Tobacco smoke dysregulates endothelial vasoregulatory transcripts in vivo.

We hypothesized that human smoking and its deleterious effects on endothelial function can be modeled by exposure of mice to tobacco smoke, and further that these changes would be reflected in gene regulation in vascular endothelium. We used for these studies a mouse strain that expresses green fluorescent protein under the control of an endothelial-specific promoter, Tie-2. Mice were exposed to sidestream smoke from reference cigarettes at 34 mg total suspended particulates/m3. After exposure for 5 days/wk for 1 and 6 wk, aortas were pooled from treatment and control groups. Endothelial cells were rapidly isolated by collagenase treatment followed by fluorescent activated cell sorting to yield populations of >95% purity. RNA isolated from >500 endothelial cells was amplified and analyzed on deeply representative long oligo microarrays. Transcripts dysregulated by >2.5-fold were confirmed by real-time PCR and selected proteins by immunofluorescent localization. In the endothelial cells, the observed more than threefold upregulation of complement factor H (Cfh), calcitonin receptor-like (Calcr1), and soluble epoxide hydrolase (Epxh2) may play a role in hypertensive responses of the vasculature to smoking. We have identified gene regulation in vivo in vascular endothelium that potentially underlies hypertensive responses to tobacco smoke.

In vivo transcriptional response of cardiac endothelium to lipopolysaccharide.

OBJECTIVE: Vascular endothelial cells must integrate stimuli from multiple sources, including plasma, leukocytes, and neighboring components of the vessel. These stimuli are difficult to recapitulate in vitro. We have developed a method to examine the in vivo regulation of gene expression in endothelial cells and have applied it to a model of sepsis. METHODS AND RESULTS: We used fluorescent-activated cell sorting to isolate highly purified endothelial cells from the hearts of transgenic mice that express green fluorescent protein driven by the endothelial-specific promoter Tie2. We treated these mice with intraperitoneal lipopolysaccharide and identified those genes within cardiac endothelium that were >3-fold dysregulated 4 and 24 hours later by microarray analysis. These findings were confirmed by real-time polymerase chain reaction and compared with in vitro regulation in a murine endothelial cell line. CONCLUSIONS: The in vivo regulation was distinct and, in general, more robust than that seen in vitro. We identified endothelial-expressed genes not previously recognized to be regulated in response to lipopolysaccharide. This approach provides insight into the cardiac-specific responses of the endothelium that contribute to the specific responses of the heart to sepsis, and can be generalized to the exploration of endothelial responses in any organ.

Short-term toxic effects of chlorobenzenes on broadbean (Vicia faba) seedlings.

The root growth, changes in superoxide dismutase (SOD, EC 1.15.1.1) activity, malonyldialdehyde (MDA) and total soluble protein level of broadbean (Vicia faba) seedlings were researched at different soil concentrations of chlorobenzene (CB), 1,2,4-trichlorobenzene (TCB) and hexachlorobenzene (HCB). The results showed that root growth of seedlings was interrupted after 5 d of 50-200 microg x g(-1) TCB treatment. During a 3 d recovery period, root growth was, however, restored to some extent although there was a delay in returning to the control level. The total soluble protein content in seedlings increased with TCB concentration and duration of exposure. Effect of TCB stress on SOD activity in seedlings displayed a significant dose-effect relationship for 1-5 d of 50-200 microg x g(-1) treatment. When broadbean seedlings were placed in clean tap water for 3 d following exposure to 5 d of TCB stress to clear tap water for 3 d, SOD activity at 50 microg x g(-1) TCB recovered towards control level (P>0.05) while a significant increase in SOD activity was observed at 100 and 200 microg x g(-1) TCB compared to control (P<0.05). The experiments also revealed that a significant increase of MDA level in seedlings occurred after 3 and 5 d of 100 and 200 microg x g(-1) TCB treatment (P<0.05 and P<0.01), and there was a positive correlation between TCB concentration and MDA level. All the above results showed that SOD activity and MDA level of broadbean seedlings might be proposed as the biomarkers for short-term TCB contamination in soil. Compared to TCB, the toxicity of 50-1000 microg x g(-1) CB or HCB in soil to broadbean seedlings was not observed after a 3 d exposure.

