Genotoxicity in lead treated human lymphocytes evaluated by micronucleus and comet assays.

Lead (Pb) which plays a significant role in modern industry is related to a broad range of physiological, biochemical, behavioural and genetical dysfunctions. Its exposure leads to an increased frequency of genetic aberrations in humans. Hence, this study was designed to assess the genotoxic effect of lead acetate at three dosage levels (10, 25 and 50 µg/mL) by employing: the Cytokinesis Block Micronucleus (CBMN) assay and the Comet assay in Peripheral Blood Lymphocyte Cultures. The results of this study revealed an increased level of DNA damage among treated groups. A significant increase in the tail length of comets and other indices was observed at 25 and 50 µg/mL concentrations comparatively. Thus, lead acetate induced single-strand breaks (SSB) and double strand breaks (DSB) in DNA, alkali-labile sites (ALS), oxidative DNA damage as well as DNA-DNA/DNA-protein/DNA-metal cross linking as evidenced by the Comet assay. The chromosome breakage, DNA misrepair, chromosome loss and telomere end fusion were determined by the Micronucleus assay. Micronucleus frequency in treated lymphocytes was significantly higher as compared to controls. Nucleoplasmic bridges increased significantly and Nuclear buds increased at higher two doses only in exposed cultures. Thus, these assays are better indices for lead induced genotoxicity and metal-nucleus interactions.

Experimental studies on insecticides commonly used in India.

Using hexachlorocyclohexane (BHC) as a model histopathological, histoenzymological, biochemical, and electrophoretic studies were undertaken to find out certain parameters for early diagnosis of liver cancer. In addition, cytogenetic studies were carried out to evaluate the effect of BHC feeding on mitotic and meiotic divisions. The results of these investigations suggest that there is a significant change in liver weight in experimental group. Histologically, liver cells follow a definite sequential cellular alteration ultimately leading to liver tumor. Histochemically, well defined pattern of glycogen accumulation and iron distribution in hepatocytes was observed. The electron-microscopic observation demonstrated prominently the proliferation of agranular endoplasmic reticulum in early stages. The distribution of certain enzymes linked with plasma membrane, lysosomes, and mitochondria showed the functional alteration of these organelles both in neoplastic nodules and tumours induced by BHC. The biochemical changes observed in gluconeogenic enzymes (G6Pase and F1,6dipase) and dehydrogenases (LDH, ICDH, and MDH) at different duration of exposure to BHC indicated decrease in enzyme activity of both gluconeogenic pathway and tricarboxylic acid cycle, linked with energy metabolism. These changes tend to recover with discontinuation of BHC but 8 months continuous feeding produces irreversible changes in G6Pase activity. Using polyacrylamide gel electrophoresis technique a change in serum proteins and LDH isoenzymes was observed. However, extrapolation of these findings to human situation needs more extensive studies, taking into account all possible variables, such as the DDT and BHC load in our environment and the body burden resulting there from.

Early changes in serum protein and liver LDH isoenzymes in mice exposed to technical grade hexachlorocyclohexane (BHC) and their possible relationship to liver tumours.

Mice were exposed to hexachlorocyclohexane (BHC) in order to study the changes in the serum protein pattern and in the LDH isoenzymes of the liver. After 2 months of exposure the protein pattern showed a new band which persisted even after the development of a tumour. The LDH isoenzymes pattern showed a gradual decrease of the faster moving LDH-1 and LDH-2 bands which later disappeared completely when hepatic tumours formed. The significance of these results is discussed.

Dominant-lethal study of technical-grade hexachlorocyclohexane in Swiss mice.

Male Swiss mice, 6-8 weeks old, were given a diet containing technical-grade hexachlorocyclohexane (BHC) at 500 ppm continuously for 4, 6 and 8 months. After the completion of the scheduled exposure period, the males were sequentially mated with 2-3 untreated virgin females at weekly intervals for 8 weeks. The females were autopsied at mid-term pregnancy for evaluation of dominant-lethal mutation. The number of dead implants, including deciduomas and dead embryos, showed a significant increase. Similarly, the percentage fertility and live embryos per female showed a decline when compared with the control

Studies on glucose-6-phosphatase, fructose-1,6-diphosphatase activity, glycogen distribution and endoplasmic reticulum changes during hexachlorocyclohexane induced hepatocarcinogenesis in pure inbred Swiss mice.

