Ali B. Ishaque

Lead-induced cytotoxicity and transcriptional activation of stress genes in human liver carcinoma (HepG2) cells.

Lead is a non-essential element that exhibits a high degree of toxicity, especially in children. Most research on lead has focused on its effects on organ systems such as the nervous system, the red blood cells, and the kidneys which are considered to be the primary targets of lead toxicity. However, the molecular mechanisms by which it induces toxicity, and carcinogenesis remain to be elucidated. In this research, we performed the MTT assay to assess the cytotoxicity, and the CAT-Tox assay to assess the transcriptional responses associated with lead exposure to thirteen different recombinant cell lines generated from human liver carcinoma cells (HepG2), by creating stable transfectants of mammalian promoter chloramphenicol (CAT) gene fusions. Study results indicated that lead nitrate is cytotoxic to HepG2 cells, showing LD50 values of 49.0 ± 18.0 mgr;g/mL, 37.5 ± 9.2 μg/mL, and 3.5 ± 0.7 μg/mL for cell mortality upon 24, 48 and 72 h of exposure, respectively; indicating a dose- and time-dependent response with regard to the cytotoxic effect of lead nitrate. A dose-response relationship was also recorded with respect to the induction of stress genes in HepG2 cells exposed to lead nitrate. Overall, six out of the thirteen recombinant cell lines tested showed inductions to statistically significant levels (p < 0.05). At 50 μg/mL of lead nitrate, the average fold inductions were: 2.1 ± 1.0, 5.4 ± 0.4, 12.1 ± 6.2, 5.0 ± 1.7, 2.5 ± 1.3, and 4.8 ± 4.5 for XRE, HSP70, CRE, GADD153, and GRP78, respectively. These results indicate the potential for lead nitrate to undergo biotransformation in the liver (XRE), to cause cell proliferation (c-fos), protein damage (HSP70, GRP78), metabolic perturbation (CRE), and growth arrest and DNA damage (GADD153). Marginal but not significant inductions were also obtained with the GSTYa (1.5 ± 0.8), and GADD45 (5.7 ± 8.1) promoters, and the NF-κB (2.0 ± 1.7) response element, indicating the potential for oxidative stress. No significant inductions (p > 0.05) were recorded for CYP1A1, HMTIIA, p53RE, and RARE.

Environmental toxicology and health effects associated with dinitrotoluene exposure

Dinitrotoluenes (DNTs) are nitroaromatic compounds appearing as pale yellow crystalline solids at room temperature. Dinitrotoluenes exist as a mixture of 2 to 6 isomers, with 2,4-DNT, and 2,6-DNT being the most significant. About 500 persons are estimated to be potentially exposed yearly to 2,4-DNT and 2,6-DNT during the production of munitions and explosives. The main route of human exposure at ammunition facilities is inhalation, but dermal contact and inadvertent ingestion can also be substantial. In factory workers, exposure to DNTs has been linked to many adverse health effects, including cyanosis, vertigo, headache, metallic taste, dyspnea, weakness and lassitude, loss of appetite, nausea, and vomiting. Other symptoms including pain or parasthesia in extremities, abdominal discomfort, tremors, paralysis, chest pain, and unconsciousness have also been reported. The primary targets of DNT toxicity are the hematopoietic system (pallor, cyanosis, anemia, and leukocytosis), the cardiovascular system (ischemic heart disease), the nervous system (muscular weakness, headache, dizziness, nausea, insomnia, and tingling pains in the extremities) and the reproductive system (reduction of sperm counts, alteration of sperm morphology, and aspermatogenesis). An association between DNT exposure and increased risk of hepatocellular carcinomas and subcutaneous tumors in rats, as well as renal tumors in mice, has been established. Epidemiologic studies of DNT toxicity have been limited to small groups of workers who had been occupationally exposed at various ammunitions production facilities. Clearly defining the health effects of DNTs with a high degree of confidence has therefore been difficult because of the multigenic nature of occupational exposure. In an attempt to update the toxicologic profile of the DNTs, we hereby provide a critical review of the environmental and toxicologic pathology of DNTs, with a special emphasis on their potential implications for public health.

