Percutaneous in vivo and in vitro absorption of lead.

Diffusion tubes were used to measure the degree of in vitro penetration of tetrabutyl lead, lead naphthanate, lead nuolate, lead acetate and lead oxide in excised guinea pig skin and human skin from autopsy. Tetrabutyl lead demonstrated the greatest penetration in skin from both guinea pig and man. Lead nuolate, lead naphthanate and lead acetate followed in descending order in the human tissue. A similar pattern occurred with guinea pig skin in most cases. There were no measurable amounts of lead oxide absorbed in either species. In vivo absorption was measured by applying 300 mg/kg tetrabutyl lead, lead nuolate, lead naphthanate or lead oxide to the shaved backs of guinea pigs for 7 d under occluded wrappings. Tetrabutyl lead was present in tissues in the highest quantities. Lead nuolate was present in greater amounts than lead naphthanate in the liver and kidneys. Lead acetate was the most poorly absorbed with the exception of lead oxide which demonstrated no absorption.     

Subacute effects of low dose lead nitrate and mercury chloride exposure on kidney of rats

Lead nitrate and mercury chloride are the most common heavy metal pollutants. In the present study, the effects of lead and mercury induced nephrotoxicity were studied in Wistar rats. Lead nitrate (LN, 45 mg/kg b.w/day) and mercury chloride (MC, 0.02 mg/kg b.w/day) and their combination were administered orally for 28 days. Four groups of rats were used in the study: control, LN, MC and LN plus MC groups. Serum biochemical parameters, lipid peroxidation, antioxidant enzymes and histopathological changes in kidney tissues were investigated in all treatment groups. LN and MC caused severe histopathological changes. It was shown that LN, MC and also co-treatment with LN and MC exposure induced significant increase in serum urea, uric acid and creatinine levels. There were also statistically significant changes in antioxidant enzyme activities (SOD, CAT, GPx and GST) and lipid peroxidation (MDA) in all groups except control group. In this study, we showed that MC caused more harmful effects than LN in rats.     

Oxidative stress and apoptotic changes in primary cultures of rat proximal tubular cells exposed to lead

Lead is a known nephrotoxic element. In this study, primary cultures of rat proximal tubular (rPT) cells were treated with different concentrations of lead acetate (0.25, 0.5 and 1 μM) to investigate its cytotoxic mechanism. A progressive loss in cell viability together with a significant increase in the number of apoptotic and necrotic cells and lactate dehydrogenase release were seen in the experiment. Simultaneously, elevation of reactive oxygen species levels and intracellular [Ca²⁺]i, depletion of mitochondrial membrane potential and intracellular glutathione were revealed during the lead exposure. In addition, apoptotic morphological changes induced by lead exposure in rPT cells were demonstrated by Hoechst 33258 staining. The apoptosis was markedly prevented by N-acetyl-l-cysteine, while the necrosis was not affected. Moreover, catalase and superoxide dismutase activities in the living cells rose significantly. In conclusion, exposure of rPT cells to low-concentration lead led to cell death, mediated by an apoptotic and a necrotic mechanism. The apoptotic death induced by oxidative stress was the chief mechanism. Meanwhile, a group of cells survived lead action, mediated by their ability to activate antioxidant defense systems.     

The effect of oral administration of Allium sativum extracts on lead nitrate induced toxicity in male mice

Lead is a common environmental occupational toxic metal, known to have indirect oxidative effects. Considering the antioxidant properties of garlic, this study was undertaken to evaluate the therapeutic efficacy of garlic extracts in terms of normalization of altered hematological, biochemical and immunological parameters, and depletion of inorganic lead burden in blood, kidney and brain tissues. Chronic lead nitrate ingestion showed a significant decline in total erythrocyte count, total leukocyte count, hemoglobin concentration, lymphocyte and monocyte content, while neutrophil content increased in lead nitrate treated group. Pb(NO₃)₂ exposure elicited a significant escalation in thiobarbituric acid reactive substances level and depletion in reduced glutathione content and antioxidant enzymes namely, superoxide dismutase and catalase in kidney and brain. Activities of aspartate transaminase, alanine transaminase, acid phosphatase and alkaline phosphatase augmented significantly in kidney and brain of lead exposed mice. Lead nitrate treatment decreased protein content while cholesterol and lead burden increased significantly. A decrease in viability of macrophage, phagocytic index, immunoglobulin level and plaque count were the salient features observed in lead exposed animals. However, oral administration of garlic extracts to Pb(NO₃)₂ treated groups attenuated the deranged parameters to some extent. This indicates that garlic can be a protective regimen for lead toxicity.     

