蒽醌染料降解菌XL-1活细胞色素及降解产物的紫外可见光谱分析

应用紫外-可见吸收光谱法研究了一株蒽醌染料降解菌活细胞色素的种类、降解反应程度以及降解产物分子结构的改变。结果表明,蒽醌染料降解菌活细胞中含有细菌叶绿素a以及细菌类胡萝卜素,降解过程中染料分子结构发生了明显的改变,分子原来的共轭结构被破坏,并有新的中间产物生成。

Effects of Diazepam,Phenobarbital,Propranolol,and Cimetidine on Diazepam Oxidizing Isoenzymes in Rat Liver Microsomes

Isolation and identification of the liver microsomal cytochrome P 450 isoen zymes responsible for the formation of diazepam main metabolites nordiazepam and temazepam in rats were studied. The effects of P 450 inducers and inhibitors on the protein contents in SDS poly acrylamide gel electrophoresis and thin layer chromatography to the corresponding diazepam me tabolizing activities of rat liver microsomes were observed. The P 450 contents were dramatically re duced by ip diazepam, cimetidine or propranolol. Diazepam and propranolol inhibited temazepam formation, high dose of propranolol also inhibited nordiazepam formation. Phenobarbital increased the P 450 contents and induced the production of both nordiazepam and temazepam. It also induced proteins with molecular weight (m) of 51 and 59 kDa in SDS PAGE and those with m ranging from 45 to 55 kDa and from 55 to 65 kDa in TLC. Propranolol inhibited both fractions, especially that of m 55～65 kDa, whereas diazepam tended to inhibit the fraction of 45～55 kDa. The protein of m 51 kDa could be mainly involved in diazepam C3 hydroxylation, whereas those of m 59 kDa could be responsible for the N demethylation of diazepam in rats.

Oxidative modification of caspase-9 facilitates its activation via disulfide-mediated interaction with Apaf-1

Intracellular reactive oxygen species （ROS） are known to regulate apoptosis. Activation of caspase-9, the initial caspase in the mitochondrial apoptotic cascade, is closely associated with ROS, but it is unclear whether ROS regulate caspase-9 via direct oxidative modification. The present study aims to elucidate the molecular mechanisms by which ROS mediate caspase-9 activation. Our results show that the cellular oxidative state facilitates caspase-9 activation. Hydrogen peroxide treatment causes the activation of caspase-9 and apoptosis, and promotes an interac- tion between easpase-9 and apoptotic protease-activating factor 1 （Apaf-1） via disulfide formation. In addition, in an in vitro mitochondria-free system, the thiol-oxidant diamide promotes auto-cleavage of caspase-9 and the caspase-9/ Apaf-1 interaction by facilitating the formation of disulfide-linked complexes. Finally, a point mutation at C403 of caspase-9 impairs both H2O2-promoted caspase-9 activation and interaction with Apaf-1 through the abolition of disulfide formation. The association between cytochrome c and the C403S mutant is significantly weaker than that between cytochrome c and wild-type caspase-9, indicating that oxidative modification of caspase-9 contributes to apoptosome formation under oxidative stress. Taken together, oxidative modification of caspase-9 by ROS can medi- ate its interaction with Apaf-1, and can thus promote its auto-cleavage and activation. This mechanism may facilitate apoptosome formation and caspase-9 activation under oxidative stress.

Anti-cancer and anti-angiogenic effects of curcumin and tetrahydrocurcumin on implanted hepatocellular carcinoma in nude mice

AIM: To determine the effect of tetrahydrocurcumin (THC) on tumor angiogenesis compared with curcumin (CUR) by using both in vitro and in vivo models of human hepatocellular carcinoma cell line (HepG2). METHODS: The 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) assay was used for testing the anti-proliferating activities of CUR and THC. In male BALB/c nude mice, 2 × 106 human HepG2 cells were inoculated onto a dorsal skin-fold chamber. One day after HepG2 inoculation, the experimental groups were fed oral daily with CUR or THC (300 mg/kg or 3000 mg/kg). On d 7, 14 and 21, the tumor microvasculature was observed using fluorescence videomicroscopy and capillary vascularity (CV) was measured. RESULTS: Pathological angiogenic features including microvascular dilatation, tortuosity, and hyper-permeability were observed. CUR and THC could attenuate these pathologic features. In HepG2-groups, the CV were significantly increased on d 7 (52.43%), 14 (69.17%), and 21 (74.08%), as compared to controls (33.04%,P < 0.001). Treatment with CUR and THC resulted in significant decrease in the CV (P < 0.005 and P < 0.001, respectively). In particular, the anti-angiogenic effects of CUR and THC were dose-dependent manner. However, the beneficial effect of THC treatment than CUR was observed, in particular, from the 21 d CV (44.96% and 52.86%, P < 0.05). CONCLUSION: THC expressed its anti-angiogenesis without any cytotoxic activities to HepG2 cells even at the highest doses. It is suggested that anti-angiogenic properties of CUR and THC represent a common potential mechanism for their anti-cancer actions.

