Targeting Nrf-2 is a promising intervention approach for the prevention of ethanol-induced liver disease.

Alcoholic liver disease (ALD) remains to be a worldwide health problem. It is generally accepted that oxidative stress plays critical roles in the pathogenesis of ALD, and antioxidant therapy represents a logical strategy for the prevention and treatment of ALD. Nuclear factor erythroid-derived 2-like 2 (NFE2L2 or Nrf-2) is essential for the antioxidant responsive element (ARE)-mediated induction of endogenous antioxidant enzymes such as heme oxygenase 1 (HO-1) and glutamate-cysteine ligase [GCL, the rate-limiting enzyme in the synthesis of glutathione (GSH)]. Activation of Nrf-2 pathway by genetic manipulation or pharmacological agents has been demonstrated to provide protection against ALD, which suggests that targeting Nrf-2 may be a promising approach for the prevention and treatment of ALD. Herein, we review the relevant literature about the potential hepatoprotective roles of Nrf-2 activation against ALD.

Impairment of Akt activity by CYP2E1 mediated oxidative stress is involved in chronic ethanol-induced fatty liver.

Protein kinase B (PKB/Akt) plays important roles in the regulation of lipid homeostasis, and impairment of Akt activity has been demonstrated to be involved in the development of non-alcoholic fatty liver disease (NAFLD). Previous studies suggest that cytochrome P4502E1 (CYP2E1) plays causal roles in the pathogenesis of alcoholic fatty liver (AFL). We hypothesized that Akt activity might be impaired due to CYP2E1-induced oxidative stress in chronic ethanol-induced hepatic steatosis. In this study, we found that chronic ethanol-induced hepatic steatosis was accompanied with reduced phosphorylation of Akt at Thr308 in mice liver. Chronic ethanol exposure had no effects on the protein levels of phosphatidylinositol 3 kinase (PI3K) and phosphatase and tensin homologue deleted on chromosome ten (PTEN), and led to a slight decrease of phosphoinositide-dependent protein kinase 1 (PDK-1) protein level. Ethanol exposure resulted in increased levels of malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE)-Akt adducts, which was significantly inhibited by chlormethiazole (CMZ), an efficient CYP2E1 inhibitor. Interestingly, N-acetyl-L-cysteine (NAC) significantly attenuated chronic ethanol-induced hepatic fat accumulation and the decline of Akt phosphorylation at Thr308. In the in vitro studies, Akt phosphorylation was suppressed in CYP2E1-expressing HepG2 (CYP2E1-HepG2) cells compared with the negative control HepG2 (NC-HepG2) cells, and 4-HNE treatment led to significant decrease of Akt phosphorylation at Thr308 in wild type HepG2 cells. Lastly, pharmacological activation of Akt by insulin-like growth factor-1 (IGF-1) significantly alleviated chronic ethanol-induced fatty liver in mice. Collectively, these results indicate that CYP2E1-induced oxidative stress may be responsible for ethanol-induced suppression of Akt phosphorylation and pharmacological modulation of Akt in liver may be an effective strategy for the treatment of ethanol-induced fatty liver.

Critical Roles of Kupffer Cells in the Pathogenesis of Alcoholic Liver Disease: From Basic Science to Clinical Trials.

Alcoholic liver disease (ALD) encompasses a spectrum of liver injury ranging from steatosis to steatohepatitis, fibrosis, and finally cirrhosis. Accumulating evidences have demonstrated that Kupffer cells (KCs) play critical roles in the pathogenesis of both chronic and acute ALD. It has become clear that alcohol exposure can result in increased hepatic translocation of gut-sourced endotoxin/lipopolysaccharide, which is a strong M1 polarization inducer of KCs. The activated KCs then produce a large amount of reactive oxygen species (ROS), pro-inflammatory cytokines, and chemokines, which finally lead to liver injury. The critical roles of KCs and related inflammatory cascade in the pathogenesis of ALD make it a promising target in pharmaceutical drug developments for ALD treatment. Several drugs (such as rifaximin, pentoxifylline, and infliximab) have been evaluated or are under evaluation for ALD treatment in randomized clinical trials. Furthermore, screening pharmacological regulators for KCs toward M2 polarization may provide additional therapeutic agents. The combination of these potentially therapeutic drugs with hepatoprotective agents (such as zinc, melatonin, and silymarin) may bring encouraging results.

Associations between the tumor necrosis factor-α gene and interleukin-10 gene polymorphisms and risk of alcoholic liver disease: A meta-analysis.

