The biphasic nature of hypoxia-induced directional migration of activated human hepatic stellate cells

Liver fibrogenesis is sustained by pro‐fibrogenic myofibroblast‐like cells (MFs), mainly originating from activated hepatic stellate cells (HSC/MFs) or portal (myo)fibroblasts, and is favoured by hypoxia‐dependent angiogenesis. Human HSC/MFs were reported to express vascular‐endothelial growth factor (VEGF) and VEGF‐receptor type 2 and to migrate under hypoxic conditions. This study was designed to investigate early and delayed signalling mechanisms involved in hypoxia‐induced migration of human HSC/MFs. Signal transduction pathways and intracellular generation of reactive oxygen species (ROS) were evaluated by integrating morphological, cell, and molecular biology techniques. Non‐oriented and oriented migration were evaluated by using wound healing assay and the modified Boyden's chamber assay, respectively. The data indicate that hypoxia‐induced migration of HSC/MFs is a biphasic process characterized by the following sequence of events: (a) an early (15 min) and mitochondria‐related increased generation of intracellular ROS which (b) was sufficient to switch on activation of ERK1/2 and JNK1/2 that were responsible for the early phase of oriented migration; (c) a delayed and HIF‐1α‐dependent increase in VEGF expression (facilitated by ROS) and its progressive, time‐dependent release in the extracellular medium that (d) was mainly responsible for sustained migration of HSC/MFs. Finally, immunohistochemistry performed on HCV‐related fibrotic/cirrhotic livers revealed HIF‐2α and haem‐oxygenase‐1 positivity in hepatocytes and α‐SMA‐positive MFs, indicating that MFs were likely to be exposed in vivo to both hypoxia and oxidative stress. In conclusion, hypoxia‐induced migration of HSC/MFs involves an early, mitochondrial‐dependent ROS‐mediated activation of ERK and JNK, followed by a delayed‐ and HIF‐1α‐dependent up‐regulation and release of VEGF. Copyright © 2012 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.