Factor VIII expression in azoxymethane-induced murine fulminant hepatic failure

Abstract

Fulminant hepatic failure (FHF) in humans produces a bleeding diathesis due in large part to a reduction in the biosynthesis of liver-derived coagulation factors. Remarkably, factor VIII procoagulant activity is elevated in most of these patients despite widespread liver cell death. FHF can be modeled in mice by administration of azoxymethane, the active ingredient found in cycad palm nuts. We compared the expression of factor VIII to other hepatic hemostatic factors in azoxymethane-induced murine FHF. Mice displayed dose-dependent decreases in all coagulation factor activities measured, including factors V, VII, VIII, and IX. At the highest dose of azoxymethane (50 g/g body weight), factor VIII activity in plasma decreased by 98% within 36 hours after treatment, which was associated with an 80% reduction in hepatic factor VIII messenger RNA (mRNA). In contrast, factor VIII mRNA levels in spleen, kidney, and lung tissue of azoxymethane-treated mice were unchanged. Cellular damage in these mice appeared to be limited to hepatocytes as evident by histologic examination. This finding is supported by 2 observations. First, hepatic mRNA levels of von Willebrand factor, which is synthe- IntroductionDespite more than 50 years of investigation, the nature of factor VIII biosynthesis in vivo remains poorly understood. [1][2][3][4] Factor VIII messenger RNA (mRNA) is present in nearly every tissue examined, including liver, spleen, lymph node, heart, brain, lung, kidney, testes, muscle, and placenta. [5][6][7] The liver clearly is an important site of synthesis because liver transplantation cures human and canine hemophilia A. [8][9][10] However, the relative contribution of hepatocytes and liver sinusoidal endothelial cells remains uncertain. 7,[11][12][13][14][15] Furthermore, the contribution of extrahepatic biosynthesis of factor VIII in health and disease states is not known.Paradoxically, there usually is a significant rise in plasma factor VIII activity in human fulminant hepatic failure (FHF). [16][17][18][19] This increase can be more than 14 times normal levels and occurs simultaneously with profound decreases in the circulating levels of vitamin K-dependent and fibrinolytic plasma proteins. 16,20,21 The analysis of this phenomenon has been hindered by the lack of a reproducible animal model of FHF. However, a murine model of FHF recently was developed using the hepatotoxin azoxymethane (AOM), which is found in cycad palm nuts on the island of Guam. 22 AOM-treated mice develop hepatocellular necrosis, elevated liver transaminase and ammonia levels in blood and hepatic encephalopathy. They die within days, with death related to the dose of AOM. In this study, we examined the expression of factor VIII and several other hemostatic factors, including factors V, VII, and IX, and von Willebrand factor (VWF) in AOM-induced murine FHF. Materials and methods MaterialsMale C57BL/6 mice (22-30 g body weight) were purchased from Charles River Laboratory (Wilmington, MA). AOM and TriReagent were purchased from Sigma (St Lou...

Targeting FVIII expression to endothelial cells regenerates a releasable pool of FVIII and restores hemostasis in a mouse model of hemophilia A

“…1 The precise site of FVIII biosynthesis and the cellular origin of the regulated releasable pool of FVIII remain unclear. [2][3][4][5] It has been proposed that synthesis of FVIII occurs in a subpopulation of endothelial cells. [6][7][8][9][10][11] Targeting FVIII expression to lung endothelial cells 12 or liver sinusoidal endothelial cells 13 has been shown to result in phenotypic correction in hemophilia A mice.…”

Decreased factor VIII levels during acetaminophen-induced murine fulminant hepatic failure

“…The decrease in fVIII activity observed in the present study corresponds well with the results previously reported in azoxymethane-treated mice in which, at the highest dose of azoxymethane tested (50 g/g body weight), fVIII activity dropped to 10% of control levels at 24 hours after administration. 10 Several factors have been shown to influence circulating fVIII levels, including VWF whose levels display a strong positive correlation with fVIII levels (reviewed in Kamphuisen et al 15 ). Accordingly, in the complete absence of VWF, fVIII levels are reduced by approximately 80% in mice.…”

“…We previously reported that fVIII levels were decreased in a murine model of FHF induced by the hepatotoxin, azoxymethane. 10 Because azoxymethane has not been proven to cause FHF in humans, we have continued to identify other models that may be more representative of human FHF pathophysiology. In the current study we sought to determine whether fVIII levels are elevated in a recently described murine model of acetaminophen-induced FHF 11 and possibly identify alternative sites of fVIII biosynthesis in vivo.…”

Factor VIII biosynthesis: new inspirations?

“…3 However, hepatocyte-specific toxins eliminate almost all factor VIII from the circulation in this species. 4 Northern blot analysis of factor VIII expression is technically difficult (since the transcript is large and not very abundant) and has yielded inconclusive results. More sensitive methods to detect factor VIII transcripts (eg, reverse transcriptase-polymerase chain reaction [RT-PCR]) have shown factor VIII mRNA in liver, spleen, and peripheral lymphocytes in humans and other species.…”

“…More sensitive methods to detect factor VIII transcripts (eg, reverse transcriptase-polymerase chain reaction [RT-PCR]) have shown factor VIII mRNA in liver, spleen, and peripheral lymphocytes in humans and other species. [4][5][6][7] Furthermore, in humans it is known that factor VIII levels remain normal or even elevated in patients with liver disease, even when other coagulation factors are greatly diminished. All of these findings point to hepatic as well as extrahepatic synthesis of factor VIII.…”