BACKGROUND: The critical roles of tumor necrosis factor-α (TNF-α) and interleukin-10 (IL-10) in the pathogenesis of alcoholic liver diseases (ALD) suggest that functional variations in the TNF-α (TNFA) and IL-10 genes may be related to individual susceptibility to ALD. As available studies examining the associations between TNFA or IL-10 polymorphisms and ALD risk have yielded conflicting results, a meta-analysis was conducted to clarify the potential relation between TNFA and IL-10 polymorphisms and the risk of ALD. METHODS: A comprehensive literature search was conducted to identify relevant studies. Pooled odds ratios and 95% confidence intervals were calculated using a random-effects model. The heterogeneity between studies was assessed using the Cochran's Q statistic and the I(2) statistic. Publication bias was assessed using funnel plots and the Egger's regression test. RESULTS: A total of 17studies and 12studies were identified and included in the meta-analysis of the associations between TNFA polymorphisms and ALD risk, and IL-10 polymorphisms and ALD risk, respectively. The pooled results showed that the "A" allele of the TNFA-238G>A polymorphism was significantly associated with an increased risk of ALD. Significant differences in the allele and genotype distributions of the IL-10-1082A>G polymorphism were detected in the comparison between ALD patients and healthy controls, but not when comparing ALD patients and alcohol dependent individuals without ALD. No significant associations between other polymorphic loci and ALD risks were detected. CONCLUSIONS: The TNFA-238G>A polymorphism was significantly associated with ALD risk, while the TNFA-308G>A polymorphism and IL-10 polymorphisms (-1082A>G and -592C>A) may not be associated with the individual susceptibility to ALD. The impact of combined TNFA and IL-10 polymorphisms on individual susceptibility to ALD needs to be investigated in future studies.

Garlic Oil Suppressed Nitrosodiethylamine-Induced Hepatocarcinoma in Rats by Inhibiting PI3K-AKT-NF-κB Pathway.

To explore the underlying mechanisms for the protective effects of garlic oil (GO) against nitrosodiethylamine (NDEA)-induced hepatocarcinoma, 60 male Wistar rats were randomized into 4 groups (n=15): control group, NDEA group, and two GO plus NDEA groups. The rats in GO plus NDEA groups were pretreated with GO (20 or 40 mg/kg) for 7 days. Then, all rats except those in control group were gavaged with NDEA for 20 weeks, and the rats in GO plus NDEA groups were continuously administered with GO. The results showed that GO co-treatment significantly suppressed the NDEA-induced increases of alpha fetal protein (AFP) level in serum, nuclear atypia in H&E staining, sirius red-positive areas and proliferating cell nuclear antigen (PCNA) expression. The molecular mechanisms exploration revealed that the protein levels of phosphatidylinositol 3 kinase (PI3K)-p85, PI3K-p110, total AKT, p-AKT (Ser473) and p-AKT (Thr308) in the liver of NDEA group rats were higher than those in control group rats. In addition, NDEA treatment induced IκB degradation and NF-κB p65 phosphorylation, and up-regulated the protein levels of downstream pro-inflammatory mediators. GO co-treatment significantly reversed all the above adverse effects induced by NDEA. These results suggested that the protective effects of GO against NDEA-induced hepatocarcinoma might be associated with the suppression of PI3K- AKT-NF-κB pathway.

Inhibition of cytochrome P4502E1 by chlormethiazole attenuated acute ethanol-induced fatty liver.

Cytochrome P4502E1 (CYP2E1) has been demonstrated to play crucial roles in chronic ethanol-induced fatty liver, while its role in acute ethanol-induced fatty liver remains unclear. The current study was designed to evaluate the effects of chlormethiazole (CMZ), a specific inhibitor of CYP2E1, on acute ethanol-induced fatty liver, and to explore the mechanisms. Mice were pretreated with single dose of CMZ (50mg/kg body weight) by intraperitoneal injection or equal volume of saline, and then exposed to three doses of ethanol (5g/kg body weight, 25%, w/v) by gavage with 12h intervals. The mice were sacrificed at 4h after the last ethanol dosing. It was found that CMZ significantly attenuated acute ethanol-induced increase of the hepatic and serum triglyceride levels, and reduced fat droplets accumulation in mice liver. Acute ethanol-induced increase of the hepatic malondialdehyde (MDA) and 4-hydroxynonenal (4-HNE) levels (two biomarkers for oxidative stress) and decrease of glutathione (GSH) level was significantly suppressed by CMZ. CMZ also suppressed ethanol-induced decline of serum adiponectin level, but did not significantly affect the serum tumor necrosis factor-α (TNF-α) and ethanol levels. Furthermore, a significant decline of p62 protein level was observed in CMZ/ethanol group mice liver compared with that of the ethanol group mice. However, acute ethanol-induced increase of peroxisome proliferator-activated receptor α (PPAR-α) protein level was suppressed by CMZ, while the protein levels of sterol regulatory element-binding protein-1c (SREBP-1) and diacylglycerol acyltransferase 2 (DGAT2) were not significantly affected by ethanol or CMZ. Collectively, the results of the current study demonstrated that CMZ could effectively attenuate acute ethanol-induced fatty liver possibly by suppressing oxidative stress and adiponectin decline, and activating autophagy, which suggest that CYP2E1 might also play important roles in acute ethanol-induced fatty liver.

