Contributions of caspase-8 and -9 to liver injury from CYP2E1-produced metabolites of halogenated hydrocarbons.

1. Drug-induced liver injury is difficult to predict at the pre-clinical stage. This study aimed to clarify the roles of caspase-8 and -9 in CYP2E1 metabolite-induced liver injury in both rats and cell cultures in vitro treated with carbon tetrachloride (CCl4), halothane or sevoflurane. The human hepatocarcinoma functional liver cell line was maintained in 3-dimensional culture alone or in co-culture with human acute monocytic leukemia cells. 2. In vivo, laboratory indices of liver dysfunction and histology were normal after administration of sevoflurane. CCl4 treatment increased blood AST/ALT levels, liver caspase-3 and -9 activities and liver malondialdehyde, accompanied by centrilobular hepatocyte necrosis. Halothane increased AST/ALT levels, caspase-3 and -8 activities (but not malondialdehyde) concomitant with widespread hepatotoxicity. In vitro, CCl4 treatment increased caspase-9 activity and decreased both mitochondrial membrane potential (MMP) and cell viability. In co-culture, halothane increased caspase-8 activity and decreased MMP and cellular viability. There were no toxic responses in CYP2E1 knockdown in monoculture and co-culture. 3. CYP2E1-inducing compounds play a pivotal role in halogenated hydrocarbon toxicity. 4. Changes in hepatocyte caspase-8 and -9 activities could be novel biomarkers of metabolites causing DILI, and in pre-clinical development of new pharmaceuticals can predict nascent DILI in the clinical stage.

The first case of human protothecosis caused by Prototheca zopfii in Japan.

This report describes a fatal case Prototheca zopfii genotype 2 infection in an immunosuppressed patient. The patient was a 62-year-old housewife who presented general malaise in April 2011. Hairy cell leukemia was highly suspected. Chemotherapy was started because the patient developed severe pancytopenia in October 2011. Itraconazole capsules (100 mg/day) and trimethoprim (320 mg/day) plus sulfamethoxazole (1600 mg/day) combinations were orally administered for prophylaxis of fungal infections. Of BacT/ALERT 3D FA aerobic culture bottles and FN anaerobic culture bottles, only FA aerobic blood culture bottles produced positive reactions when the patient developed fever in January 2012. Gram-staining of blood culture bottles revealed Gram-negative elliptical sporangia. Culturing on Sabouraud dextrose agar produced smooth and creamy white, yeast-like colonies. Partial DNA sequences of the nuclear 18S rDNA and 28S rDNA D1/D2 domains of the isolated strain were identical to those of P. zopfii genotype 2. The MICs and minimal lethal concentrations of antifungals revealed that it was susceptible to amphotericin B and itraconazole. The patient died, at which time plasma (1 â†’ 3)-ß-D-glucan was positive (131 pg/mL).

Chronological changes in circulating levels of soluble tumor necrosis factor receptors 1 and 2 in rats with carbon tetrachloride-induced liver injury.

Carbon tetrachloride (CCl4) facilitates the generation of hepatotoxins that can result in morphologic abnormalities, and these abnormalities are reasonably characteristic and reproducible for each particular toxin. It is also known that tumor necrosis factor-alpha (TNF-α) may participate in CCl4-induced liver injury (CILI). In this study, we observed the chronological changes in circulating soluble tumor necrosis factor receptors 1 and 2 (sTNF-R1 and -R2) in rats with CILI. Laboratory data; circulating levels of TNF-α, sTNF-R1, and sTNF-R2; and TNF-α levels in liver tissues were measured at various time-points. In the CCl4 group, the plasma aspartate aminotransferase (AST, 7694±3041IU/l)/alanine aminotransferase (ALT, 3241±2159 IU/l) levels peaked at 48 h after CCl4 administration, but the other laboratory data did not differ significantly from the corresponding data in the controls. Centrilobular hepatocyte necrosis and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells near the central vein area were observed via hematoxylin eosin (HE) and TUNEL staining, respectively, at 24 and 48 h after CCl4 administration. Compared to the control group, the CCl4 group did not show significantly the increased circulating TNF-α levels. But TNF-α levels in the liver tissues first peaked at 1h (5261±2253 pg/g liver), and a second peak was observed at 12h (3806±533 pg/g liver) after CCl4 administration. Compared to the control group, the CCl4 group showed significantly increased circulating levels of both sTNF-R1 (797±121pg/ml) and sTNF-R2 (5696±626 pg/ml) 1h after CCl4 administration. Since the hepatocyte apoptosis may be resulted from binding of TNF-α with TNF-R1 at 24h after administration, and consequently the circulating TNF-R2 level might be approximately 10-fold higher than the circulating TNF-R1 level. In conclusion, increased circulating levels of sTNF-R1 and -R2 potentially contribute to drug-induced liver injury, together with AST/ALT.