The structure of the exocellular polysaccharide produced by Rhodococcus sp. RHA1

Rhodococcus sp. RHA1 is a Gram-positive actinomycete capable of metabolizing a wide spectrum of organic compounds whose survival in chemically hostile environments is believed to be in part due to the production of an exocellular polysaccharide (EPS). In order to investigate the functional nature of the EPS, its structure was determined using a combinatory approach including hydrolysis, composition, and methylation, analysis methods, as well as 2D (1)H and (13)C NMR spectroscopy. The EPS was found to be a high-molecular-mass polymer of a repeating tetrasaccharide unit composed of D-glucuronic acid, D-glucose, D-galactose, L-fucose and O-acetyl (1:1:1:1:1), and has the structure:     

Effects of the environmental mammary carcinogen 6-nitrochrysene on p53 and p21(Cip1) protein expression and cell cycle regulation in MCF-7 and MCF-10A cells

The environmental pollutant 6-nitrochrysene (6-NC) is a potent mammary carcinogen in rats; it is more potent than numerous classical mammary carcinogens such as benzo[a]pyrene (BaP). The mechanisms that account for the remarkable carcinogenicity of 6-NC remain elusive. Similar to BaP, 6-NC is also known to induce DNA damage in rodents and in human breast tissues. As an initial investigation, we reasoned that DNA damage induced by 6-NC may alter the expression of p53 protein in a manner that differs from other DNA damaging carcinogens (e.g. BaP). Using human breast adenocarcinoma MCF-7 cells and immortalized human mammary epithelial MCF-10A cells, we determined the effects of 6-NC on the expression of p53 protein and its direct downstream target cyclin-dependent kinase inhibitor p21(Cip1) as well as on the cell cycle progression. Western blot analysis demonstrated that treatments of MCF-7 and MCF-10A cells with 6-NC for 12, 24 or 48h did not increase the level of total p53 protein; however, an increase of p21(Cip1) protein and a commitment increase of G(1) phase were observed in MCF-10A cells but not in MCF-7 cells. Further studies using 1,2-dihydroxy-1,2-dihydro-6-hydroxylaminochrysene (1,2-DHD-6-NHOH-C), the putative ultimate genotoxic metabolite of 6-NC, was conducted and showed a significant induction of p53 (p<0.05) in MCF-7 cells; however, this effect was not evident in MCF-10A cells, indicating the varied DNA damage responses between the two cell lines. By contrast to numerous DNA damaging agents such as BaP which is known to stimulate p53 expression, the lack of p53 response by 6-NC imply the lack of protective functions mediated by p53 (e.g. DNA repair machinery) after exposure to 6-NC and this may, in part, account for its remarkable carcinogenicity in the mammary tissue.     

MiR‐629 regulates hypoxic pulmonary vascular remodelling by targeting FOXO3 and PERP

Pulmonary arterial hypertension (PAH) is featured by the increase in pulmonary vascular resistance and pulmonary arterial pressure. Despite that abnormal proliferation and phenotypic changes in human pulmonary artery smooth muscle cells (HPASMCs) contributing to the pathophysiology of PAH, the underlying molecular mechanisms remain unclear. In the present study, we detected the expression of miR‐629 in hypoxia‐treated HPASMCs and explored the mechanistic role of miR‐629 in regulating HPASMC proliferation, migration and apoptosis. Hypoxia time‐dependently induced up‐regulation of miR‐629 and promoted cell viability and proliferation in HPASMCs. Treatment with miR‐629 mimics promoted HPASMCs proliferation and migration, but inhibited cell apoptosis; while knockdown of miR‐629 suppressed the cell proliferation and migration but promoted cell apoptosis in HPASMCs. The bioinformatics prediction revealed FOXO3 and PERP as downstream targets of miR‐629, and miR‐629 negatively regulated the expression of FOXO3 and PERP via targeting the 3’ untranslated regions. Enforced expression of FOXO3 or PERP attenuated the miR‐629 overexpression or hypoxia‐induced enhanced effects on HPASMC proliferation and proliferation, and the suppressive effects on HPASMC apoptosis. Furthermore, the expression of miR‐629 was up‐regulated, and the expression of FOXO3 and PERP mRNA was down‐regulated in the plasma from PAH patients when compared to healthy controls. In conclusion, the present study provided evidence regarding the novel role of miR‐629 in regulating cell proliferation, migration and apoptosis of HPASMCs during hypoxia.     