Microbial changes in rhizospheric soils contaminated with petroleum hydrocarbons after bioremediation.

Effects of bioremediation on microbial communities in soils contaminated with petroleum hydrocarbons are a scientific problem to be solved. Changes in dominate microbial species and the total amount of microorganisms including bacteria and fungi in rhizospheric soils after bioremediation were thus evaluated using field bioremediation experiments. The results showed that there were changed dominant microorganisms including 11 bacterial strains which are mostly Gram positive bacteria and 6 fungal species which were identified. The total amount of microorganisms including bacteria and fungi increased after bioremediation of microbial agents combined with planting maize. On the contrary, fungi in rhizospheric soils were inhibited by adding microbial agents combined with planting soybean.

[Degradation and kinetics of pyrene and benzo [a] pyrene by three bacteria in contaminated soil].

Degradation kinetics of pyrene and benzo [a] pyrene (BaP) in soil by three individual strains of bacteria were examined using the Michaelis-Menton and Monod dynamic models. There were significant differences occurred in the degradation rates of pyrene and BaP by the three individual strains. When the initial concentrations of pyrene or BaP was 50 mg/kg in soil, the degradation rates of pyrene by Bacillus sp. SB02, Zoogloea sp.SB09, and Flavobacterium sp.SB10 in 42 days were 42.69%, 32.88% and 25.07% respectively for pyrene. Similarly, degradation rates of BaP by Bacillus sp. SB02, Zoogloea sp. SB09, and Flavobacterium sp. SB10 in 42 days were 33.04%, 25.39% and 22.02% respectively for BaP. In addition, there also were significant differences occurred in the degradation rates per day of pyrene and BaP by the three individual strains. The most rapid degradation of pyrene and BaP by Bacillus sp. SB02 occurred in one week, and 20.88% of pyrene and 12.6% of BaP were degraded. However, Zoogloea sp. SB09 had a slower degradation, while Flavobacterium sp. SB10 had the slowest degradation.

[Remediation of polycyclic aromatic hydrocarbons in non-fluid medium with immobilized microorganism technique].

The remediation of polycyclic aromatic hydrocarbons (PAHs) in non-fluid medium is a difficult and urgent task for environmental scientists. Due to the particularities of non-fluid medium, conventional techniques cannot work effectively, while bioremediation techniques using free microorganism also have many limitations that restricted the use of microorganism in removing PAHs from non-fluid medium. Immobilized microorganism (IM) technique is a proven approach in increasing the stability of microorganism under unfavorable conditions, which can not only provide a special microenvironment to protect dominant microorganisms from the malignant competition of aboriginal microorganism, but also improve the tolerance against toxic compounds in environment. Biodegradation using immobilized cells has been widely investigated for numerous toxic compounds in fluid medium such as in wastewater, or in half-fluid medium such as in slurry. In this paper, based on analyzing the characteristics of IM technique and its feasibility, the problem and development of future research were presented, and the new technique using IM to remediate PAHs in non-fluid medium was put forward.

[Comparative study on ecotoxicological indexes of crops exposure to reactive X-3B red dye].

