Effects of Toluene on the Development of the Inner Ear and Lateral Line Sensory System of Zebrafish.

OBJECTIVE: The aim of this study was to explore the ototoxicity of toluene in the early development of zebrafish embryos/larvae. METHODS: Zebrafish were utilized to explore the ototoxicity of toluene. Locomotion analysis, immunofluorescence, and qPCR were used to understand the phenotypes and molecular mechanisms of toluene ototoxicity. RESULTS: The results demonstrated that at 2 mmol/L, toluene induced zebrafish larvae death at 120 hours post fertilization (hpf) at a rate of 25.79% and inhibited the rate of hatching at 72 hpf. Furthermore, toluene exposure inhibited the distance travelled and average swimming velocity of zebrafish larvae while increasing the frequency of movements. As shown by fluorescence staining of hair cells, toluene inhibited the formation of lateral line neuromasts and middle line 1 (Ml 1) neuromasts in 3 days post fertilization larvae in a concentration-dependent manner. Toluene altered the expression level of genes involved in ear development/function in zebrafish, among which the mRNA levels of cd164l2, tekt3, and pcsk5a were upregulated, while the level of otofb was downregulated, according to the qPCR results. CONCLUSION: This study indicated that toluene may affect the development of both the inner ear and lateral line systems in zebrafish, while the lateral line system may be more sensitive to toluene than the inner ear.

Methamphetamine induces hepatotoxicity via inhibiting cell division, arresting cell cycle and activating apoptosis: In vivo and in vitro studies.

Methamphetamine (METH) resulted in acute hepatic injury. However, the underlying mechanisms have not been fully clarified. In the present study, rats were treated with METH (15 mg/kg B.W.) for 8 injections (i.p.), and the levels of alanine transaminase, asparatate transaminase and ammonia in serum were significantly elevated over those in the control group, suggesting hepatic injury, which was evidenced by histopathological observation. Analysis of the liver tissues with microarray revealed differential expressions of a total of 332 genes in METH-treated rats. According to the GO and KEGG annotations, a large number of down-regulated cell cycle genes were screened out, suggesting that METH induced cell cycle arrest and deficient of cell cycle checkpoint. Related genes and proteins were confirmed by RT-qPCR and western blotting in rat livers, respectively. Moreover, treatment of Brl-3A cells with METH caused significant cytotoxic response and marked cell cycle arrest. Furthermore, overexpressions of Cidea, cleaved caspase 3 and PARP 1 in METH-treated rats indicated activation of apoptosis, while its inhibition alleviated cell death in Brl-3A cells, suggesting that activation of apoptosis took an important role in METH-induced hepatotoxicity. Taken together, the present study demonstrates that METH induced hepatotoxicity via inducing cell cycle arrest and activating apoptosis.

Sodium butyrate induces apoptosis of human colon cancer cells by modulating ERK and sphingosine kinase 2.

OBJECTIVE: To investigate the role of extracellular signal-regulated kinase (ERK) in apoptosis of human colon cancer (HCT116) cells. METHODS: After the HCT116 cells were pretreated with specific ERK inhibitor (U0126) or specific siRNA and exposed to 10 mmol/L sodium butyrate (NaBT) for 24 h, their apoptosis was detected by flow cytometry, levels of SphK2 and ERK protein were measured by Western blot, and translocation of SphK2 was assayed by immunofluorescence microscopy. RESULTS: The U0126 and siRNAs specific for SphK2 blocked the export of SphK2 from nuclei to cytoplasm and increased the apoptosis of HCT116 cells following NaBT exposure. Over-expression of PKD decreased NaBT-induced apoptosis of HCT116 cells, which was reversed by U0126. Furthermore, transfection of HCT116 cells with constitutively activated PKD plasmids recovered the U0126-blocked export of SphK2. CONCLUSION: ERK regulates the export of SphK2 and apoptosis of HCT116 cells by modulating PKD. Modulation of these molecules may help increase the sensitivity of colon cancer cells to the physiologic anti-colon cancer agent, NaBT.

An improved liquid scintillation counting method for the determination of gross alpha activity in groundwater wells.

A liquid scintillation counting (LSC) method having several advantages over the gas proportional counting (GPC) U.S. Environmental Protection Agency (EPA) Method 900.0 for the detection of gross alpha activity in drinking water was evaluated in this study. The improved method described here involves the use of nitromethane as the quench agent for establishing counting efficiencies and spillover factors, and it minimizes sample preparation. It has the advantage of achieving the regulatory detection limit of 111 mBq L(-1) with short count times (100 min) and small sample aliquot sizes. A thorough method validation study was performed by testing field samples ranging in total dissolved solids (TDS) from 0.3 mg L(-1) to 1,000 mg L(-1) and spiking each matrix from 194 mBq L(-1) to 11.6 Bq L(-1). Comparable method precision and accuracy was observed on the two types of LSC instruments tested, Perkin Elmer Quantulus 1220 and Packard 2550, with the former giving better performance. Data presented demonstrate that this efficient and high throughput LSC method is suitable for groundwater samples in excess of 1,000 mg L(-1) of TDS in contrast with the 500 mg L(-1) limit by the routine GPC method. Groundwater wells across the state of California were sampled, analyzed for gross alpha activity using the EPA- approved method and the improved LSC method, and the results were compared.

