[Effect of anatoxin-A on calcium-mediate signal system of brain tissue in mice].

OBJECTIVE: To explore the mechanism of central neurotoxity induced by anatoxin-A (ANTX-A) from the calcium-mediate signal system. METHODS: Mice were administered by i.p. at different doses of ANTX-A for one month, 0.9% of sodium chloride solution was used as the control. Using fluorescence method and colorimetry method, the intracellular calcium concentration and the activity of calcineurin (CaN, PP2B) and ATPase in brain were determined. RESULTS: Calcium concentrations in brain were increased in 12.5, 50 and 200 microg/kg bw groups compared with the control group, the difference had statistical significance (P < 0.01). The activity of PP2B of brain in 200 microg/kg bw group was enhanced compared with the control group (P < 0.05). In all ANTX-A treated groups, the activity of Na+ -K+ -ATPase of brain in mice had no obvious changes, but the activity of Ca2+ -ATPase in 50 and 200 microg/kg bw groups were significantly reduced compared with the control group ( P < 0.05). CONCLUSION: It is suggested that calcium-mediate signal system could have important regulative action in the central neurotoxity induced by antoxin-A.

[Eutrophication and pollution level of microcystin in Dianshan Lake].

A survey was conducted in Dianshan Lake to study the eutrophication indexes including total phosphorus (TP), total nitrogen (TN), pH, temperature, diaphaneity and chlorophyll-a level and dominant algae in seasons. The impacts of temperature, light, nitrogen and phosphorus on growth of and microcystin LR production by Microcystis aeraginosa strain under laboratory conditions were studied. Relationship between algal cell density and concentration of microcystin LR were studied. Results suggest that water in Dianshan Lake was eutrophicated. The suitable seasons for algae growth are the end of spring and summer. The annual average of TP and TN were 1.93 mg/L and 0.18 mg/L respectively. And 93.5 and 92.2 percent of TP and TN were higher than the criteria for the third class water body. Significant impact from agriculture was indicated since the peak of algae laged one month after the maxium use of fertilizer. The dominant algae in Dianshan Lake were cyanobacteria, bacillariophyta, cryptophyta and euglenophyta. Microcystis, anabaena and synedra, which excrete toxins and indicate water pollution, and are dominant algae species in summer. M. aeraginosa strain had a biggest growth rate at temperature of 25 degrees C and light intensity of 3 0001x, while microcystin LR production contents reached maximum at 20 degrees C and 5000lx respectively. The optimum TP and TN concentrations for growth of and toxin production by M. aeraginosa were found to be 650 micromol/L and 6.5 micromol/L respectively. TP is suspected to be the limiting factor for the growth of algae both in field and laboratory conditions. Positive correlations between total microcystin LR concentrations and algae cell density or M. aeraginosa cell densities are found. The algae cell density can be used to predict the level of algal toxins in water.

[Effects of alcohol on the expression of heat shock protein 70 and super microstructure of astrocytes in rat embryonic brain].

OBJECTIVE: To explore effects of alcohol on the expression of heat shock protein 70 (HSP70) and super microstructure of astrocytes. METHODS: Different dosage of alcohol (1, 5, 25, 50mmol/L) was administered to astrocytes of rat embryos. Effects of alcohol on expression of HSP70 were studied by immunocytochemistry, and changes of super microstructure of astrocytes were observed by electron microscope. RESULTS: The results showed that with increase of alcohol dosage, the expression of HSP70 increased, hepatin granule and survival astrocytes in 25mmol/L and 50mmol/L dosage group decreased, cell membrane and mitochondrion were damaged, and even apoptosis and necrosis occurred. CONCLUSION: It suggested that alcohol could increase expression of HSP of astrocytes, which could depress the expression of normal function protein, and mainly impaired cell membrane and mitochondrion, which could lead to apoptosis and necrosis.

[The immediate early genes expression in the PC12 cells activated by anatoxin-a].

OBJECTIVE: To explore the regulative action of IEGs when the nicotinic acetylcholine receptors (nAChRs) of PC12 cells were activated by anatoxin-a. METHODS: Using the reverse transcription polymerase chain reaction (RT-PCR) method, the mRNA gene expression of c-fos, c-jun, NGFI-A and NGFI-B were measured while PC12 cells were activated by anatoxin-a. RESULTS: As 10(-9), 10(-8) and 10(-7) mol/L anatoxin-a activated PC12 cells for an hour, or 10(-7) mol/ L anatoxin-a activated PC12 cells for 30 min, 60 min, 120 min, the intracellular gene expression of c-fos and NGFI-A increased significantly than the control group (P < 0.05), it was 2-6 times than the control group. And the gene expression of c-fos presented dose-response and time-effect relation. However, under the same condition, the gene expression of c-jun and NGFI-B did not show any remarkable changes. CONCLUSION: c-fos and NGFI-A might be involved to modulation the action of anatoxin-a activating the nAChRs of PC12 cells.