CMZ reversed chronic ethanol-induced disturbance of PPAR-α possibly by suppressing oxidative stress and PGC-1α acetylation, and activating the MAPK and GSK3ß pathway.

BACKGROUND: Cytochrome P4502E1 (CYP2E1) has been suggested to play critical roles in the pathogenesis of alcoholic fatty liver (AFL), but the underlying mechanisms remains unclear. The current study was designed to evaluate whether CYP2E1 suppression by chlormethiazole (CMZ) could suppress AFL in mice, and to explore the underlying mechanisms. METHODS: Mice were treated with or without CMZ (50 mg/kg bw, i.p.) and subjected to liquid diet with or without ethanol (5%, w/v) for 4 weeks. Biochemical parameters were measured using commercial kits. The protein and mRNA levels were detected by western blot and qPCR, respectively. Histopathology and immunohistochemical assay were performed with routine methods. RESULTS: CYP2E1 inhibition by CMZ completely blocked AFL in mice, shown as the decline of the hepatic and serum triglyceride levels, and the fewer fat droplets in the liver sections. Chronic ethanol exposure led to significant decrease of the mRNA and protein levels of peroxisome proliferator-activated receptor α (PPAR-α), which was blocked by CMZ co-treatment. CMZ co-treatment suppressed ethanol-induced oxidative stress, overproduction of tumor necrosis α (TNF-α), and decrease of protein levels of the PPAR-α co-activators including p300 and deacetylated PGC1-α. Furthermore, CMZ co-treatment led to the activation of AMP-activated protein kinase (AMPK), mitogen-activated protein kinase (MAPK), and PI3K/Akt/GSK3ß pathway. However, chronic ethanol-induced decline of acyl-CoA carboxylase (ACC) and fatty acid synthase (FAS) protein levels was partially restored by CMZ, while the activation of autophagy appeared to be suppressed by CMZ. CONCLUSION: These results suggested that CMZ suppressed chronic ethanol-induced oxidative stress, TNF-α overproduction, decline of p300 protein level and deacetylation of PGC1-α, and activated AMPK, MAPK, and PI3K/Akt/GSK3ß pathway, which might contribute to the activation of PPAR-α and account for the protection of CMZ against AFL.

Pentoxifylline for the treatment of nonalcoholic fatty liver disease: a meta-analysis of randomized double-blind, placebo-controlled studies.

AIMS: Pentoxifylline has been used to treat nonalcoholic fatty liver diseases (NAFLDs) due to its anti-tumor necrosis factor-α effects. We conducted a meta-analysis of randomized, double-blinded, placebo-controlled trials to investigate the effect of pentoxifylline on the biochemical and histological parameters of NAFLD patients. MATERIALS AND METHODS: A comprehensive literature search was conducted in the database including PubMed, Embase, ISI web of knowledge, the Cochrane Library, and Google Scholar to identify randomized, double-blind, placebo-controlled clinical trials about the effects of pentoxifylline on NAFLD. The pooled weighted mean difference (WMD) with 95% confidence interval (CI) was calculated to compare the effects of pentoxifylline and placebo. RESULTS: Five well-designed studies were retrieved. Pooled results showed that pentoxifylline significantly reduced the serum alanine transaminase activity (WMD=-27.97; 95% CI: -42.59, -13.34) and aspartate transaminase activity (WMD=-13.97; 95% CI: -23.31, -4.63) in NAFLD patients compared with placebo. In addition, pentoxifylline significantly improved steatosis (WMD=-0.68; 95% CI: -1.01, -0.34), lobular inflammation (WMD=-0.49; 95% CI: -0.86, -0.12), and fibrosis (WMD=-0.60; 95% CI: -0.99, -0.21). Furthermore, pentoxifylline also led to significant reduction in BMI (WMD=-0.51; 95% CI: -0.96, -0.06) and fasting glucose (WMD=-8.97; 95% CI: -14.52, -3.42), but did not significantly affect the serum tumor necrosis factor α and adiponectin levels when compared with placebo. CONCLUSION: Pentoxifylline could reduce the aminotransferase activities and improve the histological parameters in NAFLD patients. Large well-designed, randomized, placebo-controlled studies are needed to confirm these results.

[Effect of docosahexaenoic acid and nervonic acid on the damage of learning and memory abilities in rats induced by 1-bromopropane].

