Advertisement
Hedgehog signaling in biliary fibrosis
Linda E. Greenbaum
Division of Gastroenterology, Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania, USA.
Address correspondence to: Linda E. Greenbaum, Division of Gastroenterology, Department of Medicine, University of Pennsylvania, 600 Clinical Research Building, 415 Curie Boulevard, Philadelphia, Pennsylvania 19104, USA. Phone: (215) 573-1868; Fax: (215) 573-2024; E-mail: greenbal@mail.med.upenn.edu.
Find articles by Greenbaum, L. in: JCI | PubMed | Google Scholar
Published September 18, 2008 - More info
J Clin Invest. 2008;118(10):3263–3265. https://doi.org/10.1172/JCI37189.
© 2008 The American Society for Clinical Investigation
Related article:
Abstract
Epithelial-mesenchymal transitions (EMTs) play an important role in tissue construction during embryogenesis, and evidence suggests that this process may also help to remodel some adult tissues after injury. Activation of the hedgehog (Hh) signaling pathway regulates EMT during development. This pathway is also induced by chronic biliary injury, a condition in which EMT has been suggested to have a role. We evaluated the hypothesis that Hh signaling promotes EMT in adult bile ductular cells (cholangiocytes). In liver sections from patients with chronic biliary injury and in primary cholangiocytes isolated from rats that had undergone bile duct ligation (BDL), an experimental model of biliary fibrosis, EMT was localized to cholangiocytes with Hh pathway activity. Relief of ductal obstruction in BDL rats reduced Hh pathway activity, EMT, and biliary fibrosis. In mouse cholangiocytes, coculture with myofibroblastic hepatic stellate cells, a source of soluble Hh ligands, promoted EMT and cell migration. Addition of Hh-neutralizing antibodies to cocultures blocked these effects. Finally, we found that EMT responses to BDL were enhanced in patched-deficient mice, which display excessive activation of the Hh pathway. Together, these data suggest that activation of Hh signaling promotes EMT and contributes to the evolution of biliary fibrosis during chronic cholestasis.
Authors
Alessia Omenetti, Alessandro Porrello, Youngmi Jung, Liu Yang, Yury Popov, Steve S. Choi, Rafal P. Witek, Gianfranco Alpini, Juliet Venter, Hendrika M. Vandongen, Wing-Kin Syn, Gianluca Svegliati Baroni, Antonio Benedetti, Detlef Schuppan, Anna Mae Diehl
Cholangiopathies represent a spectrum of both congenital and acquired disorders characterized by chronic bile duct injury (reviewed in refs. 1, 2). Although cholangiocytes initially have the ability to proliferate in response to cellular injury, late-stage cholangiopathies are associated with loss of bile ducts and progressive fibrosis, resulting in cirrhosis and liver failure.
Ductular reactions, composed of small and apparently proliferating cholangiocytes in the periportal region, are detected in many forms of chronic liver injury, including those that primarily effect cholangiocytes. These cholangiocytes have been termed reactive cholangiocytes, and they acquire phenotypic behaviors not observed in the noninjured state, including secretion of inflammatory cytokines, chemoattractant proteins, and inhibitors of apoptosis. Reactive cholangiocytes are believed to arise from a progenitor cell compartment located along the terminal bile ductules and in the canal of Hering (2). However, the origin of these reactive cholangiocytes has not yet been established definitively. Evidence from experimental animal models has shown that crosstalk among reactive cholangiocytes, myofibroblasts, endothelial cells, and inflammatory cells contributes to cholangiocyte proliferation as well as the fibrotic response (1, 2). The identity and relative contribution of these paracrine mediators and the mechanisms that directly regulate cholangiocyte proliferation and fibrogenesis remain incompletely understood.
-
Potential role of epithelial-mesenchymal transition in cholestatic liver injury
In addition to secreting numerous regulatory molecules in response to cholangiocyte injury, reactive cholangiocytes lose some of their epithelial phenotypic characteristics and acquire a more mesenchymal phenotype characterized by the loss of E-cadherin and increased expression of some mesenchymal markers. During epithelial-mesenchymal transition (EMT), epithelial cells lose polarity and cell-cell communication and acquire migratory and invasive phenotypic cellular behaviors. This process is important for tissue remodeling during normal development and more recently has been recognized as a component of repair processes in fibrotic diseases and malignancy (3, 4). EMT has been implicated in fibrotic diseases of the biliary tract as a potential mechanism for disappearance of bile ductules and concomitant increase in fibrosis (5–7). How EMT might be regulated in chronic biliary diseases is not known.