Inbred Swiss mice were fed hexachlorocyclohexane (BHC) at 500 ppm dose level in diet for 2, 4, 6 and 8 months. Later BHC was discontinued for 4 months and subsequently the animals were refed BHC for 1 month. Glucose-6-phosphatase (G6Pase) and fructose-1,6-diphosphatase (FDPase) activity was studied at different time intervals accompanied with changes in glycogen distribution and endoplasmic reticulum (ER) proliferation in hepatocytes. G6Pase and FDPase showed a decline in activities on BHC feeding. The activities of these enzymes showed recovery on BHC discontinuation. The changes were progressive with duration of exposure. After 6 months exposure the biochemical changes became more resistant to recovery. Maximal changes occurred in 8 month-exposure and the changes were irreversible. Glycogen accumulation and depletion followed a definite pattern. After two months of BHC feeding, increase in parenchymal glycogen storage zones was observed. In the later stage of hepatocarcinogenesis and specially in tumors, glycogen was depleted considerably. Smooth endoplasmic reticulum (SER) proliferation was recorded around the 3rd and 4th month. The correlation between glycogen accumulation, SER proliferation, G6Pase and FDPase activity is discussed.

Distribution of isomers of BHC and related histopathology of liver in hexachlorocyclohexane (technical grade BHC) fed mice.

In an attempt to correlate body burden and related liver histopathology after exposure to 500 ppm benzene hexachloride, 6- to 8-wk-old male and female mice were fed benzene hexachloride with their mash diet for a 6-month period. The mice were divided into 8 groups (12/group; 6 experimental, 6 control) and were sacrificed by group monthly. During the first month, 3 groups of mice were sacrificed after 10, 20, and 30 days of treatment, respectively. A dose-related increase in liver weight was found in test animals (P less than .01). The histopathology of liver showed clear, oval cells; hypertrophied cells with foci; and neoplastic nodules that were apparent during the final 3- to 6-month period. Few oval and hypertrophied cells were found in control animals. The alpha isomer concentration of benzene hexachloride rose during the first 2 months of treatment, but declined after that time. A transient rise was noted for the beta isomer concentration of benzene hexachloride at the end of the 6-month period. The gamma isomer concentration was elevated during the initial 3 months of treatment, but declined during the subsequent 3 months. The results present definitive changes suggestive of precancerous states.

In utero and lactation exposure of mice to pan masala: effect on dams and pregnancy outcome.

Pan masala, a chewing mixture, is a popular alternate of areca nut/tobacco/betal quid in various parts of the world. In view of embryotoxic effects of areca nut and tobacco, it is hypothesized that in utero and lactational exposure to pan masala plain (PMP, containing areca nut as major ingredient), and pan masala with tobacco (PMT) can also have similar effects. To investigate this, pregnant female Swiss albino mice were treated with 3 and 6% of PMP and PMT from gestation day (GD) 0, 6, and 14 until lactation. They were weighed during pregnancy and lactation. At parturition, pups were counted, weighed, and measured. At weaning, dams were sacrificed for implantation count. Three percent and 6% PMT considerably reduced female fertility. Gestation length was lower in all the pan masala­treated mice, which was significant at 3 and 6% PMT treatment from GD 0. Pups born to pan masala­treated dams had significantly low birth weight at 3 and 6% PMT in GD 0 and GD 6, and 6% PMT in the GD 14 group. Sex ratio declined in the GD 0 pan masala­treated group. Neonatal death was observed in all the pan masala­treated groups from GD 0 and 6% of both PMP- and PMT-treated groups from GD 14 with respect to control. Weaning index was considerably altered in GD 0 and 14 pan masala­treated groups. Postimplantation loss was considerably high in all the pan masala­treated groups. The data points toward the in utero and lactational fetotoxic effects of pan masala treatment, mainly PMT.

Cytogenetic alterations in buccal mucosa cells of chewers of areca nut and tobacco.

OBJECTIVE: the rationale of the study was to evaluate the cytological alterations especially micronucleus (MN) and other nuclear anomalies in buccal mucosa cells of chewers to understand the genotoxic and clastogenic potential of chewing mixture (containing areca nut and tobacco as main ingredients). METHODS: the buccal cytome assay involves the examination of epithelial smear to determine micronucleated cell and other nuclear anomalies after the Feulgen plus light green staining. The assay was applied to exfoliated buccal mucosa cells of 262 subjects [non-chewers - 161 and chewers - 101 (includes 20 subjects with OSMF)] and 1000 cells per individual were examined microscopically. Nuclear anomalies were compared among chewers, non-chewers and OSMF subjects and correlated with consumption of quids per day and duration of chewing in years. RESULTS: MN cells were found significantly (p<0.0001) higher among chewers and OSMF subjects as compared to non-chewers. Further analysis indicated that MN was significantly higher in OSMF subjects with respect to even chewers. Nuclear buds were significantly higher (p<0.0001) in OSMF subjects as compared to chewers as well as non-chewers. Nuclear anomalies viz. binucleated, karyorrhexis and karyolysis were also considerably higher in OSMF subjects as compared to non-chewers. CONCLUSION: the MN and other nuclear anomalies reflected genetic damage and cytotoxicity, associated with tobacco and areca nut consumption. Further, these data reveal a risk for development of OSMF among chewers of mixture containing areca nut and/or tobacco, as all the OSMF subjects were chewers.