OBJECTIVE: To investigate the protective effects of docosahexaenoic acid (DHA) and nervonic acid (NA) on the learning and memory abilities in rats exposed to 1-bromopropane (1-BP) and their action mechanisms. METHODS: Forty male Wistar rats (specific pathogen-free) were randomly divided into 4 groups (n = 10 for each), i.e., solvent control group, 1-BP (800 mg/kg) group, NA (150 mg/kg) + 1-BP (800 mg/kg) group, and DHA (500 mg/kg) + 1-BP (800 mg/kg) group. The rats were given respective test substances by gavage for 7 d. The Morris water maze (MWM) test was performed from days 8 to 12 to evaluate the rats' learning and memory abilities. After MWM test, rats were sacrificed in the next day, and cerebral cortex was quickly dissected and homogenized in an ice bath. The supernatant of the obtained homogenate was collected to measure the content of glutathione (GSH) and malondialdehyde (MDA) and the activities of glutathione reductase (GR) and γ-glutamate cysteine ligase (γ-GCL). RESULTS: The MWM spatial navigation test showed that the 1-BP group had significantly longer escape latency and significantly longer total swimming distance compared with the control group (P<0.05), while the DHA+1-BP group had significant decreases in escape latency and total swimming distance compared with the 1-BP group (P<0.05). The spatial probe test showed that the number of platform crossings was significantly greater in the DHA+1-BP group and NA+1-BP group than in the 1-BP group (P<0.05); compared with the control group, the 1-BP group had a significantly lower ratio of time spent in the zone around the platform to total time (P < 0.05), and the ratio was significantly higher in the DHA+1-BP group than in the 1-BP group (P < 0.05). Compared with the control group, the 1-BP group had a 18.1% decrease in GSH content, and DHA could significantly reverse 1-BP-induced decrease in GSH content (P < 0.05). Compared with the 1-BP group, the DHA+1-BP group and NA+1-BP group had significantly decreased MDA content (P < 0.05), the DHA+1-BP group had significantly increased GR activity (P < 0.05), and the NA+1-BP group had significantly increased γ-GCL activity (P < 0.05). CONCLUSION: The rats exposed to 1-BP have oxidative stress in the brain and impaired cognitive function. DHA and NA can reduce 1-BP-induced cognitive function impairment in rats, possibly by increasing the activities of GR and γ-GCL and the content of GSH in the brain.

Garlic oil suppressed the hematological disorders induced by chemotherapy and radiotherapy in tumor-bearing mice.

Although the anticancer effects of garlic and its products have been demonstrated by a variety of studies; however, few studies were conducted to investigate the effects of garlic on the adverse effects of chemo/radiotherapy. In order to clarify the above question and make a more comprehensive understanding of the anticancer effects of garlic, tumor xenograft mice model was established by subcutaneous injection of H22 tumor cells, and was used for the investigation of effects of garlic oil (GO) on the chemo/radiotherapy. In the chemotherapy test, tumor-bearing mice were treated with cyclophosphamide (CTX) or CTX plus GO (25 or 50 mg/kg bw) for 14 d, while the mice received a single 5 Gy total body radiation or radiation plus GO (25 or 50 mg/kg bw) in radiotherapy test. The results showed that GO did not increase the tumor inhibitory rate of CTX/radiation, which indicated that GO could not enhance the chemo/radiosensitivity of cancer cells. However, the decrease of the peripheral total white blood cells (WBCs) count induced by CTX/radiation was significantly suppressed by GO cotreatment. Furthermore, GO cotreatment significantly inhibited the decrease of the DNA contents and the micronuclei ratio of the bone marrow. Lastly, the reduction of the endogenous spleen colonies induced by CTX/radiation was significantly suppressed by GO cotreatment. These findings support the idea that GO consumption may benefit for the cancer patients receiving chemotherapy or radiotherapy.

Garlic oil attenuated nitrosodiethylamine-induced hepatocarcinogenesis by modulating the metabolic activation and detoxification enzymes.

Nitrosodiethylamine (NDEA) is a potent carcinogen widely existing in the environment. Our previous study has demonstrated that garlic oil (GO) could prevent NDEA-induced hepatocarcinogenesis in rats, but the underlying mechanisms are not fully understood. It has been well documented that the metabolic activation may play important roles in NDEA-induced hepatocarcinogenesis. Therefore, we designed the current study to explore the potential mechanisms by investigating the changes of hepatic phase Ⅰ enzymes (including cytochrome P450 enzyme (CYP) 2E1, CYP1A2 and CYP1A1) and phase Ⅱ enzymes (including glutathione S transferases (GSTs) and UDP- Glucuronosyltransferases (UGTs)) by using enzymatic methods, real-time PCR, and western blotting analysis. We found that NDEA treatment resulted in significant decreases of the activities of CYP2E1, CYP1A2, GST alpha, GST mu, UGTs and increases of the activities of CYP1A1 and GST pi. Furthermore, the mRNA and protein levels of CYP2E1, CYP1A2, GST alpha, GST mu and UGT1A6 in the liver of NDEA-treated rats were significantly decreased compared with those of the control group rats, while the mRNA and protein levels of CYP1A1 and GST pi were dramatically increased. Interestingly, all these adverse effects induced by NDEA were simultaneously and significantly suppressed by GO co-treatment. These data suggest that the protective effects of GO against NDEA-induced hepatocarcinogenesis might be, at least partially, attributed to the modulation of phase I and phase II enzymes.

