Wendy M. Mars

Bone marrow stem cells develop into hematopoietic and mesenchymal lineages but have not been known to participate in production of hepatocytes, biliary cells, or oval cells during liver regeneration. Cross-sex or cross-strain bone marrow and whole liver transplantation were used to trace the origin of the repopulating liver cells. Transplanted rats were treated with 2-acetylaminofluorene, to block hepatocyte proliferation, and then hepatic injury, to induce oval cell proliferation. Markers for Y chromosome, dipeptidyl peptidase IV enzyme, and L21-6 antigen were used to identify liver cells of bone marrow origin. From these cells, a proportion of the regenerated hepatic cells were shown to be donor-derived. Thus, a stem cell associated with the bone marrow has epithelial cell lineage capability.

Hepatocyte growth factor, also known as scatter factor, is a complete mitogen for hepatocytes that bears sequence and structural homology with plasminogen. Because it exists in both a mitogenically inactive single-chain form and an active two-chain form, we were interested in determining whether plasminogen activators could properly cleave single-chain hepatocyte growth factor to generate active two-chain hepatocyte growth factor. Herein we report that both urokinase-type plasminogen activator and tissue-type plasminogen activator can cleave single-chain hepatocyte growth factor, generating two-chain hepatocyte growth factor. When equal quantities of plasminogen activator-treated and activator-untreated hepatocyte growth factor are compared in serum-free in vitro bioassays, the treated hepatocyte growth factor is mitotically more active. Also, urokinase-type plasminogen activator was inactive against hepatocyte growth factor molecules with a mutated cleavage site. This suggests that urokinase-type and tissue-type plasminogen activator may be natural biological regulators of hepatocyte growth factor. Because the active form of hepatocyte growth factor is a powerful stimulator of DNA synthesis and cell motility, these findings may be relevant in understanding the role of plasminogen activators in the biology of cancer invasion and metastasis.

Hepatocyte growth factor (HGF) and Wnt signaling pathways have been shown to be important in embryogenesis and carcinogenesis. The aim of this study was to elucidate the mechanism of functional similarities observed in the two pathways. We used normal rat liver, primary hepatocyte cultures and a dominant-negative Met expression system to study the effect of HGF on Wnt pathway components. We demonstrate novel association of beta-catenin and Met, a tyrosine kinase receptor of HGF, at the inner surface of the hepatocyte membrane. HGF induces dose-dependent nuclear translocation of beta-catenin in primary hepatocyte cultures that is Wnt independent. The source of beta-catenin for translocation in hepatocytes is the Met-beta-catenin complex, which appears to be independent of the E-cadherin-beta-catenin complex. To test the functionality of this association, we used a dominant-negative Met expression system that expresses only the extracellular and transmembrane regions of the beta-subunit of Met. A loss of Met-beta-catenin association resulted in abrogation of nuclear translocation of beta-catenin upon HGF stimulation. This event is tyrosine phosphorylation dependent, and the association of Met and beta-catenin is crucial for this event. We conclude that the HGF causes similar redistribution of beta-catenin as Wnt-1 in the hepatocytes and that this effect is attributable to subcellular association of Met and beta-catenin. The intracellular kinase domain of Met is essential for tyrosine phosphorylation and nuclear translocation of beta-catenin. Part of the multifunctionality of HGF might be attributable to nuclear beta-catenin and the resulting target gene expression.

Tissue-type plasminogen activator (tPA) is one of the major components in the matrix proteolytic network whose role in the pathogenesis of renal interstitial fibrosis remains largely unknown. Here, we demonstrate that ablation of tPA attenuated renal interstitial fibrotic lesions in obstructive nephropathy. Mice lacking tPA developed less morphological injury and displayed a reduced deposition of interstitial collagen III and fibronectin as well as total tissue collagen in the kidneys after sustained ureteral obstruction, when compared with their wild-type counterparts. Deficiency of tPA selectively blocked tubular epithelial-to-myofibroblast transition (EMT), but did not affect myofibroblastic activation from interstitial fibroblasts. A marked decrease in matrix metalloproteinase-9 (MMP-9) induction was found in the obstructed kidneys of tPA(-/-) mice, which led to a dramatic preservation of the structural and functional integrity of tubular basement membrane (TBM). In vitro, tPA induced MMP-9 gene expression and protein secretion in renal interstitial fibroblasts. Thus, increased tPA is detrimental in renal interstitial fibrogenesis through a cascade of events that lead to MMP-9 induction, TBM destruction, and promotion of EMT. Our findings establish a crucial and definite importance of EMT in the pathogenesis of renal interstitial fibrosis at the whole-animal level.