Arnulfo Albores

Chronic Arsenic Poisoning in the North of Mexico

1 We compared the prevalence of signs and symptoms of chronic arsenic poisoning in two rural populations. 2 The arsenic concentration in the drinking water of the exposed population was 0.41 mg/l, and 0.007 mg/l in the control population. 3 The arsenic was present mainly (70%) in its pentavalent form. 4 The objective was to quantitate health effects and risks derived from chronic ingestion of arsenic in contaminated water. 5 In the exposed population, 21.6% of the sample, showed at least one of the cutaneous signs of chronic arsenic poisoning against 2.2% in the control town. 6 Non-specific symptoms were more prevalent in the exposed population and they occurred more frequently in those individuals with skin signs. 7 The relative risk of suffering a particular manifestation of poisoning, ranged from 1.9 to 36 times higher in the exposed population. 8 We estimated the risks above mentioned, which were derived from exposure to minute quantities of arsenic in a known proportion of its oxidation states during a life time period.

Cytogenetic effects in human exposure to arsenic

The cytogenetic effects of arsenic exposure were studied among rural populations that live in the same geographical area and have similar socioeconomic status, but different degree of exposure to inorganic arsenic (As) via drinking water. A group of inhabitants of Santa Ana (408.17 micrograms/l of As in drinking water) were considered the exposed individuals and a group of inhabitants of Nazareno (29.88 micrograms/l) were considered as controls. Blood and urine samples were obtained from volunteers. Past and current exposure, health, and nutritional status as well as the presence of arsenic skin lesions were ascertained in study participants through questionnaires and physical examination. The frequencies and types of chromosomal aberrations in first-division metaphases were studied in whole blood lymphocyte cultures while the presence of micronuclei (MN) was studied in exfoliated epithelial cells obtained from the oral mucosa and from urine samples. Total arsenic (TAs) content, and the relative proportions of inorganic arsenic (IAs), and the metabolites monomethylarsonic (MMA) and dimethylarsinic (DMA) acid were determined in urine samples. Exposed individuals showed a significant increase in the frequency of chromatid and isochromatid deletions in lymphocytes and of MN in oral and urinary epithelial cells. Males were more affected than females, and a higher number of micronucleated oral cells were found among those individuals with skin lesions. The type of cytogenetic damage observed gives evidence of arsenic as a clastogenic/aneugenic carcinogen.

Chlorzoxazone, a selective probe for phenotyping CYP2E1 in humans.

The ability of human cytochromes P450 other than CYP2E1 to catalyse the 6-hydroxylation of chlorzoxazone (6-OH-CHZ) was examined in vitro using human liver microsomal preparations and in vivo using chlorzoxazone as a metabolic probe. Chlorzoxazone 6-hydroxylation activity was significantly correlated with 4-nitrophenol 2-hydroxylase activity and immunodetected CYP2E1 in 14 human liver samples (r = 0.92 and 0.81, P < 0.001, respectively). Conversely, this catalytic activity was not correlated with CYP 3A or CYP1A activities. Diethyldithiocarbamate (DEDTC), a specific CYP2E1 inhibitor, reduced chlorzoxazone 6-hydroxylase activity by 92.3 +/- 7.6% (n = 14 samples) while ketoconazole, a specific CYP3A inhibitor, reduced this activity by 8.6 +/- 6.3% (n = 14). The residual activity following preincubation with DEDTC was significantly correlated with nifedipine oxidation and tamoxifen N-demethylations, both specific to CYP3A (r = 0.76 and 0.68, respectively). Genetically produced pure human CYP2E1 and 3A4 hydroxylated chlorzoxazone with turnover numbers of 19.7 and 0.14 min(-1), respectively. Furthermore, cytochrome b5 stimulated chlorzoxazone 6-hydroxylation. From examination of the relative liver content of CYP2E1 and 3A, it can be asserted that CYP2E1 is the major enzyme involved in chlorzoxazone 6-hydroxylation and that the contribution of CYP3A is very minor. CYP2E1 activity was evaluated by the plasmatic metabolic ratio 6-OH-CHZ/CHZ (CHZ-MR) measured 2 h after ingestion of 500 mg CHZ. Smoker status did not influence the rate of CHZ hydroxylation. The CHZ-MR was 0.30 +/- 0.13 (mean +/- SD) n = 39 non-smokers versus 0.32 +/- 0.15, n = 75 smokers. This result suggests that CYP1A, inducible by cigarette smoking, is not significantly involved in chlorzoxazone hydroxylation. Women exhibited a slightly lower CHZ-MR than men (0.29 +/- 0.15, n = 44 versus 0.34 +/- 0.15 n = 49, respectively). Obesity increased CHZ-MR, especially in non-insulin-dependent diabetic individuals (0.45 +/- 0.21, n = 13 versus 0.30 +/- 0.15, n = 42 control individuals, P = 0.007). Furthermore, exposure of workers to volatile organics in a shoe factory decreased CHZ-MR (0.19 +/- 0.09, n = 10 Mexican workers versus 0.34 +/- 0.12, n = 16 Mexican control individuals, P = 0.001). Concomitant administration of grapefruit juice (known to be an inhibitor of CYP3A4) with chlorzoxazone did not significantly modify the CHZ metabolic ratio: 0.29 +/- 0.1 versus 0.31 +/- 0.1, for nine control individuals without and with grapefruit juice, respectively. In conclusion, all these results demonstrate that chlorzoxazone is a very selective probe for phenotyping CYP2E1 in humans.