Protective effects of garlic oil on hepatocarcinoma induced by N-nitrosodiethylamine in rats.

To investigate the protective effects and the possible mechanisms of garlic oil (GO) against N-nitrosodiethylamine (NDEA)-induced hepatocarcinoma in rats, Wistar rats were gavaged with GO (20 or 40 mg/kg) for 1 week, and then were gavaged with GO and NDEA (10 mg/kg) for the next 20 weeks. The changes of morphology, histology, the biochemical indices of serum, and DNA oxidative damage of liver were examined to assess the protective effects. Lipid peroxidation (LPO), antioxidant defense system, and apoptosis-related proteins were measured to investigate potential mechanisms. At the end of the study (21 weeks), GO administration significantly inhibited the increase of the nodule incidence and average nodule number per nodule-bearing liver induced by NDEA, improved hepatocellular architecture, and dramatically inhibited NDEA-induced elevation of serum biochemical indices (alanine aminotransferase , aspartate aminotransferase, alkaline phosphatase and gamma-glutamyl transpeptidase) and hepatic 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels in a dose-dependent manner. The mechanistic studies demonstrated that GO counteracted NDEA-induced oxidative stress in rats illustrated by the restoration of glutathione (GSH), superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx), glutathione-S-transferase (GST) levels, and the reduction of the malondialdehyde (MDA) levels in liver. Furthermore, the mRNA and protein levels of Bcl-2, Bcl-xl, andß-arrestin-2 were significantly decreased whereas those of Bax and caspase-3 were significantly increased. These data suggest that GO exhibited significant protection against NDEA-induced hepatocarcinogenesis, which might be related with the enhancement of the antioxidant activity and the induction of apoptosis.

PI3K/Akt pathway activation was involved in acute ethanol-induced fatty liver in mice.

Accumulating evidences support the important roles of sterol regulatory element-binding protein-1 (SREBP-1) activation in ethanol-induced fatty liver, but the underlying mechanisms for its activation are not fully understood. Recent studies have demonstrated that phosphatidylinositol 3 kinase (PI3K)/Akt pathway activation could enhance SREBP-1 activity. The current study was designed to investigate the potential roles of PI3K/Akt pathway in acute ethanol-induced fatty liver in mice. In the first experiment, mice were treated with ethanol (2.5 or 5 g/kg bw) or isocaloric/isovolumetric maltose-dextrin solution, and sacrificed at several time points after ethanol exposure. As expected, ethanol dose-dependently increased the hepatic triglyceride (TG) levels and the protein levels of the mature form of SREBP-1 (n-SREBP-1). The phosphorylation of Akt and glycogen synthase kinase-3ß (GSK-3ß) was significantly increased in mice treated with ethanol (5 g/kg bw), while the protein levels of PI3K-p85 were significantly reduced. To confirm the roles of PI3K/Akt pathway, mice were then pretreated with wortmannin (0.7 or 1.4 mg/kg bw), a specific PI3K/Akt pathway inhibitor, before exposure to ethanol. Interestingly, a dual effect of wortmannin was observed. Low dose of wortmannin significantly reduced the hepatic TG levels, while high dose of wortmannin aggravated ethanol-induced fatty liver. The ratio of LC3II/LC3I of wortmannin (1.4 mg/kg bw) group mice was significantly increased, while the p62 protein level was significantly decreased compared to those of ethanol group, which indicated that wortmannin (1.4 mg/kg bw) might suppress the lipid degradation by autophagy. These results supported the hypothesis that PI3K/Akt activation might be involved in acute ethanol-induced fatty liver, and PI3K/Akt inhibitors might have therapeutic potential for the treatment of ethanol-induced fatty liver.

Garlic oil alleviated ethanol-induced fat accumulation via modulation of SREBP-1, PPAR-α, and CYP2E1.

Garlic oil (GO) has been shown to partially attenuate ethanol-induced fatty liver, but the underlying mechanisms remain unclear. The current study was designed to evaluate the protective effects of GO against ethanol-induced steatosis in vitro and in vivo, and to explore potential mechanisms by investigating the sterol regulatory element binding protein-1c (SREBP-1c), peroxisome proliferators-activated receptor-α (PPAR-α), cytochrome P4502E1 (CYP2E1), and etc. In the in vitro study, human normal cell LO2 was exposed to 100 mM ethanol in the presence or absence of GO for 24 h. We found that ethanol increased the protein levels of n-SREBP-1c and CYP2E1, but decreased the protein levels of PPAR-α, which was significantly attenuated by GO co-treatment. In the in vivo study, male Kun-Ming mice were pretreated with single dose of GO (50-200 mg/kg body weight) at 2 h before ethanol (4.8 g/kg body weight) exposure. The changes of n-SREBP-1c, PPAR-α and CYP2E1 were paralleled well to those of in vitro study. Furthermore, GO significantly reduced the protein levels of fatty acid synthase (FAS), and suppressed ethanol-induced hepatic mitochondrial dysfunction. These results suggested that GO had the potential to ameliorate alcoholic steatosis which might be related to its modulation on SREBP-1c, PPAR-α, and CYP2E1.

