Evaluation of vascular and kidney injury biomarkers in Mexican children exposed to inorganic fluoride.

Exposure to inorganic fluoride (F) has been implicated in cardiovascular and kidney dysfunction mainly in adult populations. However, limited epidemiological information from susceptible populations, such as children, is available. In this study we evaluated the relationship of F exposure with some vascular and kidney injury biomarkers in children. A cross-sectional study was conducted in 374 Mexican schoolchildren. Dental fluorosis and F concentrations in the water and urine were evaluated. The glomerular filtration rate (eGFR) and the urinary concentrations of kidney injury molecule 1 (KIM-1) and cystatin-C (uCys-C) were examined to assess kidney injury. The carotid intima media thickness (cIMT) and serum concentrations of vascular adhesion molecule 1 (VCAM-1), intracellular adhesion molecule 1 (ICAM-1), endothelin 1(ET-1) and cystatin-C (sCys-C) were measured to assess vascular alterations. High proportions of children exposed to F were observed (79.7% above 1.2 ppm F in urine) even in the low water F exposure regions, which suggested additional sources of F exposure. In robust multiple linear regression models, urinary F was positively associated with eGFR (ß = 1.3, p = 0.015), uCys-C (ß = -8.5, p = 0.043), VCAM-1 (ß = 111.1, p = 0.019), ICAM-1 (ß = 57, p = 0.032) and cIMT (ß = 0.01, p = 0.032). An inverse association was observed with uCys-C (ß = -8.5, p = 0.043) and sCys-C (ß = -9.6, p = 0.021), and no significant associations with ET-1 (ß = 0.069, p = 0.074) and KIM-1 (ß = 29.1, p = 0.212) were found. Our findings revealed inconclusive results regarding F exposure and kidney injury. However, these results suggest that F exposure is related to early vascular alterations, which may increase the susceptibility of cardiovascular diseases in adult life.

Cadmium Complexed with ß2-Microglubulin, Albumin and Lipocalin-2 rather than Metallothionein Cause Megalin:Cubilin Dependent Toxicity of the Renal Proximal Tubule.

Cadmium (Cd2+) in the environment is a significant health hazard. Chronic low Cd2+ exposure mainly results from food and tobacco smoking and causes kidney damage, predominantly in the proximal tubule. Blood Cd2+ binds to thiol-containing high (e.g., albumin, transferrin) and low molecular weight proteins (e.g., the high-affinity metal-binding protein metallothionein, ß2-microglobulin, α1-microglobulin and lipocalin-2). These plasma proteins reach the glomerular filtrate and are endocytosed at the proximal tubule via the multiligand receptor complex megalin:cubilin. The current dogma of chronic Cd2+ nephrotoxicity claims that Cd2+-metallothionein endocytosed via megalin:cubilin causes renal damage. However, a thorough study of the literature strongly argues for revision of this model for various reasons, mainly: (i) It relied on studies with unusually high Cd2+-metallothionein concentrations; (ii) the KD of megalin for metallothionein is ~105-times higher than (Cd2+)-metallothionein plasma concentrations. Here we investigated the uptake and toxicity of ultrafiltrated Cd2+-binding protein ligands that are endocytosed via megalin:cubilin in the proximal tubule. Metallothionein, ß2-microglobulin, α1-microglobulin, lipocalin-2, albumin and transferrin were investigated, both as apo- and Cd2+-protein complexes, in a rat proximal tubule cell line (WKPT-0293 Cl.2) expressing megalin:cubilin at low passage, but is lost at high passage. Uptake was determined by fluorescence microscopy and toxicity by MTT cell viability assay. Apo-proteins in low and high passage cells as well as Cd2+-protein complexes in megalin:cubilin deficient high passage cells did not affect cell viability. The data prove Cd2+-metallothionein is not toxic, even at >100-fold physiological metallothionein concentrations in the primary filtrate. Rather, Cd2+-ß2-microglobulin, Cd2+-albumin and Cd2+-lipocalin-2 at concentrations present in the primary filtrate are taken up by low passage proximal tubule cells and cause toxicity. They are therefore likely candidates of Cd2+-protein complexes damaging the proximal tubule via megalin:cubilin at concentrations found in the ultrafiltrate.

Evaluation of kidney injury biomarkers in an adult Mexican population environmentally exposed to fluoride and low arsenic levels.