Lymphocyte replicating ability in individuals exposed to arsenic via drinking water.

A human monitoring study was carried out to explore the effect on lymphocyte proliferation of chronic exposure to arsenic (As) via drinking water. Blood and urine samples were taken from volunteers from a town where levels of As in the drinking water averaged 412 micrograms/l, and from a matched group of individuals, with similar socioeconomic status, that drank water with As average levels of 37.2 micrograms/l. Exposure was assessed by questionnaires and by determining the levels of As in urine and water samples. The evaluation of the peripheral blood lymphocyte proliferation was done at different culture times using labelling (LI), mitotic (MI) and replication indexes (RI) as endpoints. No significant differences were seen for either LI or MI, except for MI in 72 h cultures and in LI in males and females with skin lesions vs. those without lesions. Significant differences in RI were seen for exposed females but not for males. Correlations between LI and MI showed that progression from the initial S-to M-phase is altered in exposed individuals. Arsenic exposure as well as lead and mercury affect cellular immune response, making the endpoints of cell proliferation variables of interest in population monitoring study design, since they might provide information in health impairment due to exposure, which is important in risk assessment.

Microsomal transformation of organophosphorus pesticides by white rot fungi

The enzymatic mechanism for the transformationof organophosphorus pesticides (OPPs) by differentwhite-rot fungi strains was studied. With theexception of Ganoderma applanatum 8168,all strains from a collection of 17 different fungicultures were able to deplete parathion. Threestrains showing the highest activities were selectedfor further studies: Bjerkandera adusta 8258,Pleurotus ostreatus 7989 and Phanerochaetechrysosporium 3641. These strains depleted 50 to96% of terbufos, azinphos-methyl, phosmet andtribufos after four-days exposure to the pesticides.In order to identify the cellular localization of thetransformation activity, the extracellular andmicrosomal fractions of Pleurotus ostreatus7989 were evaluated in vitro. While the activitiesof ligninolytic enzymes (lignin peroxidase,manganese peroxidase and laccase) weredetected in the extracellular fraction, noenzymatic modification of any of the fivepesticides tested could be found, suggestingthe intracellular origin of the transformationactivity. In accordance with this observation themicrosomal fraction was found able to transformthree OPPs with the following rates:10 μmol mg prot-1 h-1 forphosmet, 5.7 μmol mg prot-1 h-1 forterbufos, and 2.2 μmol mg prot-1 h-1 forazinphos-methyl. The products from these reactions andfrom the transformation of trichlorfon and malathion,were identified by mass-spectrometry. These results,supported by specific inhibition experiments and thestringent requirement for NADPH during the in vitroassays suggest the involvement of a cytochrome P450.

