growth. We have recently identified a critical role for the p44/p42 mitogen-activated protein kinase (MAPK) pathway in interleukin 6 (IL-6)-stimulated growth of human cholangiocytes. Although IL-6 is a potential mitogen for cholangiocarcinoma, the role of this cytokine and its intracellular signaling pathways in cholangiocarcinoma growth is unknown. Thus, our aims were to determine the role of IL-6-mediated signaling mechanisms, and in particular the MAPK pathways, in the growth regulation of human cholangiocarcinoma.

The aim of this study was to determine whether taurocholate prevents vagotomy-induced cholangiocyte apoptosis. After bile duct ligation (BDL) + vagotomy, rats were fed taurocholate for 1 wk in the absence or presence of wortmannin. Caspase involvement was evaluated by measurement of caspase 8, 9, and 3 activities. Proliferation was determined by morphometry and PCNA immunoblots. Changes in phosphatidylinositol 3-kinase (PI3-kinase) activity were estimated by the expression of the phosphorylated Akt protein. Apically located Na+-dependent bile acid transporter (ABAT) expression and activity were evaluated by immunoblots and [3H]taurocholate uptake, respectively. Cholangiocyte apoptosis increased, whereas proliferation decreased in BDL + vagotomy rats. Taurocholate feeding prevented vagotomy effects on cholangiocyte functions, which were abolished by wortmannin. ABAT expression and activity as well as phosphorylated Akt protein expression were reduced by vagotomy but restored by taurocholate. The activities of caspase 8, 9, and 3 increased in BDL + vagotomy rats but were restored by taurocholate. The protective effect of taurocholate was associated with maintenance of ABAT activity, downregulation of caspase 8, 9, and 3, and activation of PI3-kinase. Bile acids are important in modulating cholangiocyte proliferation in denervated livers.

Although mitogen-activated protein kinase (MAPK) pathways play a key role in cell growth, their role in mediating the altered growth phenotype of transformed cells remains unclear. The p44/p42 MAPK signaling cascades are activated by mitogenic stimulation of human cholangiocytes. In contrast, the p38 MAPK pathway is activated by mitogen stimulation of malignant, but not nonmalignant cholangiocytes. Thus, our aims were to determine the role of p38 MAPK signaling in mediating the growth phenotype of transformed cholangiocytes. KMCH-1 malignant human cholangiocytes required the presence of serum for proliferation, but were able to grow in reduced serum conditions. Inhibition of p38 MAPK decreased serum-dependent proliferation of KMCH-1 cells. Furthermore, inhibition of p38 MAPK, but not of p44/p42 MAPK, reduced anchorage-independent growth of KMCH-1 cells. Although both p38 and p44/p42 MAPK are activated in response to mitogens, they have divergent effects on anchorage-independent growth. Inhibition of p38 MAPK, but not of p44/p42 MAPK signaling, decreased cell cycle progression and increased expression of the cyclin-dependent kinase inhibitor p21 WAF1/CIP1 . However, expression of p27 KIP1 or p16 INK4A was not altered by either pathway. Thus, mitogen activation of p38 MAPK decreases expression of p21 WAF1/CIP1 and mediates growth independent of anchorage signals, whereas mitogen activation of p44/p42 MAPK mediates an anchorage signal-dependent growth pathway. These data provide a link between aberrant stress-activated cell signaling and the altered growth phenotype of transformed cells that may be important for the development of therapies to limit transformed cell growth. (HEPATOLOGY 2001;33:43-51.)Cholangiocarcinoma is an aggressive malignancy for which we lack effective therapy. 1 Despite attempts at surgical resections, transplantation, chemotherapy, or radiation therapy, the prognosis of advanced cholangiocarcinoma remains dismal, and has not significantly changed over the past several decades. Thus, there exists a marked need for newer therapeutic strategies for unresectable cholangiocarcinoma. A potential strategy for this aggressive malignancy would be to inhibit proliferation of malignant biliary epithelial cells. Selective survival and proliferation of malignant transformed cells involves dysregulation of cell growth signaling pathways. Elucidation of the intracellular signaling events mediating transformed cell proliferation may thus lead to potential therapeutic targets for limiting tumor growth.Although chronic biliary tract inflammation predisposes to the development of cholangiocarcinoma, the relationship between chronic inflammation and subsequent malignant transformation remains poorly understood. 2,3 Our recent studies have shown the involvement of mitogen-activated protein kinase (MAPK) signaling cascades in mediating biliary epithelial cell proliferation in response to inflammatory mediators. 4 MAPK pathways represent a key point of signal convergence from distinct transmembrane signaling pathw...

The p38 mitogen-activated protein kinase (MAPK) signaling pathway is aberrantly expressed and maintains transformed cell growth in malignant human cholangiocytes. Because cell growth requires and is intimately related to protein synthesis, our aims were to assess the effect of p38 MAPK signaling on protein synthesis during growth of malignant human cholangiocytes. Inhibition of p38 MAPK activity during mitogenic stimulation decreased protein synthesis rates and tumor cell xenograft growth in nude mice. Altered protein synthesis resulted from decreased translational efficiency with impaired initiation of translation. Mitogenic stimulation resulted in phosphorylation of the eukaryotic initiation factor (eIF)-4E. Inhibition of p38 MAPK signaling or functional dysregulation of translation by small interfering double-stranded RNA (siRNA) to eIF-4E decreased anchorage-independent growth of malignant cholangiocytes. In conclusion, these studies identify a relationship between p38 MAPK activity and the regulation of protein synthesis during human cholangiocarcinoma growth. As protein synthesis is intimately linked to cell growth, dysregulation of translation initiation is a mechanism by which cellular p38 MAPK signaling participates in growth regulation of malignant cholangiocytes. (HEPATOLOGY 2003;38:158-166.)

Tumor necrosis factor (TNF)-alpha plays a critical role in epithelial cell injury. However, the role of TNF-alpha in mediating cholangiocyte injury under physiological or pathophysiological conditions is unknown. Thus we assessed the effects of TNF-alpha alone or following sensitization by actinomycin D on cell apoptosis, proliferation, and basal and secretin-stimulated ductal secretion in cholangiocytes from normal or bile duct-ligated (BDL) rats. Cholangiocytes from normal or BDL rats were highly resistant to TNF-alpha alone. However, presensitization by actinomycin D increased apoptosis in cholangiocytes following BDL and was associated with an inhibition of proliferation and secretin-stimulated ductal secretion. Thus TNF-alpha mediates cholangiocyte injury and altered ductal secretion following bile duct ligation. These observations suggest that cholestasis may enhance susceptibility to cytokine-mediated cholangiocyte injury.