Ecotoxicological indexes of wheat, Chinese cabbage and rice exposed to reactive X-3B red dye were studied using the experimental method of crop seed exposure. The results showed that reactions of the three crops to reactive X-3B red dye were different from each other. Based on the inhibition of seed gemination, the half effect concentration(IC50) of wheat, Chinese cabbage and rice was 8965, 7010 and 7514 mg.L-1. Based on the inhibition of root elongation, the IC50 of wheat, Chinese cabbage and rice was 6534, 5882 and 4570 mg.L-1. When seedlings were cultured for seven days after the gemination of seeds and the remaining seeds or husks were discarded, fresh and dry growing biomass of the three crops exposed to the dye was both decreased with the increasing the concentration of reactive X-3B red dye. However, the decrease rate of fresh biomass was higher than that of dry biomass. Biomass index of wheat was most sensitive to reactive X-3B red dye, followed by rice, and Chinese cabbage was most inertia when the crops were exposed to reactive X-3B red dye with the high concentration of 4500 and 5000 mg.L-1. As far as the tolerance of wheat, Chinese cabbage and rice to reactive X-3B red dye was concerned, wheat had the strongest tolerance, on the contrary rice had the wakestone.

[Acute toxicological effects of heavy metal pollution in soils on earthworms].

Acute and sub-acute lethal effects of single and combined Cu, Zn, Pb and Cd on earthworm was determined using meadow brown soil. Results indicated that the mortality of earthworm was significantly related with the concentrations of Cu and Zn in the soil (alpha = 0.05, RCu = 0.86, RPb = 0.87), and the inhibition rate of earthworm growth was significantly related with the concentration of Cu in the soil (alpha = 0.05, RCu = 0.84). There was no significant relation between the concentrations of other heavy metals tested and the death rate and the growth-inhibition rate. The degree of individual earthworm enduring the toxicity of heavy metals varied greatly. The threshold concentration of toxicity defined by statistical method, the level to conduce the death of individual earthworm was 300 mg.kg-1 for Cu, 1300 mg.kg-1 for Zn, 1700 mg.kg-1 for Pb, 300 mg.kg-1 for Cd. LC50 was 400-450 mg.kg-1 for Cu, 1500-1900 mg.kg-1 for Zn, 2350-2400 mg.kg-1 for Pb and 900 mg.kg-1 for Cd. Combined effects of single Cu, Zn, Pb and Cd to conduce more than 10% of the death rate of earthworm could result in 100% of the death rate of earthworm, thus showing the strong synergistic joint effect of the heavy metals.

[Revulsive response of protein and enzyme activity of degradative Bacilluss strains by phenanthrene].

Three strains of Bacillus sp. BAll, BA19 and BA27, which can grow when phenanthrene is taken as the sole source of carbon and energy, were isolated from an oil-contaminated soil. Their crude extract was used to test the activities of polyphenol oxidase and catalase. The results demonstrated that all the three strains did not show a significant change in the activity of both enzymes when phenanthrene was lower than 200 mg x L(-1). Under different phenanthrene concentration, the enzyme activities of strains BA19, BA27 showed a higher stabilization than BA11. Using the SDS-PAGE contrasted the difference of proteins between the strains in normal medium and the strains growing on phenanthrene, the results showed that BA27 growing on phenanthrene synthesized a new protein with 27000 dal of molecular weight. At the same time, the synthesis of some proteins was cancelled. We conjecture that the new protein induced by phenanthrene may be related to the ability of phenanthrene degradation and the stabilization of enzyme activity by the strain.

[Effect of short-term phenanthrene stress on SOD activities and MDA contents in soybean (Glycine max) seedlings].

Growth of soybean (Glycine max) seedlings, changes of superoxide dismutase (SOD) activities, and malonyldialdehyde (MDA) contents were researched during different concentration phenanthrene stress and following recovery from stress. The results showed that seedling growth was interrupted after 5 d in 200 micrograms.g-1 phenanthrene treatment. However, during the period of recovery, growth was restored, although there was a delay in returning to control levels. The dose-related response of SOD activities were observed in manner of linear for 2 d and of parabola for 5 d and 8 d at 50-200 micrograms.g-1 phenanthrene. SOD activities of seedling were significantly induced by 100 and 200 micrograms.g-1 phenanthrene for 2 d (P < 0.05 and P < 0.01), while declined significantly under phenanthrene stress for 8 d, and was 88% of control at 200 micrograms.g-1 phenanthrene (P < 0.05). When releasing soybean seedlings from phenanthrene stress for 5 d to clear tap water for 2 d or 4 d, their SOD activities at 50 and 100 micrograms.g-1 phenanthrene recovered towards control level, while a significant increase in SOD activities was observed at 200 micrograms.g-1 phenanthrene (P < 0.05). The experiments also revealed that a significant increase of MDA contents in seedlings occurred after 5 d and 8 d in 100 and 200 micrograms.g-1 phenanthrene treatment (P < 0.05 and P < 0.01), which suggested that the membrane systems were damaged by phenanthrene stress. All of the above results showed that SOD activities of soybean seedling might be suitable to be the biomarkers of phenanthrene stress.