Behavioral effects of low level neonatal lead exposure

Rats exposed to lead via maternal milk were tested at various stages of development on a number of behavioral tasks. Beginning at paturition, the dams were given either tap water, 0.02%, or 0.10% lead acetate in the drinking water. Pups from all three groups were weaned to normal chow and tap water at 21 days of age. The mean lead concentration of the dam's blood and of neonatal (20 days of age) brain and blood were all below 50 microgram/100 ml. No significant differences were found between the high lead-exposed group and controls in general as measured by wheel running over a 21 day period beginning at 30 days of age. However, there was a significant difference in wheel running behavior during the first three hr of testing. Both lead-exposed groups were found to display significantly less aggressive behavior as measured by the shock-elicited aggression test. Low level lead exposure had no discernable effect on the acquisition and subsequent reversal of a successive brightness discrimination task. Lead exposure under these conditions appears to affect some aspects of emotional behavior, while having little effect on general activity or cognitive function.     

Low-level lead exposure triggers neuronal apoptosis in the developing mouse brain

While the toxic effects of lead have been recognized for millennia, it has remained a significant public health concern due to its continued use and toxicological potential. Of particular interest is the increased susceptibility of young children to the toxic effects of lead. Although the exact mechanism(s) for lead toxicity is currently not well understood, research has established that it can be a potent NMDA antagonist. Previous research has established that exposure to NMDA antagonists during the brain growth spurt period (first 2 weeks of life in mice) can produce apoptotic neurodegeneration throughout the brain. Based on this information, the ability of lead exposure (two injections of 350 mg/kg lead 4 h apart) to produce apoptosis in the neonatal mouse brain was assessed histologically 8-24 h after treatment using activated caspase-3 immunohistochemistry, De Olmos silver technique, Nissl staining, and electron microscopy. Lead exposure produced significant neurodegeneration in the caudate/putamen, hippocampus, subiculum, and superficial and deep cortical layers of the frontal cortical regions. Further ultrastructural examination revealed cellular profiles consistent with apoptotic cell death. Statistical results showed that lead exposure significantly increased apoptotic neurodegeneration above that seen in normal controls in animals treated at postnatal day 7, but not on day 14. The results of this study may provide a basis for further elucidation of mechanisms through which the immature nervous system may be particularly susceptible to lead exposure.     

Cellular and molecular toxicology of lead. II. Effect of lead on delta-aminolevulinic acid synthetase of cultured cells.

The effect of lead nitrate on delta-aminolevulinic acid synthetase (ALA synthetase), the first and rate-limiting enzyme of the heme biosynthetic pathway, was studied by using an established rat liver cell line (RLC-GAI). Lead was shown to produce a time-dependent increase in ALA synthetase activity, with a maximum after incubation of the cells for 24 h with 10(-5) M lead nitrate. The effect of lead was not liver-specific in that increases in enzyme activity were produced in other tissue-derived cell lines. Cycloheximide but not actinomycin D, cordycepin, or hydroxyurea, at concentrations that inhibit the synthesis of protein, RNA, and DNA, prevented the lead-associated increase in ALA synthetase activity. Heme, added to the cells as hemin, also prevented the effect of lead. These results indicate that lead induced the synthesis of ALA synthetase secondary to an inhibition of the synthesis of heme.     

