Tsunemasa Nonogaki

Carbon tetrachloride-induced lethality in mouse is prevented by multiple pretreatment with zinc sulfate.

Carbon tetrachloride (CCl4) is commonly used as a chemical inducer of experimental liver injury. Several compounds have been demonstrated to attenuate the hepatic damage caused by sublethal doses of CCl4. However, rescue from lethal toxicity of CCl4 has not been reported. In the present study, we evaluated the protective effect of metallothionein (MT), an endogenous scavenger of free radicals, on CCl4-induced lethal toxicity of mice. To induce MT production in male ddY mice, we administered Zn (as ZnSO4) at 50 mg/kg as a once-daily subcutaneous injection for 3 days prior to a single intraperitoneal administration of 4 g/kg CCl4. Animals were observed for mortality every 3 hr for 24 hr after CCl4 injection. Liver damage was assessed by determining (in a subset of these mice) blood levels of alanine aminotransferase (ALT; a marker of liver injury) and liver histopathology at 6 hr after CCl4 injection. Our results showed that three times pretreatment with Zn yielded > 40-fold induction of hepatic MT protein levels compared to control group. Zn pretreatment completely abolished the CCl4-induced mortality of mice. We also found that pretreatment of mice with Zn significantly decreased the ALT levels and reduced the histological liver damage as assessed at 6 hr post-CCl4. These findings suggest that prophylaxis with Zn protects mice from CCl4-induced acute hepatic toxicity and mortality, presumably by induction of radical-scavenging MT.

Carbon Tetrachloride-Induced Nephrotoxicity in Mice Is Prevented by Pretreatment with Zinc Sulfate

Carbon tetrachloride (CCl4) is commonly used as a chemical inducer of experimental liver injury. In addition, many studies showed that CCl4 can induce kidney damage. In the current study, we evaluated the protective effect of zinc (Zn) against CCl4-induced nephrotoxicity. We hypothesized that this protective effect would result from the ability of Zn to serve as an inducer of metallothionein (MT), a known endogenous scavenger of free radicals. We administered Zn (as ZnSO4) 50 mg/kg subcutaneously once daily for 3 successive days prior to a single intraperitoneal administration of CCl4 4 g/kg in male ddY mice. Our results showed that Zn pretreatment significantly decreased creatinine and blood urea nitrogen levels and reduced renal histopathological damage at 6 h post-CCl4 injection, observations consistent with enhanced antioxidative activity in the kidney. Moreover, kidney MT levels in the Zn+CCl4-treated group decreased by greater than 70% compared with levels in the Zn-alone group, implying that MT was consumed by CCl4-induced radicals. These findings suggest that prophylaxis with Zn protects mice from CCl4-induced acute nephrotoxicity, presumably by induction of MT, which in turn scavenges radicals induced by CCl4 exposure.

Chronotoxicity of bromobenzene-induced hepatic injury in mice

The aim of the present study is to investigate whether or not bromobenzene (BB) toxicity varies with circadian periodicity. Seven-week-old male ICR mice were injected with 900 mg/kg (5.73 mmol/kg) BB intraperitoneally at 4 different time points of a day (zeitgeber time [ZT]: ZT0, ZT6, ZT12, and ZT18). Mortality was then monitored for 7 days after injection. Interestingly, mice were sensitive to BB acute toxicity at ZT6 while tolerant at ZT18. Moreover, in mice that were given a non-lethal dose of BB (540 mg (3.44 mmol)/kg), levels of alanine aminotransferase and aspartate aminotransferase, used as markers of hepatic injury, markedly increased in response to injection at ZT6, but did not increase significantly in response to injection at ZT18. In contrast, the markers of renal injury (creatinine and blood urea nitrogen), showed no significant difference in response to the two injection times. To further investigate this extreme circadian variation, we examined hepatic and renal lipid peroxidation levels, and conducted histopathological studies. Similar to our observation with alanine aminotransferase and creatinine, hepatic lipid peroxidation and histopathological changes were more pronounced than renal changes, and showed circadian variation. Our present investigation demonstrated that BB-induced mortality had clear circadian variation, and suggested that hepatic injury was one of the important factors for determination of this variation.