[Eco-toxicology of heavy metals on the inhibition of seed germination and root elongation of wheat in soils].

The eco-toxicity of Cu, Zn, Pb, and Cd in the single form was tested with four types of soils(red loam soils, meadow brown soils, chestnut soils, and dark brown soils) and their combined effect was determined with meadow brown soils. The results showed that with the same content of heavy metals, the root elongation was strongly inhabited than the seed germination, and it indicated that root was more sensitive to heavy metals pollution. The inhibition rate of heavy metals pollution on the root elongation (IRHMRE) of wheat was significantly related with the contents of organic matter (OR) and Kjedahl-nitrogen (K-N) in soils (R2OR = 0.91, R2K-N = 0.92). However, IRHMRE was not significantly related with soil pH as did with OR and K-N, so does the content of CEC (R2pH = 0.62, R2CEC = 0.60). Whether under the low concentration of Cu, Zn, Pb, and Cd resulted in an irritated effect in root elongation, or under high concentration resulted in an inhibition effect in the single form, an synergistic effect occurred in the combined form.

[Bioremediation of PAHs contaminated soil using bio-slurry reactor process].

Studies on bioremediation of PAHs contaminated soil were carried out using bio-slurry reactor process. Phenanthrene (PHE) and pyrene (PY) were chosen as the test pollutants. The results showed that the physical and chemical properties of the pollutants were key factor affecting the bio-remedying possibility of PAHs in soil. PAHs with less benzene rings and lower molecule weight were easier to be biodegraded. So, phenanthrene had higher bio-remedying possibility than pyrene. Temperature and airflow were important control factors in bio-slurry reactor process. In this research, the optimized parameters were operating temperature as 20-30 degrees C, ratio between water and soil as 2:1, airflow as 8 L.h-1.L-1, and the inoculating amount as 50 g.kg-1.

[Application of terrestrial invertebrates biomarkers in soil pollution ecology study].

Biomarkers in terrestrial invertebrates play an important role in estimating either exposure or resultant effects of pollutants in soil ecosystem, which has received increasing attention and made significant progress. The present paper has drawn three important biomarkers, i.e., lysosomes, stress protein and mettllothioneins (MTs). The lysosomal membrane stability of coelomocytes was assayed as neutral-red retention time (NRR-time) resulting from toxicant stress. Hsp70 and Hsp60 were commonly used in the stress protein families. The quantification methods of different isofoms of the mettllothioneins could specifically indicate different metal contamination. The rationale, characteristics, and relevant exemplary case of assaying the biomarkers and their promising application to ecotoxicological diagnosing in soil contamination were discussed.

[Eco-toxicological effects of heavy metals on the inhibition of seed germination and root elongation of Chinese cabbages in soils].