Modulation of ERK1/2 and p38(MAPK) by lead in the cerebellum of Brazilian catfish Rhamdia quelen

Lead (Pb2+) is a neurotoxic trace metal, widespread in aquatic environment that can change physiologic, biochemical and behavioral parameters in diverse fish species. Chemical exposure may drive modulation of mitogen-activated protein kinases (MAPKs) that are a family of highly conserved enzymes which comprise ubiquitous groups of signaling proteins playing critical regulatory roles in cell physiology. Extracellular signal-regulated kinases (ERK1/2) and p38(MAPK) control complex programs such as gene expression, embryogenesis, cell differentiation, cell proliferation, cell death and synaptic plasticity. Little information is available about MAPKs in aquatic organisms and their modulation by trace metals. The aim of this work was to determine the modulation of ERK1/2 and p38(MAPK) phosphorylation by Pb2+ in vivo and in vitro, in cerebellar slices of the catfish, Rhamdia quelen. In the in vitro model, slices were incubated for 3 h with lead acetate (1-10 microM). In the in vivo studies, the animals were exposed for 2 days to lead acetate (1 mg L(-1)). ERK1/2 and p38(MAPK) (total and phosphorylated forms) were immunodetected in cerebellar slices by Western blotting. Pb2+ added in vitro at 5 and 10 microM increased significantly the phosphorylation of both MAPKs. The in vivo exposed animals also showed a significant increase of ERK1/2 and p38(MAPK) phosphorylation without changes in the total content of the enzymes. In conclusion, the present work indicates that it is possible to evaluate the ERK1/2 and p38(MAPK) activation in the central nervous system (CNS) of a freshwater fish largely distributed in South America. Moreover, Pb2+, an important environmental pollutant may activate in vitro and in vivo ERK1/2 and p38(MAPK) enzymes. These findings are important considering the functional and ecologic implications associated to Pb2+ exposure of a freshwater fish species, such as R. quelen, and the roles of ERK1/2 and p38(MAPK) in the control of brain development, neuroplasticity and cell death.     

Beneficial effects of garlic on learning and memory deficits and brain tissue damages induced by lead exposure during juvenile rat growth is comparable to the effect of ascorbic acid

Objective: The neuroprotective effects of both garlic and ascorbic acid (AA) have been documented. In this study the effects of garlic and ascorbic acid on memory deficits and brain tissue oxidative damages induced by lead exposure was investigated. Methods: The juvenile rats were divided and treated: (1) Control, (2) Lead (lead acetate in drinking water, 8 weeks), (3) Lead – Ascorbic Acid (Lead-AA), (4) Lead – Garlic (100?mg/kg, daily, gavage) (Lead-Gar). Results: In Morris water maze (MWM), the escape latency and traveled path in the Lead group were significantly higher while, the time spent in the target quadrant (Q1) was lower than Control. Both Lead-Gar and Lead-AA groups spent more times in Q1than to lead group. There were no significant differences in swimming speed between the groups. In passive avoidance (PA) test, the time latency for entering the dark compartment by Lead group was lower than Control. Treatment of the animals by AA and garlic significantly increased the time latency. In Lead group, the total thiol concentration in brain tissues was significantly lower while, MDA was higher than Control. Treatment by both garlic and AA increased total thiol concentrations and decreased MDA. Both garlic and AA decreased the lead content of brain tissues. Conclusion: It is suggested that treatment with garlic attenuates the learning and memory impairments due to lead exposure during juvenile rat growth which is comparable to AA. The possible mechanism may be due to its protective effects against brain tissues oxidative damage as well the lowering effects of brain lead content.     

Effect of lead toxicity on coenzyme Q levels in rat tissues

Lead is a persistent and common environmental contaminant, which chiefly plays a significant role in modern industry. Coenzyme Q acts as electron and proton carrier in mitochondria and functions as an antioxidant in its reduced form (ubiquinol). To investigate the hazardous effects of lead on the coenzyme Q level, rats were injected i.p. with lead acetate (5 mg/kg b.wt. daily for 6 weeks). Our results showed that the levels of both oxidized (ubiquinone) and reduced (ubiquinol) forms of coenzyme Q₉ and Q₁₀ in serum, brain, liver and kidney of lead-treated rats are quite different depending on the organ tissue type.     