Fluoride (F) is a toxicant widely distributed in the environment. Experimental studies have shown kidney toxicity from F exposure. However, co-exposure to arsenic (As) has not been considered, and epidemiological information remains limited. We evaluated the association between F exposure and urinary kidney injury biomarkers and assessed As co-exposure interactions. A cross-sectional study was conducted in 239 adults (18-77 years old) from three communities in Chihuahua, Mexico. Exposure to F was assessed in urine and drinking water, and As in urine samples. We evaluated the urinary concentrations of albumin (ALB), cystatin-C (Cys-C), kidney injury molecule 1 (KIM-1), clusterin (CLU), osteopontin (OPN), and trefoil factor 3 (TFF-3). The estimated glomerular filtration rate (eGFR) was calculated using serum creatinine (Creat) levels. We observed a positive correlation between water and urine F concentrations (ρ = 0.7419, p < 0.0001), with median values of 1.5 mg/L and 2 µg/mL, respectively, suggesting that drinking water was the main source of F exposure. The geometric mean of urinary As was 18.55 ng/mL, approximately 39% of the urine samples had As concentrations above the human biomonitoring value (15 ng/mL). Multiple linear regression models demonstrated a positive association between urinary F and ALB (ß = 0.56, p < 0.001), Cys-C (ß = 0.022, p = 0.001), KIM-1 (ß = 0.048, p = 0.008), OPN (ß = 0.38, p = 0.041), and eGFR (ß = 0.49, p = 0.03); however, CLU (ß = 0.07, p = 0.100) and TFF-3 (ß = 1.14, p = 0.115) did not show significant associations. No interaction with As exposure was observed. In conclusion, F exposure was related to the urinary excretion of early kidney injury biomarkers, supporting the hypothesis of the nephrotoxic role of F exposure.

Sub-chronic exposure to fluoride impacts the response to a subsequent nephrotoxic treatment with gentamicin.

Fluoride is an important groundwater contaminant, and more than 200 million people are exposed to high fluoride levels in drinking water, the major source of fluoride exposure. Exposure above 2 ppm of fluoride is associated with renal impairment in humans. In rats, moderate levels of fluoride induce kidney injury at early stages in which the glomerular filtration rate (GFR) is not altered. In the present study, we investigated if sub-nephrotoxic stimulus induced by fluoride might impact the response to a subsequent nephrotoxic treatment with gentamicin. Male Wistar rats (~21 days) were exposed to 0, 15 or 50 ppm of fluoride through drinking water during 40 days. Afer that, rats were co-exposed to gentamicin (40 mg kg(-1) day(-1), 7 days). Gentamicin induced a marked decrease in the GFR and an increase in urinary levels as well as the protein and mRNA expression of biomarkers of early kidney injury, such as Kim-1. Interestingly, gentamicin nephrotoxicity was less pronounced in groups previously exposed to fluoride than in the group only treated with gentamicin. Fluoride induced Hsp72, a cytoprotective molecule, which might have improved the response against gentamicin. Moreover, fluoride decreased the expression of megalin, a molecule necessary for internalization of gentamicin into the proximal tubule, potentially reducing gentamicin accumulation. The present results suggest that fluoride reduced gentamicin-induced nephrotoxicity by inducing a compensatory response carried out by Hsp72 and by decreasing gentamicin accumulation. These findings should not be interpreted to suggest that fluoride is a protective agent as megalin deficiency could lead to serious adverse effects on the kidney physiology.

Evaluation of renal function in precarious workers exposed to heavy metals in vulnerable scenarios in the metropolitan area of San Luis Potosí, México.

The aim of the study was to evaluate renal function in three groups of precarious workers: garbage recyclers (REC), quarry workers (CAN), and brick makers (LAD). Samples of urine and blood were collected to evaluate clinical parameters and the metal levels in urine was measured using ICP-MS. REC group had the highest concentrations of chromium in urine (36.03 ± 27.2 µg/l) compared to CAN and LAD groups. Mercury concentrations were higher in the LAD group (3.7 ± 0.8 µg/l). Additionally, arsenic was detected in both CAN and REC groups (25.4 ± 26.2 and 19.09 ± 16.7 µg/l, respectively), while arsenic concentrations in LAD were higher (47.2 ± 30.8 µg/l). In kidney biomarkers, ß2-microglobulin concentrations were higher in the REC group (87867 ± 115159.5 ng/g UCr). Similarly, cystatin-C concentrations were higher in the REC group (32795.61 ± 34965.8 ng/g UCr). The data suggests that precarious workers are exposed to heavy metals and have elevated protein levels that contribute to kidney damage.

Assessment of biomarkers of early kidney damage and exposure to pollutants in artisanal mercury mining workers from Mexico.