Evaluation of genotoxicity of pan masala employing chromosomal aberration and micronucleus assay in bone marrow cells of the mice.

Pan masala is commonly consumed in south-east Asian and other oriental countries as an alternate of tobacco chewing and smoking. Genotoxic potential of pan masala (pan masala plain and pan masala with tobacco known as gutkha) was evaluated employing chromosome aberration (CA) and micronucleus (MN) assay in vivo. Animals were exposed to three different doses (0.5%, 1.5% and 3%) of pan masala plain (PMP) and gutkha (PMT) through feed for a period of 6 months and micronucleus and chromosomal aberrations were studied in the bone marrow cells. Induction of mean micronuclei in polychromatic erythrocytes (MNPCE) and normochromatic erythrocyte (MNNCE) was higher in both types of pan masala treated groups with respect to control group. Both pan masala plain and gutkha treatment significantly induced the frequency of MNPCE and MNNCE in the bone marrow cells, indicating the genotoxic potential. Furthermore, slight decline in the ratio of polychromatic erythrocytes to normochromatic erythrocytes was also noticed, suggesting the cytotoxic potential even though the ratio was statistically non significant. A dose-dependent, significant increase in chromosome aberration was observed in both types of pan masala treated mice with respect to control. However, no significant difference in micronucleus and chromosomal aberration induction was noticed between two types of pan masala exposed (PMP and PMT) groups. Results suggest that both types of pan masala, i.e. plain and gutkha, have genotoxic potential.

Poisoning of workers working in small lead-based units.

BACKGROUND: No data are available with the labor departments among the workers of small-scale lead-based units with regard to lead poisoning. One hundred and ninety-five workers were investigated for lead exposure and three were found exceeding the limit of 80 mg/dL, which required a treatment for lead poisoning. AIM: To assess the exposure and health risk in workers working in small lead-based units. SETTING AND DESIGN: Random sampling is selected from the cross-sectional medical study. METHODS AND MATERIALS: Medical examination cum biochemical/hematological investigations along with blood lead estimation were carried out in these workers. STATISTICAL ANALYSIS: Epi-Info and SPSS 16.0 were used for statistical analysis. RESULTS AND CONCLUSION: Workers' blood lead levels were brought down from 114.4, 110.0 and 120.6 mg/dL with treatment of D-penicillamine to 40 mg/dL. It may be concluded that lead poisoning is a preventable public health problem that particularly affects the industrial workers in small lead-based units.

Clinical and biochemical investigations to evolve early diagnosis in workers involved in the manufacture of hexachlorocyclohexane.

365 individuals exposed to hexachlorocyclohexane (HCH) during its manufacture were studied and compared with 146 controls. These 502 cases were divided into five groups on the basis of serum total HCH levels. Exposure was mainly through skin. Results indicate that neurological symptoms reported were due to recent exposure to HCH and were related to the intensity of exposure. ECG abnormalities were seen in exposed cases. A significant positive correlation to liver related enzymes gamma GT, LAP and Alp. P was observed. beta-globulins significantly increased along with an increase in total HCH. Changes in IgM levels were similar as observed in IgA. Remarkably high concentration of HCH residues were found in the serum samples of all exposed workers. beta-HCH contributed more than 80% of the total HCH built up. Presence of circulating immune complexes was also detected in representative samples. Further, repeated biological monitoring was advised to be carried out among such workers exposed to HCH, to evaluate signs of severe health impairment in them.

Serum hexachlorocyclohexane residues in workers engaged at a HCH manufacturing plant.

The accumulation of hexachlorocyclohexane, one of the most commonly used chlorinated insecticides, was studied in a high-risk group of people who are exposed during its manufacturing process. The serum HCH concentration was estimated by quantitating its alpha, beta, gamma and delta-isomers with the help of GLC analysis. Exposed workers involved in maintenance work were found to have 3 times higher HCH residues than the controls, while in the plant operators and supervisors the levels were 5 times higher. The most severely exposed were the handlers who are in direct contact with the insecticide. The level of serum HCH residues in them was found to be about 12 times higher than those in the controls. The percentage composition of the insecticide, with respect to the different isomers, varied up to 20% for the gamma-isomer, up to 30% for alpha-isomer and had between 60%-100% of the beta-isomer in the samples. The serum levels of the insecticides were clearly related to the workers' job-related close contact, intensity, frequency, and the duration of their exposure. The total of HCH residues in the sera studied were in the range of 0.143-1.152 ppm. The importance of these findings and the relevant protective measures are discussed.