Ebselen and diphenyl diselenide do not change the inhibitory effect of lead acetate on delta-aminolevulinate dehidratase

It is known that lead is toxic to several species of animals, and growing data support the participation of oxidative in lead toxicity. Selenium compounds, like diphenyl diselenide and Ebselen have a thiol-peroxidase like and other antioxidant properties. In this work, we determine whether these non-thiol-containing compounds with antioxidant properties could reverse the toxicity produced by Pb(2+). Lead acetate injection followed by injection with Ebselen or diphenyl diselenide did not change the levels of non-protein thiol groups (NPSH), whereas simultaneous treatment with lead plus Ebselen reduced NPSH levels in liver. Lead and Ebselen caused a marked reduction in TBARS level in kidney, whereas lead or selenium compounds did not change TBARS levels in brain or liver. Lead acetate inhibited, δ-aminolevulinate dehydratase (ALA-D) activity in blood, liver, kidney and brain. Selenium compounds did not change enzyme activity nor the inhibitory effect of lead acetate in kidney and liver. Ebselen reversed brain ALA-D inhibition caused by Pb(2+). Reactivation index for ALA-D by DTT was higher in lead-treated groups than control groups in all tissues. Lead acetate or selenium compounds did not demonstrate alteration on [(3)H]-glutamate uptake by synaptosomes, whereas lead acetate plus Ebselen showed an increase on [(3)H]-glutamate uptake. The results of the present study indicate that ALA-D inhibition antecedes the overproduction of reactive oxygen species, which is becoming well documented in the literature.     

Biochemical adaptation of wild and cultivated soybean against toxicity of lead salts

Agricultural production is becoming increasingly dependent on the environmental factors that alter soil properties, plant productivity, and product quality. Environment pollution caused by heavy metals because of human activities are among the most dangerous pollutants on the biosphere. Here, we have studied the biochemical adaptation of wild and cultivated soybeans to the simulated effects of lead nitrate and lead acetate. Lead in the form of acetate had a relevant toxic effect, as evidenced by a significant increase in the concentration of malonic dialdehyde in the treated samples relative to control samples. Catalase and peroxidase, possibly performing a signaling function, are involved in the adaptation to the toxicity of Pb salts. The studied Pb salts showed a predominant stimulating effect on the specific activity of acid phosphatases in cultivated soybean, while the ribonuclease activity changed in both Glycine species. Moreover, in wild soybean, it was mostly suppressive, except for the first day. We found that the electrophoretic spectra of acid phosphatases of soybean seedlings was highly stabile, while that of ribonucleases varied depending on the salt. On the seventh day of exposure, lead nitrate caused a decrease in the specific activity of the studied hydrolases of seedlings of cultivated and wild soybeans. A change in the number or electrophoretic mobility of multiple forms of enzymes during treatment with Pb salts was revealed, which indicates the adaptation of the plants at the molecular genetic level. These results imply that the observed enzymes can be used as sensitive indicators for predicting the effects of heavy metals on soybean.     

Light and electron microscopic study of the toxic effect of prolonged lead exposure on the seminiferous tubules of albino rats and the possible protective effect of ascorbic acid

Background

Lead has been long recognized as an occupational toxicant. Now, lead is considered to be one of the major environmental pollutants which have serious potential threat to human health. Reports of declining male fertility have renewed interest in the role of environmental and occupational exposures in the etiology of human infertility.

Objectives

The aim of the present work was to investigate the toxic effect of prolonged exposure to lead on albino rat’s seminiferous tubules and the possible protective effect of ascorbic acid (vitamin C) on lead toxicity.

Methods

The study was carried out on 40 adult male albino rats divided into three groups: a control group which comprised three subgroups; negative control which received no treatment and two positive controls receiving an oral daily dose of distilled water or ascorbic acid, respectively. Lead acetate - intoxicated group received lead acetate orally in a dose of 25 mg/kg b.w dissolved in distilled water for 3 months. While the protected group received ascorbic acid orally in a dose of 100 mg/kg b.w daily for 3 months, simultaneously with lead acetate in a similar dose to that of the intoxicated group. By the end of the experimental period, blood samples were collected for estimation of blood lead level. Fresh specimens were taken from the testis and processed for light and electron microscopic examination.