The Eco-toxicity effects of individual Cu, Zn, Pb and Cd on the inhibition of seed germination and root elongation of Chinese cabbages (Brassica pekimensis) were tested in four types of soils (red loam soils, meadow brown soils, chestnut soils and dark brown soils) and water solution. The combined effects of heavy metals pollution were determined with meadow brown soils. Results indicated that with same concentration, the inhibition rates of heavy metals on root elongation of Chinese cabbages are stronger than that on the seed germination. The inhibition effects of heavy metals on the root elongation of Chinese cabbages in soils are much lower than that in water, indicating that soils play an important role of buffering on heavy metals pollution. Inhibition rates of heavy metals on the root elongation (IRHMRE) of Chinese cabbages are significantly negative related with the contents of organic matter (OR) and Kjedahl-nitrogen (K-N) in soils, however, there is no significant related between IRHMRE and soil pH, so does the content of T-K. In the concentrations that result in the irritated effect in the single form of Cu, Zn, Pb and Cd pollution, synergic effects are produced significantly when four heavy metals are combined. As the results, the threshold values that result in the inhibition effects on root elongation in Chinese cabbages decrease markedly.

[Chlorobenze-stressing injury of the germination of soybean seed].

Effects of various doses of 1,2,4-trichlorobenze on seed germination and seedling growth of soybean were studied. The results indicated that 50-200 micrograms.g-1 of 1,2,4-trichlorobenze had no effect on the percentage of seed germination, but delayed the rate of seedling growth. The seed germination was stopped by 300 micrograms.g-1 of 1,2,4-trichlorobenze. The most activity of SOD took place on the 4th day after sowing of soybean seed in the normal condition, then tended to be in a certain level with a little alteration. Under the various dose stress of 1,2,4-trichlorobenze, the activity of SOD in soybean hypocotyl was decreased, while MDA and protein contents as well as diameter of hypocotyls increased. Seedlings growth, fresh weight(FW), dry weight(DW) were inhibited, while the ratio of FW/DW didn't change. Damage-level of wheat seedlings was increased with increasing 1,2,4-trichlorobenze concentrations. Roots were more sensitive to 1,2,4-trichlorobenze-stress than hypocotyls.

[Effects of chlorobenzene stress on seedling growth and cell division of Vicia faba].

Effects of 1, 2, 4-trichlorobenzene (TCB) stress on seedling growth, cell division and chromosomal aberration frequency of root-tip cells of Vicia faba were studied. The results indicated that the growth of the root length and mitotic index of root tip cells were successively decreased and even stopped with the increase of TCB concentrations and treatment duration. Numerical and structural chromosomal aberrations at metaphase and anaphase of root-tip cells in Vicia faba seedlings were produced by 50-300 micrograms.g-1 TCB treatment for 12-96 h. The percentage of c-mitosis, chromosomal bridge and chromosomal asymmetry array in root tip cells exposed to 50-100 micrograms.g-1 TCB for 12-24 h was up to 1.0-10.3%. The percentage of chromosomal stickness (S), chromosomal stickiness + chromosomal breakage (S + B), chromosomal stickness + chromosomal ring (S + R), chromosomal stickiness + chromosomal asymmetry array (S + A) and chromosomal stickness + chromosomal bridge (S + Be) in root tip cells reached 47.9-88.9%, and 18.1-29.6% for different kinds of chromosomal breakage at 300 micrograms.g-1 TCB for 12-96 h. Thus, the chromosomal aberration of root tip cells in Vicia faba seedlings could be used as a sensitive biomarker of monitoring soil contaminated with TCB.

[Effects of 1,2,4-trichlorobenzene stress on active oxygen in germinated soybean seeds].

Pot culture experiment on the effects of 1,2,4-trichlorobenzene (TCB) stress on active oxygen metabolism of germinated soybean seeds indicated that during the beginning (1st-3rd day) of 100-300 micrograms.g-1 TCB treatment, the respiratory peak appeared earlier, and the respiratory rate, the accumulation of superoxide radical (O2-.) and H2O2, as well as the malondialdehyde (MDA) content were all enhanced. During 1-6 days of TCB stress, peroxidase (POD) activity gradually increased, and superoxide dismutase (SOD) activity increased firstly, and decreased afterward. These results suggested that the injury effects of TCB stress on germinated soybean seeds were achieved by excessive accumulation of active oxygen resulted from the degradation of active oxygen scavenging enzyme system.