CYP1A1 gene polymorphisms and smoking status as modifier factors for lung cancer risk

Objective

Lung cancer remains the most prevalent malignancy worldwide. Susceptibility to lung cancer has been shown to be modulated by inheritance of polymorphic genes. Several metabolic enzymes are currently under investigation for their possible role in lung cancer susceptibility, including members of the cytochrome P450 (CYP) superfamily. The aim of this work was to identify the correlation between CYP1A1 m1 and m2 polymorphisms and lung cancer risk and figure its interactions with smoking as genetic modifiers in the etiology of lung cancer in the Egyptian population.

Materials and methods

One hundred and ten patients with lung cancer and one hundred and ten controls were enrolled in the study. CYP1A1 m1 and m2 polymorphisms were determined using polymerase chain reaction restriction fragment length polymorphism.

Results

Subjects carrying TC and CC genotypes of CYP1A1 m1 and AG and GG genotypes of CYP1A1 m2 were significantly more likely to develop lung cancer especially squamous cell carcinoma. The proportion of lung cancer attributable to the interaction of smoking and CYP1A1 m1 and CYP1A1 m2 polymorphisms was 32% and 52% respectively.

Conclusion

Our results revealed that CYP1A1 m1 and m2 polymorphisms contribute to smoking related lung cancer risk in the Egyptian population.     

A Whole Cell Bioreporter Approach to Assess Transport and Bioavailability of Organic Contaminants in Water Unsaturated Systems

Bioavailability of contaminants is a prerequisite for their effective biodegradation in soil. The average bulk concentration of a contaminant, however, is not an appropriate measure for its availability; bioavailability rather depends on the dynamic interplay of potential mass transfer (flux) of a compound to a microbial cell and the capacity of the latter to degrade the compound. In water-unsaturated parts of the soil, mycelia have been shown to overcome bioavailability limitations by actively transporting and mobilizing organic compounds over the range of centimeters. Whereas the extent of mycelia-based transport can be quantified easily by chemical means, verification of the contaminant-bioavailability to bacterial cells requires a biological method. Addressing this constraint, we chose the PAH fluorene (FLU) as a model compound and developed a water unsaturated model microcosm linking a spatially separated FLU point source and the FLU degrading bioreporter bacterium Burkholderia sartisoli RP037-mChe by a mycelial network of Pythium ultimum. Since the bioreporter expresses eGFP in response of the PAH flux to the cell, bacterial FLU exposure and degradation could be monitored directly in the microcosms via confocal laser scanning microscopy (CLSM). CLSM and image analyses revealed a significant increase of the eGFP expression in the presence of P. ultimum compared to controls without mycelia or FLU thus indicating FLU bioavailability to bacteria after mycelia-mediated transport. CLSM results were supported by chemical analyses in identical microcosms. The developed microcosm proved suitable to investigate contaminant bioavailability and to concomitantly visualize the involved bacteria-mycelial interactions.     

Cytochrome P450 expression is moult stage specific and regulated by ecdysteroids and xenobiotics in the crab Carcinus maenas

The shore crab Carcinus maenas has a high capacity for metabolizing polycyclic aromatic hydrocarbons (PAHs). Cytochrome P450 (CYP) enzymes are involved in this metabolism and also have a role in development and reproduction. This investigation is a systematic gene expression analysis of six CYPs in C. maenas. Expression of CYP2 and CYP3-like genes was predominant in hepatopancreas, while expression of CYP4-like genes was predominant in gills and epidermis. Expression of all six CYP genes fluctuated over the moult cycle in the hepatopancreas and structurally related genes were regulated coordinately. The study suggests that hepatopancreas is a major site of CYP gene expression in C. maenas confirming previous biochemical studies showing that this tissue is the major compartment for CYP mediated xenobiotic metabolism in crustaceans. In addition, the data show that CYP2 and CYP3 related genes respond to ecdysteroid and xenobiotic treatment, while those related to CYP4 genes do not and likely are involved in a more general physiological function such as fatty acid metabolism. The developmental variations of CYP expression suggest a molecular mechanism for the stage specific susceptibility of crabs exposed to environmental pollutants.     