Retinal ganglion cells of high cytochrome oxidase activity in the rat

Retinal ganglion cells in the rat were studied using the heavy metal intensified oytochrome oxidase and horseradish peroxidase histochemieal methods. The results show that a population of large retinal ganglion cells was consistently observed with the eyto3hrome oxidase staining method in retinas of normal rats or rats which received unilateral thalamotomy at birrth. These oytochrome oxidase rich ganglion cells appeared to have large somata, 3-6 primary dendrites and extensive dendritic arbors, and are comparable to ganglion cells labeled by the wheat germ agglutinin conjugated to horseradish peroxidase (WGA-HRP). However, the morphological details of some of the cells revealed by the cytoahrome oxidase staining method are frequently better than those shown by the HRP histochemieal method. These results suggest that the mit03hondrial enzyme oytoohrome oxidase can be used as a simple but reliable marker for identifying and studying a population of retinal ganglion cells with high metabolie rate in the rat.

Comparative transcriptomic insights into the mechanisms of electron transfer in Geobacter co-cultures with activated carbon and magnetite

Both activated carbon and magnetite have been reported to promote the syntrophic growth of Geobacter metallireducens and Geobacter sulfurreducens co-cultures, the first model to show direct interspecies electron transfer (DIET); however, differential transcriptomics of the promotion on co-cultures with these two conductive materials are unknown. Here, the comparative transcriptomic analysis of G. metallireducens and G. sulfurreducens co-cultures with granular activated carbon (GAC) and magnetite was reported. More than 2.6-fold reduced transcript abundances were determined for the uptake hydrogenase genes of G. sulfurreducens as well as other hydrogenases in those co-cultures to which conductive materials had been added. This is consistent with electron transfer in G. metallireducens-G. sulfurreducens co-cultures as evinced by direct interspecies electron transfer (DIET). Transcript abundance for the structural component of electrically conductive pili (e-pili), PilA, was 2.2-fold higher in G. metallireducens, and, in contrast, was 14.9-fold lower in G. sulfurreducens in co-cultures with GAC than in Geobacters co-cultures without GAC. However, it was 9.3-fold higher in G. sulfurreducens in co-cultures with magnetite than in Geobacters co-cultures. Mutation results showed that GAC can be substituted for the e-pili of both strains but magnetite can only compensate for that of G. sulfurreducens, indicating that the e-pili is a more important electron acceptor for the electron donor strain of G. metallireducens than for G. sulfurreducens. Transcript abundance for G. metallireducens c-type cytochrome gene GMET_RS14535, a homologue to c-type cytochrome gene omcE of G. sulfurreducens was 9.8-fold lower in co-cultures with GAC addition, while that for OmcS of G. sulfurreducens was 25.1-fold higher in co-cultures with magnetite, than in that without magnetite. Gene deletion studies showed that neither GAC nor magnetite can completely substitute the cytochrome (OmcE homologous) of G. metallireducens but compensate for the cytochrome (OmcS) of G. sulfurreducens. Moreover, some genes associated with central metabolism were up-regulated in the presence of both GAC and magnetite; however, tricarboxylic acid cycle gene transcripts in G. sulfurreducens were not highly-expressed in each of these amended co-cultures, suggesting that there was considerable redundancy in the pathways utilised by G. sulfurreducens for electron transfer to reduce fumarate with the amendment of GAC or magnetite. These results support the DIET model of G. metallireducens and G. sulfurreducens and suggest that e-pili and cytochromes of the electron donor strain are more important than that of the electron acceptor strain, indicating that comparative transcriptomics may be a promising route by which to reveal different responses of electron donor and acceptor during DIET in co-cultures.