[Effects of garlic oil, age and sex on n-hexane metabolism in rats].

OBJECTIVE: To investigate effects of garlic oil (GO), age and sex on n-hexane metabolism in rats. METHODS: The Wistar rats were used as experimental animals. (1) Intragastric administration: n-hexane group (3000 mg/kg n-hexane), GO treated group (80 mg/kg GO ig. an hour earlier than 3000 mg/kg n-hexane), then blood was taken from tails of rats at 8, 12, 16, 20, 24, 28, 32 h points after n-hexane administration. (2) Intraperitoneal injection: n-hexane group (1000 mg/kg n-hexane), GO treated group (80 mg/kg GO ig. an hour earlier than 1000 mg/kg n-hexane), then took blood was taken from tails of rats at 8, 12, 16, 20, 24, 28 h points after n-hexane injection. (3) 7 rats each group of 6, 8, 10 weeks age were administrated by 3000 mg/kg n-hexane intragastrically, then were taken blood from tails at 16, 20, 24 h points after administration. (4) 7 male and 7 female rats of 8 weeks age were administrated by 3000 mg/kg n-hexane intragastrically, then were taken blood from tails at 16, 20, 24, 28 h points after administration. The gas chromatography was used to determine the metabolite 2, 5-hexanedione concentration of n-hexane in serum and 2, 5-hexanedione concentration was compared between GO and no GO treated rats, different ages and different sexes. RESULTS: (1) Intragastric administration: 2, 5-hexanedione concentrations in serum of n-hexane group and GO treated group had the peak 19.2 and 12.3 µg/ml at 20h and 24 h points. Compared with n-hexane group, the serum 2, 5-hexanedione concentration of GO treated group was lower at time points prior to peak and 2, 5-hexanedione eliminating process was slower after peak. (2) Intraperitoneal injection: effects of GO on the serum 2, 5-hexanedione concentrations was very similar to intragastric administration, 2, 5-hexanedione concentrations in serum of n-hexane group and GO treated group had the peak 15.0 and 6.7 µg/ml at 12 h and 16 h points. (3) Comparison of the serum 2, 5-hexanedione concentrations of different weeks age rats: The serum 2, 5-hexanedione concentrations of 6, 8, 10 weeks age rats were 25.5, 15.0, 12.8 µg/ml each (8, 10 weeks age significantly lower than 6 weeks age) at 16 h point; at 20 h point, they were 24.7, 18.3, 15.0 µg/ml each (10 weeks age significantly lower than 6 weeks age); at 24 h point, they were 11.0, 14.7, 8.1 µg/ml each (10 weeks age significantly lower than 8 weeks age). (4) Comparisons of the serum 2, 5-hexanedione concentrations of different sex rats: the serum 2, 5-hexanedione concentrations of male and female rats were 22.5, 17.2 µg/ml each at 16 h point (different significantly); at 20, 24, 28 h points, they were 27.6, 22.9 µg/ml, 24.6, 19.1 µg/ml, 19.1, 13.8 µg/ml each (different non-significantly). CONCLUSION: GO reduces production of 2, 5-hexanedione in serum generated by n-hexane in rats; the metabolic capacity of low age rats on n-hexane is stronger than high age ones.

[The effects of tea polyphenols on the injury of fibrinolytic functions induced by high-methionine dietary in rats].

