Facilitation of survival in a rat fulminant hepatic failure model by combination therapy using recombinant G-CSF and tacrolimus.

The mortality rate of fulminant hepatic failure (FHF) is high because of retarded liver regeneration. Recombinant human granulocyte colony-stimulating factor (rHuG-CSF) and tacrolimus are known to be immunosuppressive and supportive to liver regeneration. We investigated the effects of their combination therapy in a rat FHF model with a 68% partial hepatectomy and 24% liver necrosis. All rats without drug pretreatment died within 55 h. The median time was prolonged from 37 to 52 h by rHuG-CSF (250 microg/kg/day s.c. on days -5 to 0) and to 46 h by tacrolimus (0.5 mg/kg/day i.m. on days -2 to 0). Notably, the combination therapy facilitated DNA biosynthesis and survival prolongation, with a median of 77 h. The interferon-gamma (IFN-gamma) protein levels and natural killer cell (NK) activity in the liver were low at 12 h, and no further inhibition was detected by any treatment. Tacrolimus significantly upregulated the mRNA levels of insulin receptors and transforming growth factor-alpha (TGF-alpha), whereas rHuG-CSF did not. Regarding tissue remodeling-related factors, rHuG-CSF upregulated mRNA levels of vascular endothelial growth factor (VEGF) and matrix metalloproteinase- 9 (MMP-9), whereas tacrolimus did not. The combination treatment upregulated protein levels of both insulin receptors and VEGF. These results suggest that tacrolimus improves the hepatocyte replication and rHuG-CSF contributes to tissue reconstitution, and this combination therapy directly facilitates liver regeneration in the FHF model.

G-CSF-induced evacuation of sinusoidal NK cells and the facilitation of liver regeneration in a partial hepatectomy.

Since liver regeneration after partial hepatectomy (PHx) is known to improve by pretreatment with recombinant human G-CSF (rhG-CSF), we investigated the mechanism by evaluating the distribution and activity of sinusoidal NK cells. F344 rats were treated with rhG-CSF (250 microg/kg/day) for 5 days before PHx. Pretreatment with rhG-CSF improved the serum ALT levels and DNA biosynthesis of the remnant liver tissues at 20 h after PHx. Notably, the rhG-CSF pretreatment decreased the number of NK cells in the liver determined by immunohistochemistry using anti-NKR-P1A mAb before and at 20 h after PHx with no significant change in the NK activity per cell base, while also increasing the number of NK cells in the peripheral blood detected by flow cytometry. The rhG-CSF induced a pre-PHx downregulation of the IL-12p70 protein levels, while also promoting the post-PHx reduction of the protein levels of IL-12p70 and IFN-gamma. Conversely, rhG-CSF had no effect on the pre-PHx mRNA levels or the PHx-induced upregulation of mRNA levels of TNF-alpha, IL-1beta, IL-6, TGF-beta, IL-10, HGF, and c-Met determined by real-time RT-PCR. These results strongly suggest that rhG-CSF-induced facilitation of liver regeneration is achieved by immunoregulation through the intrahepatic IL-12 downregulation and evacuation of sinusoidal NK cells.

Downregulation of interleukin-12p35 and upregulation of interleukin-12p40 mRNA expression in heart allografts by blood transfusion from granulocyte colony-stimulating factor-treated donors.

Pretransplant treatment of recipients with recombinant human granulocyte colony-stimulating factor (rhG-CSF, 250 microg/kg/day s.c. for 5 days) facilitates heart allograft acceptance in tacrolimus-treated rat recipients. We examined effectiveness of transfusion of in vivo rhG-CSF-treated blood since rhG-CSF induces immunoregulatory cells in human blood. DA heart grafts were transplanted into tacrolimus (2 mg/kg i.m. on day 0)-treated Lewis recipients. Although graft survival prolongation by blood transfusion on day 0 from rhG-CSF-treated syngeneic Lewis was comparable to that in directly rhG-CSF-pretreated recipients (p = 0.22), transfusion of rhG-CSF-treated allogeneic DA blood was much more effective (p = 0.0016). Intragraft cytokine mRNA levels were measured by reverse transcription and real-time polymerase chain reaction at 24 h after transplantation. IL-12p35 expression was downregulated by both treatments. Notably, IL-12p40 was upregulated by rhG-CSF-treated DA blood transfusion but downregulated by transfusion of rhG-CSF-treated isogeneic blood. Differential expression of IL-12 subunits was associated with facilitation of graft acceptance by rhG-CSF-treated donor blood transfusion.