The Kinetic Signature of Toxicity of Four Heavy Metals and Their Mixtures on MCF7 Breast Cancer Cell Line

This study evaluated the kinetic signature of toxicity of four heavy metals known to cause severe health and environmental issues—cadmium (Cd), mercury (Hg) lead (Pb) arsenic (As)—and the mixture of all four metals (Mix) on MCF7 cancer cells, in the presence and absence of the antioxidant glutathione (GSH). The study was carried out using real time cell electronic sensing (RT-CES). RT-CES monitors in real time the electrical impedance changes at the electrode/culture medium interface due to the number of adhered cells, which is used as an index of cell viability. Cells were seeded for 24 h before exposure to the metals and their mixtures. The results showed that in the presence and absence of cellular glutathione, arsenic was the most cytotoxic of all five treatments, inducing cell death after 5 h of exposure. Lead was the least cytotoxic in both scenarios. In the presence of cellular GSH, the cytotoxic trend was As > Cd > MIX > Hg > Pb, while in the absence of GSH, the cytotoxic trend was As > Hg > MIX > Cd > Pb. The findings from this study indicate the significance of glutathione-mediated toxicity of the metals examined—particularly for mercury—and may be clinically relevant for disorders such as autism spectrum disorder where decreased glutathione-based detoxification capacity is associated with increased mercury intoxication.

Land Use–Land Cover Changes in the Lower Eastern Shore Watersheds and Coastal Bays of Maryland: 1986–2006

Abstract Changes in land use influence surface-water quality and thus present a potential threat to coastal ecosystem health. Land use–land cover changes (LULCC) in the lower Eastern Shore watersheds of Maryland have been rapid in the last decade, with increase in real estate development an obvious indicator. The objective of this study was to evaluate the extent of historical LULCC in the lower Eastern Shore watershed and coastal bays of Maryland from 1986 to 2006. Land use–land cover data were derived by supervised classification of Landsat TM 5 satellite imagery acquired in 1986, 1996, and 2006 using the Anderson level-1 classification system in Environment for Visualizing Images (ENVI 4.5), while LULCCs were detected in an Arc-GIS 9.2 environment. The results showed that while urban and forest lands increased by 121.8% and 8.5%, respectively, in the lower Eastern Shore from 1986 to 2006, croplands and wetlands decreased by 19.6% and 21.3%, respectively. Area covered by surface water increased by 10%, submerging mostly wetlands of 150 km2 in 17 of the 23 subwatersheds studied. The loss of these coastal wetlands is attributable in part to the changing climate and the resultant sea-level rise and in part to the activities of the invasive rodent Nutria (Myocastor coypus), reported to be a major menace in Marylands Eastern Shore. The declining wetlands have serious ecological health implications for the Chesapeake Bay and its watersheds for several species and thus require urgent attention. More intensive and frequent monitoring of this delicate estuarine ecosystem is suggested.

Land Use–Land Cover Changes and Sewage Loading in the Lower Eastern Shore Watersheds and Coastal Bays of Maryland: Implications for Surface Water Quality

ABSTRACT Aighewi, I.T.; Nosakhare, O.K., and Ishaque, A.B., 2013. Land use–land cover changes and sewage loading in the lower Eastern Shore watersheds and coastal bays of Maryland: Implications for surface water quality. Changes in land use and cover influence surface water quality and thus are a potential threat to water systems and coastal ecosystem health. The objectives of this study were to evaluate the influence of historical land use–land cover (LULC) changes and point-source sewage discharge on surface water quality of some lower Eastern Shore watersheds of Maryland. LANDSAT data for 1986–2006 was acquired and classified using Anderson level-1 classification system in ENVI 4.5, whereas LULC changes were detected in Arc-GIS 9.2 environment. Historical water monitoring and climatic data were obtained from Maryland Department of Environment–Chesapeake Bay Program, and National Oceanic and Atmospheric Administration, respectively. Nutrient loading data from wastewater treatment plants were obtained from Maryland Department of Environment and data analyzed using regression analysis and principal component analysis (PCA). The result shows that total phosphorous levels in the surface waters decreased significantly (p < 0.05) during the 20-year study period. However, using PCA, we determined that the declining P trend was attributable to the decreasing agricultural land use rather than the sewage discharge from wastewater treatment plants. The increase in urban land use and the resultant runoff to the water bodies may explain the declining trends in dissolved oxygen levels observed during the period, with serious implications for eutrophication. An empirical model developed for P in surface waters (r2  =  0.90) showed that the combination of point and nonpoint sources and land use/cover change factors are good predictors. The incorporation of quantitative LULC data component as well as point-source nutrient loading into existing models is highly recommended for a more holistic assessment of land-use influence on water systems in general.

