Contribution of HGF to Liver Regeneration by Strong Mobilization of Bone Marrow Stem Cells.

Cell plasticity of bone marrow stem cells to hepatocytes is known, however, the details are still unclear. hepatocyte growth factor (HGF) promotes an increase in liver stem cells in severely injured liver, but intervention to bone marrow stem cell is unclear. We examined a role of HGF in bone marrow stem cell-mediated liver regeneration in order to obtain effective liver regeneration.

First, we found that the phenotype of stem cells, which can differentiate into hepatocytes, is Lin⁻c-kit⁺Sca-1⁺CD34⁻ in bone marrow or Lin⁻c-kit⁺Sca⁻1⁺CD34⁺ in peripheral blood mobilized by G-CSF. We transplanted single Lin⁻c-kit⁺Sca-1⁺CD34⁻ bone marrow cell harvested from male EGFP mouse to female wild type mouse, and then, using this GFP-chimera-mouse, we found that bone marrow origin GFP⁺ and Y chromosome⁺ hepatic cells were present in liver after acute liver damage. Next, single Lin⁻c-kit⁺Sca-1⁺CD34⁺ peripheral blood cell, which was mobilized in peripheral blood by administration of G-CSF, was transplanted into the portal vein of the wild type mouse which was given hepatic damage. We found that the GFP-positive cells also expressed albumin.

Second, we investigated whether the HGF can mobilize stem cells from bone marrow to peripheral blood. In peripheral blood of HGF transgenic mice, 1.1% developed CD34⁺ cells and 20±3 colony forming cells of 1X10⁶ peripheral blood mononuclear cells were shown. Colony forming cells were found in the mouse into which an HGF-expressing adenovirus was administered. After injection of rHGF to mice, a significant time-dependent increase of percentage of CD34⁺ cells in the PB was noted at the first 3 hours and CD34⁺ cells were increased in dose-dependent manner of rHGF and reached plateau level at 100 m/kg. The mice having transplantation with PB cells from 100 mg HGF-treated mice for 4days showed engraftment 2 months after transplantation. Upon activation of rHGF in mouse MS-5 stromal cells, phosphorylation c-Met and SCF were up-regulated, while VCAM -1, MMP-9, SDF -1 or CXCR4 were not changed. SCF level in conditioned media was also increased after the HGF stimulation.

Finally, we examined whether the bone marrow stem cells in PB mobilized by HGF transdifferentiate into hepatocytes. Using GFP-chimera-mice given acute liver injury after administration of retorolusine and CCl⁴, the levels of GFP⁺ cells in liver of GFP-chimera-mice 2 months after treatment by PBS and HGF were 2.2±1.4% and 12.7±3.6%, respectively (p<0.01). In conclusion, HGF can mobilize stem cells with long-term engraftment capabilities from bone marrow to peripheral blood, resulting in contribution to liver regeneration.