Confirmation of an acute no-observed-adverse-effect and low-observed-adverse-effect level for copper in bottled drinking water in a multi-site international study.

In a double blind, 3x3 factorial (volumexdose) study, 70 adult females (18-60 years of age) at four different international sites (total pooled n=269) were given 100, 150, or 200ml of bottled drinking water with 0.4, 0.8, or 1.2mg of copper (Cu) as the sulfate salt once each week. Two additional doses (0 and 1.6mg Cu) were added at the 200ml volume to determine a dose-response relationship and corroborate previously reported results. All subjects completed a questionnaire at 0, 0.25, and 1h post-dosing that screened for positive gastrointestinal (GI) effects (nausea, vomiting, abdominal pain, and diarrhea). Nausea was the most prevalent symptom reported and was generally reported within the first 15min (water volume, p<0.032; copper dose, p<0.0001; and water volumexcopper interaction, p<0.97). As volume increased, the effect of Cu-induced nausea decreased; as Cu dose increased, the incidence of nausea increased. At 200ml, a significant increase in reported incidence of nausea at 0.25h occurred at 1.2 mg Cu (6mg Cu/L), indicating a NOAEL of 0.8mg Cu (4mg Cu/L) for adult females. These data confirm a previously determined human acute NOAEL for Cu added to distilled water, and provide additional, controlled human data for determining safe concentrations of Cu in drinking water.

[Distribution of microcystin-LR and anatoxin-A and their influence factors in Dianshan Lake during summer and autumn].

Six different sampling locations in Dianshan Lake were selected to study the distributions of algaes and algae toxins and their influencing factors in summer and autumn. Total nitrogen (TN), total phosphorous (TP), Fe, chlorophyla(Chla) and COD in water samples were determined. At the same time, water temperature (TEM), pH value, illumination, turbidity degree (TUR) of water body were recorded. In addition, the total number and attribute of algae cell and the content of microcystin-LR(MC-LR) and anatoxin-A(ANTX-A) were checked. The results showed that the pollution of both toxins was most serious in July. The MC-LR were mostly distributed in Wangxiang, but the ANTX-A were mostly distributed in Jishuigang. Generalized estimating equations (GEE) regression analysis showed that total number of algae cell was significantly correlated with TN, TP and Chla (P < 0.05). The distribution of MC-LR was significantly affected by total number of algae cells and TEM (P < 0.01), but TUR mainly affected the distributions of ANTX-A(P < 0.01). There was weak correlation between both toxins (P < 0.05). In short, there was a serious pollution of MC-LR, but ANTX-A pollution was light. The factors influencing the distributions of both algae toxins were not completely consistent.

The carcinogenic potential of extractable organic matter from urban airborne particles in Shanghai, China.

The genotoxicity of extractable organic matter (EOM) from airborne particles in Shanghai has been determined using short-term bioassays. EOM samples were investigated using cell morphological transformation and two-stage model of mouse skin tumorigenicity assays to detect their carcinogenic activity. DNA adducts were detected using the 32P-postlabeling technique. The results showed that EOMs induced cell morphological transformation and played a role in tumor-initiating carcinogenesis. The EOMs of airborne particles from different districts of Shanghai had similar carcinogenic activity except the result of sample E (at downtown of Shanghai) was relatively high. The polycyclic aromatic hydrocarbon (PAH) fraction makes a major contribution to carcinogenic activity according to the results of cell morphological transformation assay. DNA adducts were also detected in skin, liver, and kidney of mouse after treatment with EOMs. It is suggested that the urban airborne particles in Shanghai, which show carcinogenic potential and genotoxic activity in our bioassays, may be responsible for the increased incidence of lung cancer in Shanghai in last few years.

[Study on guideline of alga in source water].

Toxic bloom in the sources of drinkable water has been a public health problem. Hindered by the complicated analysis procedure, very limited efforts has been done in investigating toxin level. Patronized by the Chinese Academy of Preventive Medicine, a study on alga hygiene criteria of drinkable water sources in China was conducted. Available literature was reviewed and ecological research and toxicological tests were carried out. The criteria on alga biomass in drinkable water source was formulated by referring the criteria of microcystin--LR in drinking water recommended by WHO and Ministry of Health, China. Three kinds of criteria were worked out: primary criteria, which is to ensure safety, is 1.0 x 10(4) per liter, secondary criteria, which should be alert, is 2.1 x 10(5) per liter, and third--rate criteria, which might be toxic, is 1.2 x 10(6) per liter. It is suggested that these criteria are valuable and feasible in routine work.

[Study on the unscheduled DNA synthesis in primary rat hepatocyte induced by source water, tap water and active carbon-treated tap water].