Results

Biochemical analysis demonstrated significant increase in the blood lead level in lead intoxicated group compared to the control group, while ascorbic acid - protected group revealed significant decrease in the blood lead level. Light microscopic examination of lead treated group revealed loss of normal archicture of the testicular tissue in the form of thin walled seminiferous tubules with wide lumen and vacuolations in the spermatogenic epithelium mostly separating primary spermatocytes from spermatogonia and surrounding nuclei of Sertoli cells. Apoptotic bodies were found among the basal part of the spermatogenic epithelium. Ultrastructural examination of the same group revealed degenerating cells with cytoplasmic vacuolations, apoptotic cells with heterochromatic nuclei and dense cytoplasm, irregularities in the basal lamina with increased collagen deposition and shrunken myoid cells. Late differentiating spermatids showed deformed head with widening of the subacrosomal space and redundant acrosome, in addition to the abnormalities in the tail of developing sperms. On the other hand, ascorbic acid - protected group showed amelioration of most of the degenerative changes depicted in the lead intoxicated group.

Conclusion

Lead had an injurious effect on the testis of the exposed animals. Simultaneous administration of ascorbic acid was efficient in preventing most of such toxic effect and decreasing the blood lead level. Therefore, ascorbic acid can be recommended for protection against lead intoxication.     

Simultaneous effect of lead and cadmium on granulosa cells: a cellular model for ovarian toxicity

Lead (Pb) and cadmium (Cd) are known reproductive toxicants, which accumulate in granulosa cells of the ovary. Female Charles foster rats were treated with sodium acetate (control), lead acetate and cadmium acetate either alone or in combination at a dose 0.05 mg/kg body weight intra-peritoneally for 15 days daily. Animals were killed at proestrous stage and granulosa cells were isolated from the ovaries. Binding of (125)I-luteinizing hormone ((125)I-LH), (125)I-follicle stimulating hormone ((125)I-FSH) and 17beta-hydroxysteroid dehydrogenase activity were measured. As these receptors are localized on the surface of the cell membrane, we also estimated the membrane parameters of these cells. Our results demonstrated that both lead and cadmium caused a significant reduction in gonadotropin binding, which altered steroidogenic enzyme activity of granulosa cells. These changes exhibited a positive correlation with membrane changes of the granulosa cells.     

Lead in blood and tissues of mice after administration of low lead doses

Lead levels in whole blood could be determined reliably up to a lower limit of 2 g/100 ml blood, using a modified micromethod of the graphite tube furnace technique. Lead contents of various tissues were also determined by using the automated graphite tube furnace after wet ashing of the organs with nitric acid in autoclaves.Animal experiments with mice showed no measurable increase in blood lead level after a single, 10- or 30-days oral administration of lead in doses of 10–1000 g lead acetate/kg body weight/day. However, these doses led to a rise in tissue lead content. There was a clear dependence of tissue lead content on type of organ examined, lead dose and duration of lead exposure.According to our experiments, the threshold dose which leads to a long-term increase in tissue lead content is assumed to be about 100 g lead acetate/kg body weight/day, orally administered.We are thankful to Prof. Dr. H. Rüssel, Hannover and Dr. M. Fleischer, Saarbrücken for helping in comparison studies.We would also like to thank Mr. H. Dick and Miss. Ch. Hecker for the technical and laboratory assistance.     

Acute lead poisoning of the pigeon induced by single,intraperitoneal administration of lead acetate

A single dose of lead acetate (either 30 mg/kg or 150 mg/kg) was intraperitoneally (i.p.) adminstered to adult feral pigeons, Columba livia var and the effects of calcium disodium ethylenediamine tetraacetate (CaNa₂EDTA), (0, 150, 300, 600 mg/kg), administered i.p. twice a week in the ensuing period were observed. Lead acetate caused dose related mortality and decreases in weight, hematocrit and -aminolevulinic acid dehydratase activity (ALA-D). Acute toxicity of lead acetate in the pigeon, when given intraperitoneally, appeared approximately equivalent to that in the rat and mouse in terms of LD₅₀. Blood lead (blood Pb) levels observed during the lethal stage were five to ten times less than those reported for chronic oral lead poisoning in the pigeon. Biological implications of elevated levels of blood Pb observed in the feral pigeon in the urban Tokyo area are discussed. CaNa₂EDTA induced dose related recovery in ALA-D in 30 mg/kg group, and reduction of blood Pb levels in the group dosed with 150 mg/kg of lead acetate.     

