Impaired arachidonic acid-mediated dilation of small mesenteric arteries in Zucker diabetic fatty rats

Arachidonic acid (AA) is a precursor of important vasoactive metabolites, but the role of AA-mediated vasodilation in Type 2 diabetes is not known. Using Zucker diabetic fatty (ZDF) rats, we examined the effects of AA in small mesenteric arteries preconstricted with endothelin. In ZDF rat mesenteric arteries, 1 microM AA produced only one-third the amount of dilation as in vessels from lean control animals. In lean control rats, the effect of AA was significantly and predominantly inhibited by the lipoxygenase inhibitors baicalein and cinnamyl-3,4-dihydroxy-cyanocinnamate (CDC). However, baicalein and CDC had no effect on AA-mediated dilation in ZDF rat mesenteric arteries. The major [3H]AA metabolite produced by isolated mesenteric arteries in both lean and ZDF rats was 12-hydroxyeicosatetraenoic acid (12-HETE), but the amount of [3H]12-HETE produced by ZDF rat vessels was only 36% of that of control vessels. In addition, 12-HETE produced similar amounts of dilation in lean and ZDF rat mesenteric arteries. Immunoblot analysis showed an 81% reduction in 12-lipoxygenase protein in ZDF rat mesenteric arteries. Immunofluorescence labeling showed strong nitrotyrosine signals in ZDF rat mesenteric arteries that colocalized with 12-lipoxygenase in endothelium, and 12-lipoxygenase coprecipitation with anti-nitrotyrosine antibodies was enhanced in ZDF rat vessels. We conclude that AA-mediated relaxation in ZDF rat small mesenteric arteries is impaired due to reduced 12-lipoxygenase protein and activity. Increased oxidative stress and nitration of 12-lipoxygenase may underlie the impairment of AA-mediated relaxation in small mesenteric arteries of diabetic rats.     

雪莲类PEBP基因在大肠杆菌中的表达及鉴定

本实验旨在构建雪莲类PEBP基因原核表达载体,在大肠杆菌中表达并纯化类PEBP基因所编码的蛋白,为进一步研究奠定基础.将雪莲类PEBP基因开放阅读框序列克隆到原核表达载体pET30( )上,转化感受态表达菌株BL21(DE3),低浓度IPTG低温诱导融合蛋白的表达,纯化产物,Western blotting鉴定目的蛋白.IFTG低诱导PEBP,经SDS-PAGE分析,其相对分子量约为28 kD,与预期相符,表达量约占菌体蛋白的26.8%,并且通过亲和层析纯化了重组融合蛋白,Western blotting鉴定为阳性.成功构建了原核表达载体pET-PEBP,获得了高效表达产物,并为进一步研究雪莲类PEBP基因的抗冻功能打下基础.     

Molecular diversity of 16S rRNA and gyrB genes in copper mines

The molecular diversities of the microbial communities from four sites impacted by acid mine drainage (AMD) at Dexing Copper Mine in Jiangxi province of China were studied using 16S rRNA sequences and gyrB sequences. Of the four sampled sites, each habitat exhibited distinct geochemical characteristics and the sites were linked geographically allowing us to correlate microbial community structure to geochemical characteristics. In the present study, we examined the molecular diversity of 16S rRNA and gyrB genes from water at these sites using a PCR-based cloning approach. We found that the microbial community appears to be composed primarily of Proteobacteria, Acidobacteria, Actinobacteria, Nitrospira, Firmicutes, Chlorella and unknown phylotypes. Of clones affiliated with Nitrospira, Leptospirillum ferrooxidans, Leptospirillum ferriphilum and Leptospirillum group III were all detected. Principal-component analysis (PCA) revealed that the distribution of the microbial communities was influenced greatly by geochemical characteristics. The overall PCA profiles showed that the sites with similar geochemical characteristics had more similar microbial community structures. Moreover, our results also indicated that gyrB sequence analysis may be very useful for differentiating very closely related species in the study of microbial communities. H. Yin and L. Cao contributed equally to this work.     