Altered Urinary Porphyrin Excretion in a Human Po p ulation Chronically Exposed to Arsenic in Mexico

1 A detailed study of the urinary excretion pattern of porphyrins in humans chronically exposed to As via drinking water was performed using high performance liquid chromatography (HPLC) 2 Thirty-six individuals (15 men and 21 women) were selected from a town which had 0.400 mg L -1 of As in drinking water. The control group consisted of thirty-one individuals (13 men and 18 women) whose As concentration in drinking water was 0.020 mg L-1. 3 The major abnormalities in the urinary porphyrin excretion pattern observed in arsenic-exposed individuals were: (a) significant reductions in coproporphyrin III excretion resulting in decreases in the COPRO III/COPRO I ratio, and (b) significant increases in uroporphyrin excretion. Both alterations were responsible for the decrease in the COPRO/URO ratio. 4 No porphyrinogenic response was found in individuals with urinary As concentrations below 1,000 μg of As g-1 of creatinine, However, as arsenic concentrations exceeded this value, the excretion of porphyrins (except coproporphyrin III) increased proportionally. 5 The prevalence of clinical signs of arsenicism showed a direct relationship to both As concentration in urine and time-weighted exposure to As. A direct relationship between time-weighted exposure and alterations in urinary porphyrin excretion ratios was also observed. 6 The alterations found are compatible with a lower uroporphyrinogen decarboxylase activity in arsenic-exposed individuals. However, the similarities in the urinary porphyrin excretion pattern between As-exposed individuals and Dubin-Johnson syndrome patients suggest that impairments in the excretion of coproporphyrin isomers may also contribute to the pattern observed.

Fluoride levels in well-water from a chronic arsenicism area of Northern Mexico.

This paper reports on the concentrations and geographical relationships between fluoride and total arsenic in 129 water wells of the Región Lagunera, Mexico, where arsenic has caused severe health effects. Fluoride concentrations ranged from less than 0.5 to 3.7 mg liter(-1); 25 samples (19.4%) had levels above 1.5mg liter(-1), the current WHO and Mexican drinking water standard, whereas 45 (34.9%) had levels below 0.5 mg liter(-1). The range of total arsenic concentrations was 0.008-0.624 mg liter(-1) and 64 (50%) had levels above 0.050 mg liter(-1), the current WHO standard. A linear regression analysis of arsenic and fluoride concentrations showed a highly positive correlation (r = 0.774), consistent with their geographical distribution. The highest concentrations of both elements were found in the northeastern part of the Región, mostly corresponding to rural areas, whereas the lowest concentrations were found in the southwestern part of the Región, as well as in the cities of Torreón in the state of Coahuila, and Gómez Palacio and Lerdo in the state of Durango. In consequence, people exposed to high arsenic concentrations are also exposed to fluoride at levels above the drinking water standard. The possibility of interactions between both elements is also discussed.

Cadmium and mercury toxicity in a human fetal hepatic cell line (WRL-68 cells)

The toxic effects of cadmium (Cd) and mercury (Hg), as chloride salts, were studied using an hepatic human fetal cell line (WRL-68 cells). From viability curves and the proliferative capacity of the cell in the presence of the metal, three different cell treatments were chosen, (1) 0.5 microM of the metal chloride for 24 h (acute low dose treatment), (2) 0.5 microM of the metal chloride for 7 days (chronic treatment), and (3) 5 microM of the metal chloride for 24 h (acute high dose treatment). WRL-68 cells grown in the presence of Cd exhibited the same proliferative curve as control cells, whereas in the case of Hg, the cells increased their proliferative capacity. Both metals produced ultrastructural alterations in different degrees, mainly observed as mitochondrial and RER structural changes, depending of the treatment and concentration of the metal used. Cytotoxicity was assessed by measuring the release of lactate dehydrogenase from the cells. Acutely high dose-treated cells showed the highest value for this parameter, and Cd-treated cells presented higher lactate dehydrogenase release than the Hg-treated ones. Cell damage was also measured by alanine aminotransferase (ALAT) and aspartate aminotransferase (ASAT) activities. Acute high dose Cd treatment caused the highest value of enzymatic release. Lipid peroxidation was significantly different with respect to control cells in chronic and acute high dose treatments with both metals. Metallothionein (MT) induction in response to Hg treatment was not detected. However, a dramatic induction of this protein occurred in Cd-treated cells. WRL-68 cells differentially respond to Cd and Hg making this hepatic fetal human cell line a useful tool in investigating the mechanism of toxicity of these heavy metals.