OBJECTIVE: To study the protective impact of tea polyphenols (TP) on the injury of fibrinolytic functions induced by high-methionine dietary in rats. METHODS: 50 male Wistar rats were divided by stratified based on body weight into 5 groups with 10 in each group: namely control group, model group, low-dose TP group, medium-dose TP group and high-dose TP group. The rats in model group and TP groups were fed with 3% methionine dietary, control group rats with routine diet. In addition, rats in low-dose, medium-dose and high-dose TP groups were treated with TP at 50, 100 and 200 mg/kg dosage respectively by gavages every day, control group and model group rats were given with same amount distilled water. The animals were sacrificed after 8 weeks. The levels of tissue-type plasminogen activator (t-PA) and type-1 plasminogen activator inhibitor (PAI-1) in plasma were determined by ELISA assays, mRNA levels of t-PA and PAI-1 in aortic arch were detected by RT-PCR, t-PA and PAI-1 expression in aortic arch were detected by immunohistochemistry strept-avidin-biotin complex (SABC). RESULTS: After experiment, the t-PA expression of aortic arch in control group, model group, low-dose TP group, medium-dose TP group and high-dose TP group were 133.03 ± 10.14, 95.46 ± 11.08, 111.97 ± 11.91, 130.23 ± 10.80, 139.39 ± 9.41 (F = 14.15, P < 0.01), respectively, and the PAI-1 expression were 90.91 ± 8.67, 166.76 ± 12.18, 139.63 ± 12.71, 134.66 ± 13.19, 109.49 ± 10.82 (F = 31.44, P < 0.01). The t-PA concentration of plasma were (10.69 ± 1.26), (6.13 ± 0.92), (8.56 ± 1.19), (9.69 ± 0.92), (11.97 ± 1.08) ng/ml, respectively (F = 41.98, P < 0.01), and the PAI-1 concentration of plasma were (6.31 ± 0.81), (16.98 ± 1.27), (11.39 ± 0.82), (8.46 ± 0.67), (8.08 ± 0.91) ng/ml, respectively (F = 207.74, P < 0.01). The mRNA levels of t-PA in aortic arch were 1.12 ± 0.02, 0.75 ± 0.14, 1.01 ± 0.09, 0.95 ± 0.08, 1.05 ± 0.13 (F = 5.77, P < 0.05), and the mRNA levels of PAI-1 in aortic arch were 1.25 ± 0.11, 1.74 ± 0.06, 1.23 ± 0.05, 1.09 ± 0.14, 1.23 ± 0.04 (F = 23.56, P < 0.01). CONCLUSION: The results indicate that TP seems to have regulatory function on transcription and protein levels of t-PA and PAI-1, in addition to maintaining the balance between PAI-1 and t-PA and healing the injury of fibrinolytic functions in rats induced by high-methionine dietary.

[Protective effects of garlic oil on n-hexane-induced neurotoxicity in rats via inhibition of hepatic alcohol dehydrogenase activity].

OBJECTIVE: To study the protective effects of garlic oil (GO) on the peripheral nerve injuries induced by n-hexane. METHODS: Male Wistar rats were randomly divided into four groups (10 rats in each group): the control, the n-hexane treatment (2000 mg/kg), the low dose GO, and the high dose GO groups. The rats in the low and high doses of GO groups were pretreated with GO (40 and 80 mg/kg) before exposure to n-hexane (2000 mg/ kg), while the animals of the n-hexane treatment group were given normal saline and then 2000 mg/ kg n-hexane. The rats were exposed to GO and n-hexane 6 times a week for 10 weeks. The gait scores and staying time on the rotating rod for all rats were detected every two weeks. The rats were sacrificed at the end of ten weeks, then the levels of alcohol dehydrogenase (ADH), maleic dialdehyde (MDA), reduced glutathione (GSH), glutathione peroxidase(GSH-Px), total antioxidation capacity(T-AOC) and the ability of inhibition of *OH in livers were examined. RESULTS: The gait scores increased significantly and the time staying on the rotating rod obviously decreased in rats of n-hexane treatment group, as compared with control group (P < 0.05 or P < 0.01). In the hepatic tissues of n-hexane group, the levels of MDA and ADH significantly increased, the activities of GSH-Px, T-AOC and the ability of inhibition of *OH obviously decreased, as compared to control group (P < 0.05 or P < 0.01). In 2 GO groups, the gait scores and the staying time on the rotating rod were significantly improved, the levels of MDA and ADH significantly decreased, the activities of GSH-Px, T-AOC and the ability of inhibition of *OH obviously increased, as compared with n-hexane group (P < 0.05 or P < 0.01 ). CONCLUSION: ADH could play an important role in the protective effects induced by garlic oil on the peripheral nerve injuries produced by n-hexane.

[The role of CYP2E1 in the protection of garlic oil's from n-hexane-induced neurotoxicity].

OBJECTIVE: To study the role of CYP2E1 in the protective effects and mechanism of garlic oil (GO) on the peripheral nerve injuries induced by n-hexane. METHODS: Fifty male Wistar rats were randomly divided into five groups (n = 10): the control, the GO (80 mg/kg) control, the n-hexane (2000 mg/kg) model, the low dose GO (40 mg/kg) plus n-hexane, and the high dose GO (80 mg/kg) plus n-hexane groups. All rats were treated by intragastric administration 6 times a week for 10 weeks. The gait scores were determined every two weeks for monitoring the peripheral neurotrosis. All rats were sacrificed in 10 weeks, the activities and expression levels of hepatic CYP2E1 and 2, 5-HD in serum were examined. RESULTS: As compared with control group, the content and activity of hepatic CYP2E1 in GO control group reduced by 83.1% and 48.3% respectively (P < 0.01), the content and activity of hepatic CYP2E1 in model group increased by 112.5% and 72.2% respectively (P < 0.01). As compared with model group, the contents of hepatic CYP2E1 in low dose and high dose GO groups reduced by 32.9% and 39.1% respectively, the activities of hepatic CYP2E1 in low dose and high dose GO groups reduced by 27.4% and 44.5% respectively (P < 0.01); the contents of serum 2,5-HD in low dose and high dose GO groups reduced by 47.7% and 78.7% respectively (P < 0.01). The gait scores in model, low dose and high dose GO groups were significantly lower than that in control group, but the gait scores in low dose and high dose GO groups were significantly lower than that in model group (P < 0.05). CONCLUSION: Garlic oil can effectively reduce the peripheral neurotrosis induced by n-hexane due to the decreased content and activity of hepatic CYP2E1, resulting in the reduced formation of 2, 5-HD from n-hexane.