Factors influencing the electrokinetic dispersion of PAH-degrading bacteria in a laboratory model aquifer

Despite growing interest in the electro-bioremediation of contaminated soil it is still largely unknown to which degree weak electric fields influence the fate of contaminant-degrading microorganisms in the sub-surface. Here we evaluate the factors influencing the electrokinetic transport and deposition of fluorene-degrading Sphingomonas sp. LB126 in a laboratory model aquifer exposed to a direct current (DC) electric field (1 V cm(-1)) typically used in electro-bioremediation measures. The influence of cell size, cell membrane integrity, cell chromosome contents (all assessed by flow cytometry), cell surface charge and cell hydrophobicity on the spatial distribution of the suspended and matrix-bound cells after 15 h of DC-treatment was evaluated. In presence of DC the cells were predominantly mobilised by electroosmosis to the cathode with an apparent velocity of 0.6 cm h(-1), whereas a minor fraction only of the cells augmented was mobilised to the anode by electrophoresis. Different electrokinetic behaviour of individual cells could be solely attributed to intra-population heterogeneity of the cell surface charge. In the absence of DC by contrast, a Gaussian-type distribution of bacteria around the point of injection was found. DC had no influence on the deposition efficiency, as the glass beads in presence and absence of an electric field retained quasi-equal fractions of the cells. Propidium iodide staining and flow cytometry analysis of the cells indicated the absence of negative influences of DC on the cell wall integrity of electrokinetically mobilised cells and thus point at unchanged physiological fitness of electrokinetically mobilised bacteria.     

An Integrated Case Study for Evaluating the Impacts of an Oil Refinery Effluent on Aquatic Biota in the Delaware River: Advanced Chemical Fingerprinting of PAHs

More than one thousand samples were collected and analyzed to evaluate the potential impact of Motiva's oil refinery effluent on the receiving water, sediment, and biota of the Delaware River. The data collected from these samples were used with advanced chemical fingerprinting of polycyclic aromatic hydrocarbons (PAHs) in Motiva's oil refinery effluent to differentiate Motiva-related PAHs in sediment and biota from other sources. The PAHs released from the refinery between 1999 and 2002 were dominated by petrogenic 4-ring PAHs. Specifically, the refinery signature exhibited relatively high levels of fluoranthenes/pyrenes with two (FP2) and three (FP3) alkyl groups and benz(a)anthracene/chrysenes with two (BC2), three (BC3), and four (BC4) alkyl groups. This PAH signature, attributed to accelerated degradation of low molecular weight PAHs in the Motiva wastewater treatment plant, exhibited little variability over time relative to the background patterns in the Delaware River. This distinctive feature of the Motiva effluent allowed the identification of this source in other samples. Water and sediment samples identified a range of PAH characteristics associated with the Delaware River urban background signature. These characteristics included varying levels of 2- to 3-ring PAHs (likely from weathered automotive fuel, marine fuel, or bilge tank discharges), pyrogenic 4- to 6-ring PAHs (from partially combusted organic material like soot), and perylene (diagenetic product of terrestrial plant decomposition). The Motiva hydrocarbon signature was only evident at moderate to low levels in selected near-field sampling stations for sediment, bivalves, and effluent/nearfield water. PAHs in the river sediments beyond the near-field area were consistently associated with samples containing the Delaware River urban background signature, and exhibited little to no effect from the Refinery.     

Petroleum-degrading microbial numbers in rhizosphere and non-rhizosphere crude oil-contaminated soil

Phytoremediation can be a cost-effective and environmentally acceptable method to clean up crude oil-contaminated soils in situ. Our research objective was to determine the effects of nitrogen (N) additions and plant growth on the number of total hydrocarbon (TH)-, alkane-, and polycyclic aromatic hydrocarbon (PAH)-degrading microorganisms in weathered crude oil-contaminated soil. A warm-season grass, sudangrass (Sorghum sudanense (Piper) Stapf), was grown for 7 wk in soil with a total petroleum hydrocarbon (TPH) level of 16.6 g TPH/kg soil. Nitrogen was added based upon TPH-C:added total N (TPH-C:TN) ratios ranging from 44:1 to 11:1. Unvegetated and unamended controls were also evaluated. The TH-, alkane-, and PAH-degrading microbial numbers per gram of dry soil were enumerated from rhizosphere and non-rhizosphere soil for vegetated pots and non-rhizosphere soil populations were enumerated from non-vegetated pots. Total petroleum-degrading microbial numbers were also calculated for each pot. The TH-, alkane-, and PAH-degrading microbial numbers per gram of dry soil in the sudangrass rhizosphere were 3.4, 2.6, and 4.8 times larger, respectively, than those in non-rhizosphere soil across all N rates. The presence of sudangrass resulted in significantly more TH-degrading microorganisms per pot when grown in soil with a TPH-C:TN ratio of 11:1 as compared to the control. Increased plant root growth in a crude oil-contaminated soil and a concomitant increase in petroleum-degrading microbial numbers in the rhizosphere have the potential to enhance phytoremediation.