To study the mutagenicity of source water with different turbidities, tap water and active carbon-treated tap water, unscheduled DNA synthesis in primary rat hepatocyte was detected. Increased unscheduled DNA synthesis was found in a dose-depend manner inducted by tap water, source water and tap water treated with 5, 10 and 15 milligrammes active carbon per liter water. The results also showed that the mutagenicity of the source water increased with the increasing of its turbidity, and mutagenicity of the tap water is higher than that of the source water as well. There was no unscheduled DNA synthesis in tap water followed by the treatment with 20 milligrammes active carbon per liter water. It is suggested that the development of the water treatment technics including sedimentation, filtration and treatment with active carbon is needed for decreasing or eliminating the mutagenicity of tap water.

[Effects of microcystins on cell cycle and expressions of c-fos and c-jun].

OBJECTIVES: To investigate the effects of microcystins on cell cycle and expressions of c-fos and c-jun, and explore the potential carcinogenic mechanisms of Microcystins. METHODS: Microcystic cyanobacteria extraction (MCE) purified by Sep-Pak C(18) cartridge was added into the media and co-incubated with SHE cell for various periods. Immunohistochemistry assay was applied to detect the expressions of c-fos and c-jun at 1, 3, 6 hr time point, and cell cycle at 6, 12, 24 hr point were analyzed by flow cytometry respectively. RESULTS: Sustained up-regulated expression of c-fos and c-jun were induced by MCE during the experimental period, and 5 - 6 folds increased expression were observed at 6 hour point after treatment. As much as 44.8 per cent of cells were induced to entry S-phase from resting G(0)/G(1). CONCLUSION: Up-regulating the expression of transcript factor such as c-fos and c-jun thus to induce cell abnormal proliferation may be the potential carcinogenic mechanisms of microcystins.

[Study on the estrogenic effect of the organic extracts from tap water and source water].

In order to study the estrogenic effects of the organic extracts from tap water and source water, two in vitro assay and one in vivo short-term assay were performed to detect the estrogen-like activity of the component of water. The results showed that at the concentration of 50 ml relevant to real water, the recombinant yeast assay and MCF-7 cell proliferation assay could both detected the estrogen-like activity and with increase of concentration, the component exhibited an increasing estrogenic effects. The dose inducing the maximum effect in cell proliferation assay was 250 ml and in recombinant yeast assay was 1250 ml. Exceed this dose with increasing concentration, the estrogenic effects of component decreased both in two assay. In uterrtropic assay, doses of 10 L/kg and 50 L/kg increased the weight of uterus of 18 day ICR female mice significantly. All above findings indicated that organic extracts from source water exhibited the estrogenic effects. The series of 3 bioassay process displayed sensitivity and practicability.

Genotoxic activity of extractable organic matter from urban airborne particles in Shanghai, China.

The aim of this research is to investigate the impact of air pollution on the population in Shanghai. The genotoxicity of extractable organic matter (EOM) from the air particles was investigated by the means of the Salmonella plate incorporation assay, rat hepatocyte unscheduled DNA repair assay, and mice micronuclei test. The airborne particles were collected in 13 locations during the summer of 1992 and winter of 1993. The crude extracts were fractionated by acid-base partitioning into acid, base and neutral fractions. The neutral fractions were further fractionated by resin-silica gel column chromatography into three subfractions. The induction of revertants with the crude extracts was higher in winter samples than in summer samples. Both indirect-acting and direct-acting mutagenicity were observed. The mutagenicity was detected with TA98, but was not detected with TA100. The mutagenic activity was the greatest in the acid, aromatic and polar fractions from summer samples. The fractions from the winter samples did not show clear differences. There was no substantial location-related variance in the mutagenic potencies of EOM, but substantial location- or time-related variances in the mutagenic potencies of the airborne particles per cubic meter air were found. While rat hepatocyte unscheduled DNA synthesis (UDS) assay revealed genotoxicity for all the samples, there was no big variance in the genotoxicity of the fractions. The mouse micronuclei test showed results similar to the UDS assay. The difference of locality did not have statistical significance.

[Sub-acute hepatotoxicity of low doses of microcystins].

In order to study sub-acute hepatotoxicity of low doses of microcystins in vivo, as well as to understand the mechanisms of hepatotoxicity of microcystins, eighty Spague-Dawley rats were injected with microcystins intraperitoneally at the doses of 0, 4, 8 and 12 micrograms.(kg.d)-1, respectively, for 35 days. Then blood and liver samples were used for assay. Several enzymatic levels and pathological changes were detected. Both terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) and immunohistochemical methods were employed to study the apoptosis and proliferating cell nuclear antigen (PCNA). It was shown that the activity of serum gamma-glutamyltransferase (GGT) and concentrations of whole blood glutathione (GSH) decreased, serum activities of lactate dehydrogenase (LDH) and aminotransferase (AST) increased after exposure to MC. No significant change of concentration of serum alanine aminotransferase (ALT) was observed in the tested groups. Characteristic morphological alterations and active proliferation as well as apoptosis of hepatotocytes were observed in the treated groups. It was suggested that oxidative injury and apoptosis of hepatocytes induced by microcystins may be the mechanisms of its hepatotoxicity.