Calcium-deficient diet attenuates carbon tetrachloride-induced hepatotoxicity in mice through suppression of lipid peroxidation and inflammatory response

The aim of this study is to investigate whether a Ca-deficient diet has an attenuating effect on carbon tetrachloride (CCl4)-induced hepatotoxicity. Four-week-old male ddY mice were fed a Ca-deficient diet for 4 weeks as a part of the experimental protocol. While hypocalcemia was observed, there was no significant change in body weight. The CCl4-exposed hypocalcemic mice exhibited a significant decrease in alanine aminotransferase and aspartate aminotransferase activities at both 6 h and 24 h even though markers of renal function remained unchanged. Moreover, lipid peroxidation was impaired and total antioxidant power was partially recovered in the liver. Studies conducted in parallel with the biochemical analysis revealed that hepatic histopathological damage was attenuated 24 h post CCl4 injection in hypocalcemic mice fed the Ca-deficient diet. Finally, this diet impaired CCl4-induced inflammatory responses. Although upregulation of Ca concentration is a known indicator of terminal progression to cell death in the liver, these results suggest that Ca is also involved in other phases of CCl4-induced hepatotoxicity, via regulation of oxidative stress and inflammatory responses.

Sasa veitchii extract reduces obesity-induced insulin resistance and hepatic steatosis in obese mice fed a high-fat diet

ABSTRACT The aim of this study was to investigate the therapeutic effect of Sasa veitchii leaf extract (SE) on features of obesity induced by a high-fat diet (HFD), such as hyperglycemia, insulin resistance, and inflammatory response. Four-week-old male ddY mice were freely fed HFD or control normal diet for 12 weeks; half was given SE in addition twice per day in weeks 8–12. Glucose and insulin intolerance were estimated, and body weight measured, weekly throughout the study. Following the experiment, the mice were fasted for 16 h, euthanized, and plasma was collected. Liver and epididymal adipose tissue was collected and weighed. Treatment with SE significantly decreased body weight, adipose tissue weight, plasma glucose, insulin, leptin, and tumor necrosis factor α compared with HFD groups, and markedly reduced the impairment of glucose and insulin tolerance in obese mice. Furthermore, hepatic steatosis and hepatic insulin receptor substrate were improved by treatment with SE. Our findings demonstrate that SE may reduce obesity-induced glucose and insulin tolerance, not only by suppressing inflammatory responses but also by improving insulin signaling.

Potentiating effect of acetaminophen and carbon tetrachloride-induced hepatotoxicity is mediated by activation of receptor interaction protein in mice

Abstract When multiple drugs or chemicals are used in combination, it is important to understand the risk of their interactions and predict potential additive effects. The aim of the current study was to investigate the molecular mechanism(s) accounting for the additive/synergistic effect of combination treatment with acetaminophen (APAP) and carbon tetrachloride (CCl4). Mice were intraperitoneally administered vehicle or 100 mg/kg (5 mL/kg) APAP and 30 min after vehicle or 15 mg/kg (5 mL/kg) CCl4. Sixteen hours after treatment, mice from each group were sacrificed and the livers were removed. CCl4 administration caused slight glycogen depletion; this effect was more pronounced following co-administration of APAP and CCl4. ATP and NADPH levels showed the same trend as glycogen levels. The levels of receptor interacting protein 1 and 3 increased following combination treatment with APAP and CCl4. In contrast, levels of the glutamate cysteine ligase catalytic subunit and glutamate cysteine ligase modifier subunits were not significantly affected by combination treatment. APAP and CCl4 co-administration potentiated the phosphorylation of c-Jun N-terminal kinase and p38 kinases, although phosphorylated activation of extracellular signal-regulated kinase was not changed. Our results suggest that APAP and CCl4 co-administration potentiates hepatotoxicity in an additive/synergistic manner via receptor interacting protein activation.

Suppressive effect of kamebakaurin on acetaminophen-induced hepatotoxicity by inhibiting lipid peroxidation and inflammatory response in mice

BACKGROUND Kamebakaurin (KA) is an ent-kaurane diterpenoid known to have anti-inflammatory potential. In the current study, we investigated whether pretreatment with KA could ameliorate acetaminophen (APAP)-induced hepatotoxicity by inhibiting the anti-inflammatory response in mice. METHODS Seven-week-old C57BL/6J mice were orally administered KA or olive oil emulsion for seven days. Twenty-four hours after the last KA or olive oil administration, the mice were intraperitoneally injected with 400mg/kg APAP or saline under feed deprived condition. The mice from each group were euthanized and bled for plasma analysis 24h after the injection. RESULT APAP increased plasma levels of hepatic injury markers (i.e., alanine aminotransferase and aspartate aminotransferase), lipid peroxidation, and pro-inflammatory cytokines. Pretreatment with KA reduced the magnitude of APAP-induced increases in plasma levels of hepatic injury markers, lipid peroxidation, and inflammatory response. In addition, KA exhibited antioxidant capacity in a dose-dependent manner, with slight reactive oxygen species scavenging activity. CONCLUSION Our results indicate that KA has the ability to protect the liver from APAP-induced hepatotoxicity, presumably by both inhibiting the inflammatory response and oxidative stress.

