Protective Effects of Voluntary Exercise on Hepatic Fat Accumulation Induced by Dietary Restriction in Zucker Fatty Rats

Weight control based on dietary restriction (DR) alone can cause lipid metabolic failure and progression to fatty liver. This study aimed to investigate the effect of exercise on preventing DR-induced hepatic fat accumulation in Zucker fatty (ZF) rats by focusing on the relationship between adipose tissue lipolysis and hepatic fat uptake. Six-week-old male ZF rats were randomly assigned to obese, DR, or DR with exercise (DR + Ex) groups. The DR and DR + Ex groups were fed a restricted diet, with the latter also undergoing voluntary exercise. After 6 weeks, hepatic fat accumulation was observed in the DR group, whereas intrahepatic fat was markedly reduced in the DR + Ex group. Compared with the obese (Ob) group, the DR group exhibited 2.09-fold expression of hepatic fatty acid translocase (FAT)/CD36 proteins (p < 0.01) and 0.14-fold expression of hepatic fatty acid-binding protein (FABP)1 (p < 0.01). There were no significant differences between the DR + Ex group and the Ob group. FAT/CD36 and hepatic triglyceride (TG) expression levels were strongly positively correlated (r = 0.81, p < 0.001), whereas there was a strong negative correlation between FABP1 and hepatic TG expression levels (r = −0.65, p < 0.001). Our results suggest that hepatic fat accumulation induced by DR in ZF rats might be prevented through exercise-induced modifications in FAT/CD36 and FABP1 expression.     

Reaction of α-tocopherol in heated bulk phase in the presence of methyl linoleate (13S)-hydroperoxide or methyl linoleate

α-Tocopherol and methyl (9Z, 11E)-(S)-13-hydroperoxy-9, 11-octadecadienoate (13-MeLOOH) were allowed to stand at 100°C in bulk phase. The products were isolated and identified as methyl 13-hydroxyoctadecadienoate (1), stereoisomers of methyl 9,11,13-octadecatrienoate (2), methyl 13-oxo-9, 11-octadecadienoate (3), epoxy dimers of methyl linoleate with an ether bond (4), a mixture of methyl (E)-12, 13-epoxy-9-(α-tocopheroxy)-10-octadecenoates and methyl (E)-12, 13-epoxy-9-(α-tocopheroxy)-11-(α-tocopheroxy)-9-octadecenoates (5), a mixture of methyl 9-(α-tocopheroxy)-10,12-octadecadienoates and methyl 13-(α-tocopheroxy)-9, 11-octadecadienoates (6), α-tocopherol spirodiene dimer (7), and α-tocopherol trimer (8). α-Tocopherol and 13-MeLOOH were dissolved in methyl myristate, and the thermal decomposition rate and the distributions of reaction products formed from α-tocopherol and 13-MeLOOH were analyzed. α-Tocopherol disappeared during the first 20 min, and the main products of α-tocopherol were 5 and 6 with the accumulation of 1–4 which were the products of 13-MeLOOH. The results indicate that the alkyl and alkoxyl radicals from the thermal decomposition of 13-MeLOOH could be trapped by α-tocopherol to produce 5 and 6. The reaction products of α-tocopherol during the thermal oxidation of methyl linoleate were compounds 6 and 7. Since the radical flux during the autoxidation might be low, the excess α-tocopheroxyl radical reacted with each other to form 7.     

Bioaccumulation of organochlorines in crows from an indian open waste dumping site: evidence for direct transfer of dioxin-like congeners from the contaminated soil