[Protective effect of garlic oil given at different time against acute liver injury induced by CCl4].

OBJECTIVE: To observe and compare the protective effect of garlic oil against carbon tetrachloride (CCL)-induced acute liver injury. METHODS: The experiments include 4 preventive groups and 2 therapeutic groups. In every preventive and therapeutic group, the mice were randomized into 6 groups with 15 each, including one negative control group, one solvent control group, one CCl4 model group and 3 garlic oil groups (25, 50, and 100 mg/kg body weight). Before given a single gavage of CCl4 (80 mg/kg), the mice were pretreated with garlic oil by gavage in preventive group 1 (30 days, once daily), preventive group 2 (5 days, once daily), preventive group 3 (ahead of 2 h, once), preventive group 4 (immediately, once) or the vehicle (corn oil, 10 ml/kg) in solvent control group. In therapeutic groups, the mice were gavaged garlic oil 2 h (once, in therapeutic 1) or for 5 days (once daily, in therapeutic 2) after administration CCl. After 24 h of the last administration, blood was collected and centrifuged at 2500 r/min at 4 degrees C for 10 min, and serum was removed to measure ALT and AST activities. The liver was dissected, weighed to calculate the liver coefficient (relative liver weight). At the same time, the liver samples were studied by histological examinations. RESULTS: Compared with negative group, the liver coefficient and the activities of ALT and AST in serum of model group were increased remarkably (P < 0.01). Compared with CCl model group, the liver coefficient and the activities of ALT and AST in serum were decreased significantly (P < 0.01) by garlic oil dose-dependently in each preventive group. Simultaneously, histological assessment showed that garlic oil effectively alleviated hepatocyte injuries induced by CCl4. Comparing the preventive effects of garlic oil in every group, it was better in preventive group 3 than others. However, all indexes and histological examinations in therapeutic group 1 did not show the difference with those of CCl4 model group. In therapeutic group 2, all indexes recovered after 5 d of CCl4 administration. CONCLUSIONS: Garlic oil can prevent acute liver injury induced by CCl4 and the effect is better in ahead of 2 h group than others.

[Effects of tacrolimus (FK506) on heat-shock proteins 70, Bcl-2 and Bax expression in nervous tissue of acrylamide-induced rat].

OBJECTIVE: To investigate the effects of tacrolimus (FK506) on behavioral function and heat-shock proteins (HSP70) expression in nervous tissues of acrylamide (ACR)-induced rats. METHODS: Totally 40 health Wistar rats were randomly divided into control group, model group, low and high doses of FK506 groups. All four groups were treated five times per week for four weeks. Gait score was measured every week. And rats were sacrificed on day 28, the cerebrum, spinal cord and sciatic nerve were dissected, and homogenized in ice bath, then the levels of HSP70 and Bcl-2, Bax were analyzed by western bloting. RESULTS: Compared with the ACR model group, the gait score in low and high doses of FK506 groups decreased by 30.1% and 47.7% respectively in the 4th week. In the cerebrum and sciatic nerve pellet, the level of HSP70 in the FK506 groups increased by 11.6%, 33.3% and 56.3%, 58.5% (P < 0.01), but no significant changes existed in spinal cord. The level of Bcl-2 in the sciatic nerve pellet increased by 39.1% (P < 0.01) but no significant changes existed in the cerebrum and spinal cord from low dose of FK506 group. And the level of Bax in the spinal cord pellet markedly increased by 46.8% but not in cerebrum and sciatic nerve pellet; Whereas in the tissues mentioned above, the levels of Bcl-2 were enhanced remarkably by 16.3%, 14.8% and 56.0% (P < 0.01) in the high dose of FK506 group. And the level of Bax in the cerebrum and spinal cord pellet markedly increased by 16.4% and 40.2% but not in sciatic nerve. The values of Bcl-2/Bax in low and high doses of FK506 groups clearly increased by 15.9%, 33.3%, 36.9% and 30.1%, 49.1%, 60.1% (P < 0.01). CONCLUSION: The administration of FK506 has dramatically neuroprotective effects against the development of ACR neuropathy, which may be related to up-regulating the expression of HSP70 and Bcl-2 with down-regulating the expression of Bax.