The new incorporation bio-treatment technology of bromoamine acid and azo dyes wastewaters under high-salt conditions

The accelerating effect of quinones has been studied in the bio-decolorization processes, but there are no literatures about the incorporation bio-treatment technology of the bromoamine acid (BA) wastewater and azo dyes wastewaters under high-salt conditions (NaCl, 15%, w/w). Here we described the BA wastewater as a redox mediator in the bio-decolorization of azo dye wastewaters. Decolorization of azo dyes was carried out experimentally using the salt-tolerant bacteria under the BA wastewater and high-salt conditions. The BA wastewater used as a redox mediator was able to increase the decolorization rate of wastewater containing azo dyes. The effects of various operating conditions such as dissolved oxygen, temperature, and pH on microbial decolorization were investigated experimentally. At the same time, BA was tested to assess the effects on the change of the Oxidation–Reduction Potential (ORP) values during the decolorization processes. The experiments explored a great improvement of the redox mediator application and the new bio-treatment concept.     

Overexpression of phytoene synthase gene from Salicornia europaea alters response to reactive oxygen species under salt stress in transgenic Arabidopsis

A phytoene synthase gene SePSY was isolated from euhalophyte Salicornia europaea L. The 1655 bp full-length SePSY has an open reading frame of 1257 bp and encodes a 419-amino acid protein. The overexpression of SePSY enhanced the growth of transgenic Arabidopsis. When the plants were exposed to 100 mM NaCl, the photosynthesis rate and photosystem II activity (Fv/Fm) increased from 92% to 132% and from 9.3% to 16.6% in the transgenic lines than in the wild-type, respectively. The transgenics displayed higher activities of SOD and POD and lower contents of H(2)O(2) and MDA than the WT. In conclusion, the transgenic lines showed higher tolerance to salt stress than WT plants by increased photosynthesis efficiency and antioxidative capacity. This is the first report about improving the salt tolerance by genetic manipulation of carotenoid biosynthesis.     

脂肪酶产生菌的筛选及鉴定研究

为寻找合适的产脂肪酶野生菌,以期能高效的催化合成生物柴油。通过添加橄榄油作为惟一碳源进行富集培养,然后以透明圈平板筛选法从油污土壤样品中成功地筛选到了一株酶活力值为10.12U/ml产脂肪酶菌株。通过对该菌株表型、生理生化特征分析及16S rRNA部分序列的系统进化发育分析鉴定该菌为芽胞杆菌属,定名为BacilluspumilusB2。     

Proteomic characterization and functional analysis of outer membrane vesicles of Francisella novicida suggests possible role in virulence and use as a vaccine

We have isolated and characterized outer membrane vesicles (OMVs) from Francisella. Transport of effector molecules through secretion systems is a major mechanism by which Francisella tularensis alters the extracellular proteome and interacts with the host during infection. Outer membrane vesicles produced by Francisella were examined using TEM and AFM and found to be 43-125 nm in size, representing another potential mechanism for altering the extracellular environment. A proteomic analysis (LC-MS/MS) of OMVs from F. novicida and F. philomiragia identified 416 (F. novicida) and 238 (F. philomiragia) different proteins, demonstrating that OMVs are an important contributor to the extracellular proteome. Many of the identified OMV proteins have a demonstrated role in Francisella pathogenesis. Biochemical assays demonstrated that Francisella OMVs possess acid phosphatase and hemolytic activities that may affect host cells during infection, and are cytotoxic toward murine macrophages in cell culture. OMVs have been previously used as a human vaccine against Neisseria meningitidis . We hypothesized that Francisella OMVs could be useful as a novel Francisella vaccine. Vaccinated BALB/C mice challenged with up to 50 LD50 of Francisella showed statistically significant protection when compared to control mice. In the context of these new findings, we discuss the relevance of OMVs in Francisella pathogenesis as well as their potential use as a vaccine.     