Artisanal mercury mining (AMM) is an informal economic activity that employs low technology and limited protection, and poses a risk to workers and their families; due to the extraction process, these scenarios involve exposure to complex mixtures of pollutants that synergistically aggravate the health of miners and people living near the site. Although mercury is the predominant pollutant, there are others such as polycyclic aromatic hydrocarbons (PAHs), toluene, arsenic, and lead which have been classified as nephrotoxic pollutants. Therefore, the purpose of this research was to evaluate the association between exposure to a complex mixture of pollutants (mercury, lead, arsenic, PAHs, and toluene) and kidney damage in artisanal Hg mining workers through early kidney damage proteins (KIM-1, OPN, RBP-4, NGAL, and Cys-C). The results demonstrate the presence of OH-PAHs at concentrations of 9.21 (6.57-80.63) µg/L, hippuric acid as a biomarker of exposure to toluene, As and Pb (655. 1 (203.8-1231) mg/L, 24.05 (1.24-42.98) g/g creatinine, and 4.74 (2.71-8.14) g/dL, respectively), and urinary Hg (503.4 (177.9-878.7) g/g creatinine) in the study population. As well as biomarkers of kidney damage, NGAL and RPB-4 were found in 100% of the samples, KIM-1 and Cys-C in 44.1%, and OPN in 41% of the miners. Significant correlations were found between several of the evaluated pollutants and early kidney damage proteins. Our results demonstrate the application of the early kidney damage biomarkers for the assessment of damage caused by the exposure to mixtures of pollutants and, therefore, the urgent need for monitoring in AMM areas.

Renal adaptive response to exposure to low doses of uranyl nitrate and sodium fluoride in mice.

BACKGROUND: Despite their differences in physicochemical properties, both uranium (U) and fluoride (F) are nephrotoxicants at high doses but their adverse effects at low doses are still the subject of debate. METHODS: This study aims to improve the knowledge of the biological mechanisms involved through an adaptive response model of C57BL/6 J mice chronically exposed to low priming doses of U (0, 10, 20 and 40 mg/L) or F (0, 15, 30 and 50 mg/L) and then challenged with acute exposure of 5 mg/kg U or 7.5 mg/kg NaF. RESULTS: We showed that an adaptive response occurred with priming exposures to 20 mg/L U and 50 mg/L F, with decreased levels of the biomarkers KIM-1 and CLU compared to those in animals that received the challenge dose only (positive control). The adaptive mechanisms involved a decrease in caspase 3/7 activities in animals exposed to 20 mg/L U and a decrease in in situ VCAM expression in mice exposed to 50 mg/L F. However, autophagy and the UPR were induced independently of priming exposure to U or F and could not be identified as adaptive mechanisms to U or F. CONCLUSION: Taken together, these results allow us to identify renal adaptive responses to U and F at doses of 20 and 50 mg/L, probably through decrease apoptosis and inflammatory cell recruitment.

Evaluation of hydroxylated metabolites of polycyclic aromatic hydrocarbons and biomarkers of early kidney damage in indigenous children from Ticul, Yucatán, Mexico.

Polycyclic aromatic hydrocarbons (PAHs) are environmental persistent chemicals, produced by the incomplete combustion of solid fuels, found in smoke. PAHs are considered carcinogenic, teratogenic, and genotoxic. Children are susceptible to environmental pollutants, particularly those living in high-exposure settings. Therefore, the main objective of this study was to evaluate the exposure to PAHs through hydroxylated metabolites of PAHs (OH-PAHs), 1-hydroxynaphtalene (1-OH-NAP), and 2-hydroxynaphtalene (2-OH-NAP); 2-,3-, and 9-hydroxyfluorene (2-OH-FLU, 3-OH-FLU, 9-OH-FLU); 1-,2-,3-, and 4-hydroxyphenanthrene (1-OH-PHE, 2-OH-PHE, 3-OH-PHE, 4-OH-PHE); and 1-hydroxypyrene (1-OH-PYR), as well as kidney health through biomarkers of early kidney damage (osteopontin (OPN), neutrophil gelatinase-associated lipocalin (NGAL), α1-microglobulin (α1-MG), and cystatin C (Cys-C)) in children from an indigenous community dedicated to footwear manufacturing and pottery in Ticul, Yucatán, Mexico. The results show a high exposure to PAHs from the found concentrations of OH-PAHs in urine in 80.5% of the children in median concentrations of 18.4 (5.1-71.0) µg/L of total OH-PAHs, as well as concentrations of kidney damage proteins in 100% of the study population in concentrations of 4.8 (3-12.2) and 7.9 (6.5-13.7) µg/g creatinine of NGAL and Cys-C respectively, and 97.5% of the population with concentrations of OPN and α1-MG at mean concentrations of 207.3 (119.8-399.8) and 92.2 (68.5-165.5) µg/g creatinine. The information provided should be considered and addressed by the health authorities to establish continuous biomonitoring and programs to reduce para-occupational exposure in the vulnerable population, particularly children, based on their fundamental human right to health.

In Vivo Comparison of the Phenotypic Aspects and Molecular Mechanisms of Two Nephrotoxic Agents, Sodium Fluoride and Uranyl Nitrate.