To assess the significance of waste dumping sites as a source of chemical contamination to ecosystems, we analyzed the residue levels of polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), polychlorinated biphenyls (PCBs), and other organochlorines in the breast muscle of crows from a dumping site in the south of Chennai city, South India. Crows from the dumping site contained significantly higher total TEQs (60 +/- 27 pg/g lipid wt) than those from the reference sites (26 +/- 18 pg/g lipid wt). Especially, certain dioxin-like coplanar PCB congeners (Co-PCBs), such as CB-77 and CB-105, whose source is commercial PCBs,were significantly higher in crows from the dumping site than those from the reference sites. Profiles of PCDDs/DFs and Co-PCBs in crows from the dumping site were similar to those of soil at the same site, which was confirmed by principal component analysis. Furthermore, significant positive correlations were obtained between the congener-specific bioconcentration factors (BCFs) of PCDDs/DFs estimated from concentrations in crows and soil from the dumping site and the theoretical BCFs calculated from water-particle and lipid-water partitioning coefficients. On the other hand, the estimated BCFs had significant negative correlations with the molecular weight of PCDDs/DFs, indicating that molecular size limits their bioaccumulation. These results suggest that dioxin-like congeners in the soil of the dumping site were transferred directly to the crows through the ingestion of on-site garbage contaminated with soil, rather than through trophic transfer in the ecosystem. The present study provides insight into the ecological impacts of dumping sites.     

Effects of micafungin on the morphology of Aspergillus fumigatus

The morphological effects of micafungin, a member of a new class of antifungal agents candins, on growing hyphae of Aspergillus fumigatus were studied by the use of Nomarski microscopy, scanning electron microscopy and thin-section electron microscopy. Micafungin at concentrations of 0.001-0.1 microg ml(-1) strongly inhibited the in vitro growth of this fungus and induced striking changes in the hyphal morphology, depending on the drug concentration and the length of the incubation period. The changes observed included increased formation of branches on the lateral walls, disruption of the tips of both hyphal cells and branches, and crushing and collapse of whole hyphae. In addition, micafungin was also effective in damaging membranous structures, including disruption of the cell membrane, partial loss of nuclear membranes and expansion of endoplasmic reticula. From these results, it was concluded that micafungin primarily affected the normal formation of cell walls and septa of growing hyphae. The inhibition of apical growth of hyphae accompanied by excessive lateral branching that was followed by disruption of both hyphal tips and branch tips ultimately led to the destruction of whole hyphae. This cytological effect of micafungin on the hyphal growth of A. fumigatus can explain its potent anti-Aspergillus activity.     

Environmental fate of bisphenol A and its biological metabolites in river water and their xeno-estrogenic activity

Monitoring of bisphenol A [BPA; 2,2-bis(4-hydroxyphenyl)propane] and its biological metabolites [4,4'-dihydroxy-alphamethylstilbene (DHMS), 2,2-bis(4-hydroxyphenyl)-1-propanol (BPA-OH), 2,2-bis(4-hydroxyphenyl)propanoic acid (BPA-COOH), and 2-(3,4-dihydroxyphenyl)-2-(4-hydroxyphenyl)propane (3-OH-BPA)] in river waters was performed by solid-phase extraction and GC/MS determination. The concentrations of BPA, BPA-COOH, BPA-OH, and 3-OH-BPA in the river water ranged from 2 to 230 (8.8 x 10(-12) to 1.0 x 10(-9) M), from 5 to 75 (1.9 x 10(-11) to 2.9 x 10(-10) M), from 3 to 16 (1.2 x 10(-11) to 6.6 x 10(-11) M), and from 3 to 11 (1.2 x 10(-11) to 4.5 x 10(-11) M) ng L(-1), respectively. DHMS, an intermediate in the main degradation pathway of BPA, was not detected in any water sample. Under the aerobic conditions in the river water, BPA disappeared within 8 d of incubation, but BPA-COOH, BPA-OH, and tetraol remained in the supernatant after 14 d of incubation. For the xeno-estrogenic activity of BPA and the metabolites, their ability to bind to recombinant human estrogen receptor alpha in competition with fluorescence-labeled 17beta-estradiol was measured. Fifty percent inhibitory concentrations (IC50) of BPA, DHMS, 3-OH-BPA, and BPA-OH were approximately 1 x 10(-5), 1 x 10(-6), 3 x 10(-5), and 1 x 10(-2) M, respectively. In human cultured MCF-7 breast cancer cells, BPA increased cell numbers in a dose-dependent manner at concentrations from 10(-7) to 10(-5) M. For the BPA metabolites, DOHMS, 3-OH-BPA, and BPA-COOH caused the cells proliferation at concentrations from 10(-9) to 10(-6), from 10(-7) to 10(-6), and from 10(-5) to 10(-4) M, respectively. BPA-OH did not cause MCF-7 cells proliferation. These results indicate that BPA is mainly metabolized through oxidative rearrangement by bacteria in the river water, and intermediate bisphenols via minor metabolic pathways exist in river water. The presence of the bisphenols having the xeno-estrogenic effect suggests the necessity of monitoring those in river water, in the effluent waters from sewage plants, or in landfill leachate.     

