Wilson disease is caused by accumulation of Cu(2+) in cells, which results in liver cirrhosis and, occasionally, anemia. Here, we show that Cu(2+) triggers hepatocyte apoptosis through activation of acid sphingomyelinase (Asm) and release of ceramide. Genetic deficiency or pharmacological inhibition of Asm prevented Cu(2+)-induced hepatocyte apoptosis and protected rats, genetically prone to develop Wilson disease, from acute hepatocyte death, liver failure and early death. Cu(2+) induced the secretion of activated Asm from leukocytes, leading to ceramide release in and phosphatidylserine exposure on erythrocytes, events also prevented by inhibition of Asm. Phosphatidylserine exposure resulted in immediate clearance of affected erythrocytes from the blood in mice. Accordingly, individuals with Wilson disease showed elevated plasma levels of Asm, and displayed a constitutive increase of ceramide- and phosphatidylserine-positive erythrocytes. Our data suggest a previously unidentified mechanism for liver cirrhosis and anemia in Wilson disease.

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Background. There is an ongoing discussion about the risks of di(2-ethylhexyl) phthalate (DEHP) exposure for the general population as well as for specific subgroups in various medical settings. Haemodialysis patients certainly belong to the group with the highest exposure taking into account the repeated treatments over a long period of time. Many studies have shown that DEHP metabolites are more active with regard to cellular responses than DEHP itself. Although 4-heptanone has been shown to be a DEHP metabolite in rats, this has never been tested in humans. On the other hand, 4-heptanone was reported to be associated with diabetes mellitus. Methods. After establishing analytical methods for all postulated metabolites, we analysed (i) plasma samples from 50 patients on haemodialysis and 50 controls; (ii) urine samples from 100 diabetic patients and 100 controls; and (iii) urine samples from 10 controls receiving DEHP intravenously. Results. 4-Heptanone concentrations in urine did not differ between controls (128.6±11.4 mg/l, mean± SEM) and diabetic patients (131.2±11.6 mg/l) but were significantly elevated in plasma from haemodialysis patients (95.9±9.6 mg/l) compared with controls (10.4±0.5 mg/l). Exposure to DEHP led to a significant increase (P<0.001) of the metabolite 4-heptanone and all the proposed intermediates in urine of healthy persons within 24 h. Conclusions. These studies show that 4-heptanone is not associated with diabetes but is a major DEHP metabolite in humans. Studies concerning the toxicity of DEHP in haemodialysis patients and other highly exposed groups should therefore include 4-heptanone together with DEHP and its primary metabolites mono(2-ethylhexyl) phthalate (MEHP) and 2-ethylhexanol.

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Background:Sorafenib has recently been shown to reduce tumour growth in hepatoblastoma (HB) xenografts. The effect of a combined administration with cytostatic agents was now investigated.Methods:Cell viability after treatment with sorafenib and different cytostatic agents was evaluated in two HB cell lines (HUH6 and HepT1) using MTT assay. ERK signalling was investigated by western blot, NOXA expression by rt-PCR, and formation of DNA adducts using immunocytology. NMRI mice bearing subcutaneous HUH6-derived tumours were treated with sorafenib alone or in combination with cisplatin. Tumour progression, viability, apoptosis, and vascularisation were monitored by tumour volume, AFP levels, TUNEL assay, and CD31 immunostaining, respectively.Results:The combination of sorafenib and cisplatin led to a remarkable decrease in cell viability. The cisplatin-induced enhanced ERK1/2 activation, but not NOXA expression and the formation of DNA adducts was partly abrogated by sorafenib. In HB xenografts, both, sorafenib and alternated application of sorafenib and cisplatin significantly reduced tumour growth (P<0.05). Levels of AFP were lower in both treated groups (P=0.08). Relative apoptotic areas were increased (P=0.003). Mean vascular density was the lowest in the sorafenib/CDDP group (P=0.02).Conclusion:The combination of sorafenib with cisplatin might be a promising treatment option for high risk or recurrent HB.