Multiplex PCR and Reverse Line Blot Hybridization Assay (mPCR/RLB)

Multiplex PCR/Reverse Line Blot Hybridization assay allows the detection of up to 43 molecular targets in 43 samples using one multiplex PCR reaction followed by probe hybridization on a nylon membrane, which is re-usable. Probes are 5'' amine modified to allow fixation to the membrane. Primers are 5'' biotin modified which allows detection of hybridized PCR products using streptavidin-peroxidase and a chemiluminescent substrate via photosensitive film. With low setup and consumable costs, this technique is inexpensive (approximately US$2 per sample), high throughput (multiple membranes can be processed simultaneously) and has a short turnaround time (approximately 10 hours).The technique can be utilized in a number of ways. Multiple probes can be designed to detect sequence variation within a single amplified product, or multiple products can be amplified simultaneously, with one (or more) probes used for subsequent detection. A combination of both approaches can also be used within a single assay. The ability to include multiple probes for a single target sequence makes the assay highly specific.Published applications of mPCR/RLB include detection of antibiotic resistance genes^1,2, typing of methicillin-resistant Staphylococcus aureus^3-5 and Salmonella sp⁶, molecular serotyping of Streptococcus pneumoniae^7,8, Streptococcus agalactiae⁹ and enteroviruses^10,11, identification of Mycobacterium sp¹², detection of genital^13-15 and respiratory tract¹⁶ and other¹⁷ pathogens and detection and identification of mollicutes¹⁸. However, the versatility of the technique means the applications are virtually limitless and not restricted to molecular analysis of micro-organisms.The five steps in mPCR/RLB are a) Primer and Probe design, b) DNA extraction and PCR amplification c) Preparation of the membrane, d) Hybridization and detection, and e) Regeneration of the Membrane.     

Wld(S) reduces paraquat-induced cytotoxicity via SIRT1 in non-neuronal cells by attenuating the depletion of NAD

Wld(S) is a fusion protein with NAD synthesis activity, and has been reported to protect axonal and synaptic compartments of neurons from various mechanical, genetic and chemical insults. However, whether Wld(S) can protect non-neuronal cells against toxic chemicals is largely unknown. Here we found that Wld(S) significantly reduced the cytotoxicity of bipyridylium herbicides paraquat and diquat in mouse embryonic fibroblasts, but had no effect on the cytotoxicity induced by chromium (VI), hydrogen peroxide, etoposide, tunicamycin or brefeldin A. Wld(S) also slowed down the death of mice induced by intraperitoneal injection of paraquat. Further studies demonstrated that Wld(S) markedly attenuated mitochondrial injury including disruption of mitochondrial membrane potential, structural damage and decline of ATP induced by paraquat. Disruption of the NAD synthesis activity of Wld(S) by an H112A or F116S point mutation resulted in loss of its protective function against paraquat-induced cell death. Furthermore, Wld(S) delayed the decrease of intracellular NAD levels induced by paraquat. Similarly, treatment with NAD or its precursor nicotinamide mononucleotide attenuated paraquat-induced cytotoxicity and decline of ATP and NAD levels. In addition, we showed that SIRT1 was required for both exogenous NAD and Wld(S)-mediated cellular protection against paraquat. These findings suggest that NAD and SIRT1 mediate the protective function of Wld(S) against the cytotoxicity induced by paraquat, which provides new clues for the mechanisms underlying the protective function of Wld(S) in both neuronal and non-neuronal cells, and implies that attenuation of NAD depletion may be effective to alleviate paraquat poisoning.     

Cadmium Is More Toxic on Volume Bone Mineral Density than Tissue Bone Mineral Density

It has been showed that Cd induces low areal bone mineral density, but we do not know the effect of Cd on cubic bone density. This study was aimed to investigate the effects of Cd on volumetric bone mineral density (VBMD) and tissue bone mineral density (TBMD) in male rats. Twenty-four Sprague-Dawley male rats were randomly divided into four groups that were given cadmium chloride by subcutaneous injection at doses of 0, 0.1, 0.5, and 1.5?mg/kg body weight for 8?weeks, respectively. Then, microcomputed tomography scanning was performed on the proximal tibia, and region of interest was reconstructed using microview software. The VBMD, bone volume fraction of rats treated with 1.5?mg Cd/kg, were significantly decreased compared to control (p?相似文献