Previously, Omenetti and colleagues reported that the hedgehog signaling pathway is activated in both cholangiocytes and fibroblastic cells in experimental models of biliary injury and in livers of patients with primary biliary cirrhosis (PBC) (8, 9). Because the hedgehog signaling pathway is known to be a positive effector of EMT in other tissues, the authors hypothesized that hedgehog signaling pathways could also be involved in EMT in response to cholestatic liver injury and fibrosis (10, 11).
The study by Omenetti and colleagues in the current issue of the JCI (12) tests this hypothesis, demonstrating that the mesenchymal marker S100A4 is expressed in a subset of ductular and fibroblastic cells in livers of patients with PBC, confirming previous findings (5, 6). Using a rat model of reversible chronic bile duct obstruction, they further show that expression of this mesenchymal marker is induced following bile duct obstruction. They confirm that this activation of a mesenchymal marker occurs within cholangiocytes by demonstrating both upregulation of mesenchymal markers and decreased expression of epithelial markers in cholangiocytes isolated from the livers of bile duct–ligated rats. Using the same approach, they demonstrate that cholangiocytes isolated from bile duct–ligated livers express hedgehog proteins and these hedgehog proteins are coexpressed with mesenchymal markers in ductular cells from the livers of patients with PBC.
These findings define a novel association between the expression of hedgehog proteins and mesenchymal markers in cholangiocytes after bile duct injury (12). To establish a functional relationship between hedgehog proteins and EMT, the authors used an in vitro system in which a cholangiocyte cell line is cocultured with a hepatic stellate cell line to test whether paracrine hedgehog signaling not only affects expression of mesenchymal markers but also increases the migratory behavior of cholangiocytes. They demonstrate that soluble factors expressed by hepatic stellate cells induce expression of mesenchymal markers in cholangiocytes. More importantly, using carefully performed cell migration assays, the authors show that soluble factors from hepatic stellate cells increase cholangiocyte motility and migration, two hallmarks of the mesenchymal phenotype. When conditioned medium from hepatic stellate cells was treated with hedgehog-neutralizing antibodies, the expression of mesenchymal markers and cholangiocyte migratory behavior were significantly reduced. This experiment provides what is believed to be the first functional evidence that hepatic stellate cells can influence cholangiocyte migratory behavior through a mechanism that is dependent upon hedgehog-mediated paracrine signaling (Figure 1).
Figure 1Model of paracrine signaling in a ductular reaction in the cholestatic liver. (A) Schematic of the portal triad of the portal vein, hepatic artery, and bile duct in the normal liver. Hepatic stellate cells, the site of vitamin A storage, are located in the Disse space near hepatocytes. Portal myofibroblasts are also in close proximity to the bile ducts. (B) After bile duct ligation and cholestatic liver injury, the liver responds with a ductular reaction and the expansion of the biliary tree. In their study in this issue of the JCI, Omenetti and colleagues (12) propose that hepatic stellate cells produce and release sonic hedgehog (Hh) in response to the injury, which acts on a subpopulation of bile duct epithelial cells known as cholangiocytes to activate several markers of mesenchymal cells. The authors suggest that cholangiocytes then undergo EMT and migrate into the parenchyma. The latter hypothesis remains to be proven in vivo. Also, additional signals from the portal myofibroblasts likely contribute to the activation of bile duct epithelial cells.
To investigate whether alterations in hedgehog signaling are associated with increased EMT in vivo, Omenetti and colleagues performed bile duct ligation on mice haploinsufficient for the hedgehog inhibitor patched (Ptc) (12); these mice exhibit increased hedgehog signaling activity. As predicted, the livers of the mice showed enhanced fibrogenesis in response to bile duct ligation and elevated expression of the hedgehog transcription factor Gli2 and several mesenchymal markers. In the future, it will be important to establish that the change in cellular behavior of the cholangiocyte cell line, especially with respect to cell migration, seen in vitro can also be linked to hedgehog signaling in vivo.
Copyright © 2024 American Society for Clinical Investigation
ISSN: 0021-9738 (print), 1558-8238 (online)