Combined lead acetate and disulfiram treatment-induced alterations of glial fibrillary acidic protein (GFA) immunoreactive astrocytes in brain smears

Dithiocarbamates are known to form lipid-soluble complexes with lead and greatly increase brain lead levels. The present study was undertaken to investigate whether lead acetate, when administered together with disulfiram (Antabuse, metabolite of dithiocarbamate) during development, would induce morphological changes in brain astrocytes. Female Sprague-Dawley rats were given 0.25% lead acetate in the drinking water from day one of pregnancy and this treatment was continued after birth until the litters were 4 weeks old. In addition, some dams received disulfiram in a dose of 0.1 mmol/kg p.o. twice weekly and after parturition this dose was given s.c. directly to the offspring twice a week. Lead acetate and disulfiram treatments were discontinued at weaning and animals sacrificed 3 weeks later. Samples of parietal cortex, hippocampal formation and cerebellar cortex were dissected out and smeared onto glass-slides and astrocytes were visualized in toto using immunohistochemistry with antibodies against glial fibrillary acid protein (GFA), enabling morphometric analysis with a computerized image analyser. Animals treated with lead acetate showed a minor increase in the size of the GFA-immunoreactive astrocytes in parietal cortex smears, while animals treated with disulfiram showed no difference in size or form compared to controls. However, in combined lead acetate and disulfiram-treated animals a profound increase in astrocyte size and an increase in the number of processes of the individual GFA-immunoreactive astrocytes could be demonstrated in parietal cortex. No significant changes were noted in GFA-immunoreactive astrocytes of hippocampal smears following the different treatments, while GFA-immunoreactive astrocytes in cerebellar cortex smears were significantly smaller and had reduced number or processes following the combined lead acetate and disulfiram treatment compared to lead acetate treatment or controls. It is concluded that combined exposure to lead acetate and disulfiram during development induces regionally specific changes in GFA-immunoreactive astrocyte morphology. Furthermore, the present study demonstrates the usefulness of smear preparations combined with computerized image analysis to study the morphology of GFA-immunoreactive astrocytes as an index of toxic effects in CNS.     

母体铅暴露对仔鼠海马中一氧化氮/环鸟苷酸的影响

目的：通过研究母体铅暴露对发育期仔鼠海马组织NO／cGMP水平变化，分析母体铅暴露对子鼠海马NO／CGMP途径的影响。方法：将母鼠从孕期第1天至仔鼠出生第20天分别饮用双蒸水、200、100、50mg/L醋酸铅溶液，检测20日龄和60日龄仔鼠物血铅、脑铅水平及海马组织nNOS神经原细胞、NO、NOS、cGMP浓度。结果：20日龄染毒组仔鼠的血铅、脑铅浓度与对照组相比有显著性增高，60日龄仔鼠的血铅与对照组相比，差异无显著性，但脑铅仍比对照组高；20日龄和60日龄染毒组仔鼠海马组织nNOS阳性神经元强度值、吸光度（A）、一氧化氮、NOS、cGMP浓度与对照组相比有显著降低，一氧化氮、NOS、cGMP三者相关性较好。结论：母体铅暴露可损伤仔鼠海马组织NO／cGMP途径，这种损伤在停止铅接触一段时间后仍持续存在。     

Effects of lead(II) nitrate and a dithiocarbamate fungicide on the rat lung.

The pulmonary toxicity of two potential environmental pollutants was studied in rats 1, 7 and 30 days after a single intratracheal instillation of lead nitrate and Dithane M-45 (mancoceb), either individually or in various combinations. The cell count, protein, phospholipids and lactate dehydrogenase level were determined in the bronchoalveolar lavage fluid, as were the protein, phospholipids and acid phosphatase contents in the lung tissue. Lead nitrate and Dithane M-45 induced acute inflammation reactions with different features. The effects of mixtures of lead nitrate and Dithane M-45 were found to be different from those of the individual components.