Because of their nephrotoxicity and presence in the environment, uranium (U) and fluoride (F) represent risks to the global population. There is a general lack of knowledge regarding the mechanisms of U and F nephrotoxicity and the underlying molecular pathways. The present study aims to compare the threshold of the appearance of renal impairment and to study apoptosis and inflammation as mechanisms of nephrotoxicity. C57BL/6J male mice were intraperitoneally treated with a single dose of U (0, 2, 4 and 5 mg/kg) or F (0, 2, 5, 7.5 and 10 mg/kg) and euthanized 72 h after. Renal phenotypic characteristics and biological mechanisms were evaluated by urine biochemistry, gene/protein expression, enzyme activity, and (immuno)histological analyses. U and F exposures induced nephrotoxicity in a dose-dependent manner, and the highest concentrations induced severe histopathological alterations as well as increased gene expression and urinary excretion of nephrotoxicity biomarkers. KIM-1 gene expression was induced starting at 2 mg/kg U and 7.5 mg/kg F, and this increase in expression was confirmed through in situ detection of this biomarker of nephrotoxicity. Both treatments induced inflammation as evidenced by cell adhesion molecule expression and in situ levels, whereas caspase 3/7-dependent apoptosis was increased only after U treatment. Overall, a single dose of F or U induced histopathologic evidence of nephrotoxicity renal impairment and inflammation in mice with thresholds under 7.5 mg/kg and 4 mg/kg, respectively.

Fluoride exposure is associated with altered metabolism of arsenic in an adult Mexican population.

Arsenic (As) and fluoride (F) are two common groundwater toxicants. The toxicity of As is closely related to As metabolism, and several biological and environmental factors have been associated with As modification. However, limited information about the effect of F exposure on the modification of the As metabolism profile has been described. The aim of this study was to assess the interaction effect of AsF coexposure on the As metabolism profile in an adult population environmentally exposed to low-moderate As levels. A cross-sectional study was conducted in 236 adults from three Mexican communities. F and As concentrations were quantified in water samples. The concentrations of urinary F and As species [inorganic arsenic (iAs), monomethylated arsenic (MAs) and dimethylated arsenic (DMAs)] were also determined and used as exposure biomarkers. As species percentages and methylation indices were estimated to evaluate the As methylation profile. Our results showed a relationship between the water and urine concentrations of both contaminants and, a significant correlation between the As and F concentrations in water and urine samples. A statistically significant interaction of F and As exposure on the increase in MAs% (ß = 0.16, p = 0.018) and the decrease in DMAs% (ß = -0.3, p = 0.034), PMI (ß = -0.07, p = 0.052) and SMI (ß = -0.13, p = 0.097) was observed. These findings indicate that drinking water is the main source of AsF coexposure and suggest that F exposure decreases As methylation capacity. However, additional large and prospective studies are required to confirm our findings, and to elucidate the involved mechanisms of interaction and their implications in adverse health effects.

Chronic Kidney Disease in Children Aged 6-15 Years and Associated Risk Factors in Apizaco, Tlaxcala, Mexico, a Pilot Study.

INTRODUCTION: Tlaxcala, a small state in central Mexico, has the highest prevalence of chronic kidney disease (CKD) deaths in population aged 5-14 in Mexico, most of them with unknown etiology. OBJECTIVE: To determine the prevalence of CKD in apparently healthy pediatric population in Apizaco, Tlaxcala. METHODS: A cross-sectional pilot study was carried out in children deemed as healthy; subjects with previous diagnosis of CKD were excluded. Informed consent was obtained in all cases. A physical examination was performed, a questionnaire was applied. Blood and urine samples were obtained for serum creatinine, urinalysis, and microalbumin/creatinine ratio. A second and third evaluation was performed after 6 and 18 months in those found with urinary anomalies/CKD to confirm the diagnosis. RESULTS: One hundred and nine subjects completed physical examination, which are the biological samples. Median age was 12 years. CKD stage 2 was confirmed in 5 subjects in the sixth month confirmation visit (4.6%). One patient accepted renal biopsy and Alport Syndrome was found. In a robust multivariate analysis, the risk factors related to reduction in the glomerular filtration rate were males -5.15 mL/min/1.73 m2 (p = 0.002), older participants as by -1.58 mL/min/1.73 m2 per year (p < 0.0001), and among participants living close to a river -3.76 mL/min/1.73 m2 (p = 0.033). DISCUSSION/CONCLUSION: The prevalence of CKD in the population studied in Apizaco Tlaxcala was confirmed in 4.6 cases per 100 inhabitants between 6 and 15 years. Males, older age, and living close to a river were the risk predictive factors. More studies are needed to determine the causes of the high CKD prevalence in this population.