Epimorphin regulates bile duct formation via effects on mitosis orientation in rat liver epithelial stem-like cells.

Junnian Zhou,Yunfang Wang,Lipeng Qin,Chen Sang,Yali Jia,Wen Yue,Chun Yang,Jing Wang,Hailei Yao,Qinghua Hu,Lei Zhao,Shuangshuang Shi,Fengyuan Zhuang,Xue Nan,Xuetao Pei,Joseph Najbauer

doi:10.1371/journal.pone.0009732

Abstract

Understanding how hepatic precursor cells can generate differentiated bile ducts is crucial for studies on epithelial morphogenesis and for development of cell therapies for hepatobiliary diseases. Epimorphin (EPM) is a key morphogen for duct morphogenesis in various epithelial organs. The role of EPM in bile duct formation (DF) from hepatic precursor cells, however, is not known. To address this issue, we used WB-F344 rat epithelial stem-like cells as model for bile duct formation. A micropattern and a uniaxial static stretch device was used to investigate the effects of EPM and stress fiber bundles on the mitosis orientation (MO) of WB cells. Immunohistochemistry of liver tissue sections demonstrated high EPM expression around bile ducts in vivo. In vitro, recombinant EPM selectively induced DF through upregulation of CK19 expression and suppression of HNF3α and HNF6, with no effects on other hepatocytic genes investigated. Our data provide evidence that EPM guides MO of WB-F344 cells via effects on stress fiber bundles and focal adhesion assembly, as supported by blockade EPM, β1 integrin, and F-actin assembly. These blockers can also inhibit EPM-induced DF. These results demonstrate a new biophysical action of EPM in bile duct formation, during which determination of MO plays a crucial role.

Highlights

Duct formation (DF) is an important process in development and regeneration of many epithelial organs including lung, kidney, mammary glands and liver, and is known to be regulated by diffusible morphogens and elements of the insoluble extracellular matrix [1]
Our results may extend the previous observations that bile ducts, hepatic blood vessels and mesenchyme development is cross-regulated [2] since EPM is highly expressed on mesenchyme around both bile ducts and blood vessels. These results suggest that EPM may be involved in maintaining normal morphogenesis and/or regenerating bile ducts in vivo, and prompted us to investigate whether EPM plays a role in the duct-like differentiation of WB-F344 cells into biliary lineage
Understanding how hepatic precursor cells differentiate into bile ducts is crucial for epithelial morphogenesis studies and for the development of future therapies for hepatobiliary diseases

Summary

Introduction

Duct formation (DF) is an important process in development and regeneration of many epithelial organs including lung, kidney, mammary glands and liver, and is known to be regulated by diffusible morphogens and elements of the insoluble extracellular matrix [1]. Intrahepatic bile ducts, a series of tubules transporting bile produced by hepatocytes to the gallbladder, are an important duct system within the liver. The lumen of these bile ducts is lined with biliary epithelial cells which share a common origin with hepatocytes [2]. The mechanism for controlling bile duct formation, especially the effect of biophysical properties, remains largely unknown. While it is known that extracellular matrix complexes such as Matrigel can combine with soluble growth factors to meet the minimum requirements for DF of hepatoblasts in vitro [3], little is known about the function of single matrix proteins in DF

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