Vitamin D3-induced hypercalcemia increases carbon tetrachloride-induced hepatotoxicity through elevated oxidative stress in mice

The aim of this study was to determine whether calcium potentiates acute carbon tetrachloride (CCl4) -induced toxicity. Elevated calcium levels were induced in mice by pre-treatment with cholecalciferol (vitamin D3; V.D3), a compound that has previously been shown to induce hypercalcemia in human and animal models. As seen previously, mice injected with CCl4 exhibited increased plasma levels of alanine aminotransferase, aspartate aminotransferase, and creatinine; transient body weight loss; and increased lipid peroxidation along with decreased total antioxidant power, glutathione, ATP, and NADPH. Pre-treatment of these animals with V.D3 caused further elevation of the values of these liver functional markers without altering kidney functional markers; continued weight loss; a lower lethal threshold dose of CCl4; and enhanced effects on lipid peroxidation and total antioxidant power. In contrast, exposure to V.D3 alone had no effect on plasma markers of liver or kidney damage or on total antioxidant power or lipid peroxidation. The potentiating effect of V.D3 was positively correlated with elevation of hepatic calcium levels. Furthermore, direct injection of CaCl2 also enhanced CCl4-induced hepatic injury. Since CaCl2 induced hypercalcemia transiently (within 3 h of injection), our results suggest that calcium enhances the CCl4-induced hepatotoxicity at an early stage via potentiation of oxidative stress.

Non-toxic Level of Acetaminophen Potentiates Carbon Tetrachloride-Induced Hepatotoxicity in Mice

A wide range of medications are routinely used to maintain and improve human health. Hence, it is essential that we understand and predict adverse effects caused by the combined use of multiple medications. In the present study, we investigated whether the combination of carbon tetrachloride (CCl4) and acetaminophen (APAP) had a detrimental effect on the liver. Mice injected with APAP (100 mg/kg) showed no significant changes in hepatic injury markers (alanine aminotransferase and aspartate aminotransferase), histopathological findings, pro-inflammatory cytokine levels, or hepatic oxidative stress. In contrast, a single injection of CCl4 (15 mg/kg) led to a significant increase in hepatic injury, in addition to an increase in pro-inflammatory cytokine levels and oxidative stress. Co-administration of APAP and CCl4 resulted in exacerbation of these hepatic injuries. Our results suggest that a non-toxic dose of APAP has the potential to increase CCl4-induced liver damage and oxidative stress.

Methyl dehydroabietate counters high fat diet-induced insulin resistance and hepatic steatosis by modulating peroxisome proliferator-activated receptor signaling in mice

The aim of this study was to investigate the therapeutic effects of methyl dehydroabietate (mDA) on adipocyte differentiation in 3T3-L1 preadipocytes and obesity characteristics induced by high-fat diet (HFD) in mice. Adipocyte differentiation in 3T3-L1 cells was evaluated after 14 days of incubation with mDA. mDA enhanced adipocyte differentiation in 3T3-L1 cells. For the in vivo evaluation, five-week-old male C57BL/6J mice were fed HFD or normal CE-2 diet (control) for eight weeks. During the experimental period, mice were administered mDA (50 mg/kg, p.o.) as an olive oil emulsion (containing 10% ethanol), and body weights were measured weekly. At the end of the experiment, the mice were euthanized after 16 h fasting period, and plasma samples were collected. The liver, kidney, and epididymal adipose tissues were collected and weighed. It significantly decreased body weight, adipose tissue weight, and plasma levels of glucose, insulin, leptin, and pro-inflammatory cytokines compared with that in the HFD group, and markedly reduced the impairment in glucose tolerance in obese mice. Furthermore, mDA reduced HFD-induced adipocyte hypertrophy and the formation of hepatic lipid droplets. Moreover, it induced the expression of proliferator-activated receptor alpha (PPARα) in the liver and PPARγ in the adipose tissues. Our findings demonstrate that mDA reduces obesity-induced glucose and insulin tolerance by inducing PPAR expression.