Increase of striatal dopamine release by cadmium in nursing rats and its prevention by dexamethasone-induced metallothionein

Repeated daily intraperitoneal (i.p.) administrations of cadmium (CdCl2, 1 mg/kg per day for 5 days) increased striatal dopamine (DA) release (180% of controls) and turnover (150% of controls) in 13-day-old rats. Cd treatment also increased striatal metallothionein (MT) content (161%), Cd (127%) and lipid peroxidation (LPO, 190%). In addition, Cd treatment decreased striatal tyrosine hydroxylase (TH) activity (-28%), and such an effect may result from D-2 receptor blockade as a consequence of excessive dopamine release, since sulpiride (a specific D-2 receptor antagonist) administration to Cd-treated rats abolished the effect of Cd on TH. No effect was observed on striatal monoamine oxidase (MAO) activity. Dexamethasone (Dx) treatment increased striatal MT content and caused no effect on either DA release or turnover. However, Dx administration prevented the effects caused by Cd, including the increased DA release and enhanced striatal lipid peroxidation. These results indicate that toxic effects on the brain are to be expected as a result of Cd exposure and that Dx administration can attenuate them.

Induction of metallothionein synthesis by zinc in cadmium pretreated rats

The ability of zinc (Zn) salts to induce the synthesis of metallothionein (MT) in liver, kidney and pancreas of rats pretreated with cadmium (Cd) salts was investigated. Twenty-four hours after either CdCl2 (2.0 mg Cd/kg, s.c.) or saline pretreatment, rats were injected with saline, CdCl2 (2.0 mg Cd/kg, s.c.) or ZnSO4 (20 mg Zn/kg, s.c.) and the concentrations of MT and MT-1 mRNA in tissues subsequently measured. After a single injection of Cd salts, concentrations of MT and MT-1 mRNA were significantly increased in liver as compared to control. With two injections of Cd, the accumulation of MT in liver was approximately twice the levels of MT following a single injection of Cd. In kidney, MT and MT-1 mRNA expression were significantly increased only after two injections of Cd and in the pancreas, Cd injections did not alter either MT content or MT-1 mRNA expression. Treatment with Zn salts increased MT concentrations in both liver and pancreas. However, the pancreas was the most responsive to injections of Zn salts as compared to the liver in terms of increases in both protein concentration and MT-1 mRNA expression. When Zn injection was preceded by a Cd injection, induction as measured by MT-1 mRNA and MT concentrations were approximately additive in liver. In kidney, although Cd or Zn treatment separately had no effect on MT or MT-1 mRNA content, injection of Cd followed by Zn resulted in significantly increased levels of renal MT and MT-1 mRNA. Fractionation of liver cytosols on a Sephadex G-75 column revealed that in animals receiving two injections of Cd, virtually all the Cd was associated with MT whereas Zn was distributed between both high molecular weight (HMW) proteins and MT. In animals receiving both Cd and Zn injections, cytosolic Cd was still bound predominantly to the MT fraction, while the proportion of cytosolic Zn associated with MT increased. The results of this study suggest that, treatment with Cd salts followed by Zn salt injection can induce further synthesis of MT in liver, kidney and pancreas with subsequent binding of both Zn and Cd to the intracellular MT.