Differential Toxicological Interaction among Arsenic, Cadmium, Lead, and Mercury on MCF 7 Cell Line

Evaluation of joint toxic action of metal ion mixtures is one of the priority research areas due to the simultaneous occurrence of metals in the environment and the health risk they posed to humans and the environment as a mixture. Individual and composite mixture acute toxicities of arsenic (As), cadmium (Cd), mercury (Hg), and lead (Pb), which are among the top eight toxic chemicals, were characterized at varying concentrations. MCF 7 cell lines were exposed to individual and composite mixtures containing the four metal ions in the proportion of their EPAs MCL for 24 hours, and the concentration-response data were generated spectrofluorometrically. Acute toxicities were estimated based on the uptake of fluorescence diacetate dye. Toxicological interactions among the four metals were profiled, based on computed interactive index. Results demonstrated that the toxicity of each of the metal ions was enhanced in the composite mixture, and the metals demonstrated differential interactions in a concentration dependent manner. Lead, the least toxic among the four metals, showed the highest enhancement (23-to 64-fold) in toxicity when in the mixture. Interaction among the four metals was largely additive although there was slight departures form additivity at the two extremes of the concentration range.

Separate and Joint Effects of Polycyclic Aromatic Hydrocarbons (PAH)and Polychlorinated Biphenyls (PCB) on Aromatase CYP19A TranscriptionLevel in Atlantic Tomcod (Microgradus tomcod)

Ovarian cytochrome 19A (CYP19A) expression is recognized as a useful biomarker for exposure of fish to environmental contaminants such as PAHs and PCBs. In this study, a laboratory approach using Atlantic tomcod (Microgradus tomcod) from the Hudson River was used to evaluate the additive and interactive effects of a PAH (benzo[a]pyrine) and a PCB mixture (Aroclor 1242) with respect to their effects on various metrics of reproduction. The experimental design was a two-way factorial with each treatment at 0, 0.1 and 1 ppm and replicated three times. Fish embryos were subjected to a short term aqueous exposure (DMSO as vehicle) whereas larvae were exposed via repeated feedings of contaminated prey (Artemia). Quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to evaluate the mRNA expression level of the ovarian aromatase CYP19A as a biomarker to assess reproductive stress in Microgadus tomcod. When tested alone, expression of aromatase CYP19A was significantly up regulated at the higher level of PCBs but no effect was observed from benzo-a-pyrene (B[a]P). In the PCB/PAH combined treatment, both low-PCB with both low/high levels of PAH and high-PCB with both low/high levels of PAH treatment groups had no significant effect on aromatase CYP19A transcript levels compared with the control group. Gonadosomatic Index (GSI) of reproductively mature females showed that only the high-PCB treatment group exhibited significant gonadal loss. The results confirm the expectation that transcription of both PCB and PAH responsive genes are upregulated since they both exert their toxic effects through the aryl hydrocarbons receptor (AHR) pathway

The Effect of L-Buthionine Sulfoximine on the Toxicities and Interactions of As, Cd, Hg and Pb and their Composite Mixture on MCF 7 Cell Line

The effect of intracellular level of GSH on the cytotoxicity and interaction of four environmentally relevant metals arsenic, cadmium, mercury and lead (As, Cd, Hg, and Pb) was investigated. LButhionine Sulfoximine (LBSO) was used to inhibit the intracellular level of GSH in MCF 7 cells. Both individual and combined cytotoxicities of the four metals on the cells were assayed by spectrofluorometric counting of the surviving cells after 24-hour exposure. Exposure of the cells to three of the studied metals: As, Cd, and Hg resulted in the production of significantly (p<0.5) higher level of cellular GSH relative to the control. However, cells exposed to Pb with or without pretreatment with LBSO exhibited about 50% decrease in cellular GSH. Individual metal toxicity was higher in GSH-depleted cells relative to GSH-rich cells; However, the effect of GSH depletion was slightly metal selective as As and Hg exhibited toxicities. Cells exposed to the composite mixture of all four metals indicated additive and antagonistic interactions in GSH depleted cells and GSH rich respectively.