Determination of acrylamide in foods by GC/MS using 13C-labeled acrylamide as an internal standard

A method was developed for the determination of trace amounts of acrylamide (AA) in foods. The method includes the addition of 13C-labeled acrylamide-1-13C (AA-1-13C) as an internal standard, extraction with water, bromination, clean-up with a Florisil cartridge column, dehydrobromination and GC/MS analysis in the selected ion monitoring (SIM) mode. Bromination of AA to 2,3-dibromopropionamide (2,3-DBPA) was done using potassium bromide and potassium bromate under an acidic condition. 2,3-DBPA was converted to 2-bromopropenamide (2-BPA) by dehydrobromination with triethylamine before GC/MS analysis. The recoveries of AA from spiked potato chips, corn snack, pretzel and roasted tea were 97-105%, and their relative standard deviations were 0.8-3.9%. The detection limit of AA in foods was 9 ng/g. The method was applied to thirty-one foods purchased from retail markets. AA was found in potato chips at the level of 466-3,340 ng/g, and in other foods at the level of ND-520 ng/g.     

Cleanup of food samples with pre-packed multi-layer silica gel column for the analysis of PCDDs,PCDFs and dioxin-like PCBs

The cleanup procedure for the determination of polychlorinated dibenzo-p-dioxins (PCDDs), dibenzofurans (PCDFs) and dioxin-like polychlorinated biphenyls (PCBs) in food samples using a disposable pre-packed multi-layer silica gel column (multi-layer dioxin tube; D-tube) was evaluated. The blank test showed the need for conditioning of the column with n-hexane. To compare the method with the D-tube and the conventional method for the analyses of actual food samples, seven food samples (spinach, komatsuna, rice, salmon, beef, egg and butter) were extracted by shaking with acetone-n-hexane or n-hexane after alkaline treatment, and then the extracts were cleaned up by use of the D-tube or the prepared conventional column, followed by several column chromatographic steps. Both cleanup procedures gave similar values at each isomeric concentration level and showed similar efficiency with favorable recoveries. The results suggest that the D-tube is applicable to cleanup for the analysis of PCDD/Fs and dioxin-like PCBs in foods.     

Replication of hepatitis C virus in cultured non-neoplastic human hepatocytes

We established a replication system for hepatitis C virus (HCV) using the PH5CH non-neoplastic human hepatocyte line that had been immortalized with simian virus 40 large T antigen. In cells inoculated with sera derived from two HCV-positive blood donors, positive-stranded HCV RNA was detected up to 30 days postinoculation (p.i.). Semi-quantitative analysis of HCV RNA revealed that HCV multiplied during the period of culture. Sequence analysis of the HCV hypervariable region 1 (HVR1) in both cases indicated that HVR1 populations from the cells at 8 days p.i. were apparently different from those of the original inocula. HVR1 populations in infected cells became homogeneous or just a few species were selected over time. These results suggest that HCV is replicating in the human hepatocyte PH5CH cells. This culture system will be useful for detailed studies of the biological effects of HCV in human hepatocytes.     

Specific RNA aptamers to NS3 protease domain of hepatitis C virus

In order to isolate RNA aptamers that bind specifically to NS3 protease domain (delta NS3) of hepatitis C virus, we carried out in vitro selection procedure using RNA pool that had 30 N random core region. After repeating nine cycles of selections and amplifications, a pool of RNAs that bind specifically to the delta NS3 were selected. A comparative analysis of 45 clones that were isolated from 9th cycle revealed three main classes that contain the conserved loop sequences GANUGGGAC. Moreover, the predominant class of aptamer (class I and III) appear to inhibit the protease activity efficiently.