Occurrence and distribution of 210Pb and 210Po in selected California groundwater wells.

Groundwater wells from across the State of California were sampled and analyzed for Pb and Po. The separation method involved Fe(OH)3 precipitation from a 5-L groundwater sample followed by electrodeposition of Po on a nickel disk. The resulting solution was passed through an ion-exchange resin column for the isolation of Pb. De-ionized water spiked at a concentration range from 4.92 mBq L(-1) to 755 mBq L(-1) with these radionuclide standards showed excellent accuracy and precision of the method. In the groundwater wells, overall activity of Pb ranged from 3.7 mBq L(-1) to 1,481 mBq L(-1) and the Po activity ranged from 0.25 mBq L(-1) to 555 mBq L(-1). Of the select wells tested, 27% for Pb and 19% for Po were above the proposed maximum contamination limits for these radionuclides, which are set at 37 mBq L(-1) and 26 mBq L(-1), respectively. From a public health perspective this is a concern, since the drinking water screening levels for gross alpha is at 555 mBq L(-1) and gross beta is at 1,850 mBq L(-1). At such high screening levels Pb and Po will not be captured, and this situation was found in several of the wells studied. The occurrence of Pb and Po are not correlated within the sources, however; the polonium concentrations were always lower than the lead concentrations. Activities of Pb measured from wells two years apart clearly demonstrated the continuous flux of groundwater within aquifers.

Occurrence of 224Ra, 226Ra, 228Ra, gross alpha, and uranium in California groundwater.

One hundred and twelve groundwater wells sampled from all the major aquifers in California were analyzed for 224Ra, 226Ra, 228Ra, gross alpha, and uranium. The results showed that radium is found in relatively low concentration, 1.56 x 10(-2)-1.23 Bq L(-1) (0.42-33 pCi L(-1)) for 224Ra, 2.2 x 10(-3)-0.81 Bq L(-1) (0.06-22 pCi L(-1)) for 226Ra, and 8.5 x 10(-3)-1.31 Bq L(-1) (0.23-35 pCi L(-1)) for 228Ra in California groundwater. Uranium was found at the highest concentration on both mass and activity basis and was correlated with the gross alpha measurement. Short-lived radioisotopes showed no significant contribution to gross alpha measurements. There was a strong correlation between 224Ra and 228Ra activities, suggesting the latter to be an indicator for the occurrence of the former. Comparison of 226Ra to 238U, 224Ra to 226Ra, and 226Ra to 228Ra showed scattered data indicating no correlation between each of these isotope pairs. Approximately 4% of the wells were found to exceed the U.S. Environmental Protection Agency (EPA) established maximum contaminant level for total radium of 0.185 Bq L(-1) (5 pCi L(-1)). Analysis of 228Ra by gamma-ray spectroscopy was in good agreement with the U.S. EPA-approved procedure.

Receptor binding assay for paralytic shellfish poisoning toxins: optimization and interlaboratory comparison.

A receptor binding assay (RBA) for detection of paralytic shellfish poisoning (PSP) toxins was formatted for use in a high throughput detection system using microplate scintillation counting. The RBA technology was transferred from the National Ocean Service, which uses a Wallac TriLux 1450 MicroBeta microplate scintillation counter, to the California Department of Health Services, which uses a Packard TopCount scintillation counter. Due to differences in the detector arrangement between these 2 counters, markedly different counting efficiencies were exhibited, requiring optimization of the RBA protocol for the TopCount instrument. Precision, accuracy, and sensitivity [limit of detection = 0.2 microg saxitoxin (STX) equiv/100 g shellfish tissue] of the modified protocol were equivalent to those of the original protocol. The RBA robustness and adaptability were demonstrated by an interlaboratory study, in which STX concentrations in shellfish generated by the TopCount were consistent with MicroBeta-derived values. Comparison of STX reference standards obtained from the U.S. Food and Drug Administration and the National Research Council, Canada, showed no observable differences. This study confirms the RBA's value as a rapid, high throughput screen prior to testing by the conventional mouse bioassay (MBA) and its suitability for providing an early warning of increasing PSP toxicity when toxin levels are below the MBA limit of detection.