Genetic polymorphisms of GRIN2A and GRIN2B modify the neurobehavioral effects of low-level lead exposure in children.

Lead (Pb) is neurotoxic and children are highly susceptible to this effect, particularly within the context of continuous low-level Pb exposure. A current major challenge is identification of children who may be uniquely susceptible to Pb toxicity because of genetic predisposition. Learning and memory are among the neurobehavioral processes that are most notably affected by Pb exposure, and modification of N-methyl-D-aspartate receptors (NMDAR) that regulate these processes during development are postulated to underlie these adverse effects of Pb. We examined the hypothesis that polymorphic variants of genes encoding glutamate receptor, ionotropic, NMDAR subunits 2A and 2B, GRIN2A and GRIN2B, exacerbate the adverse effects of Pb exposure on these processes in children. Participants were subjects who participated as children in the Casa Pia Dental Amalgam Clinical Trial and for whom baseline blood Pb concentrations and annual neurobehavioral test results over the 7 year course of the clinical trial were available. Genotyping assays were performed for variants of GRIN2A (rs727605 and rs1070503) and GRIN2B (rs7301328 and rs1806201) on biological samples acquired from 330 of the original 507 trial participants. Regression modeling strategies were employed to evaluate the association between genotype status, Pb exposure, and neurobehavioral test outcomes. Numerous significant adverse interaction effects between variants of both GRIN2A and GRIN2B, individually and in combination, and Pb exposure were observed particularly among boys, preferentially within the domains of Learning & Memory and Executive Function. In contrast, very few interaction effects were observed among similarly genotyped girls with comparable Pb exposure. These findings support observations of an essential role of GRIN2A and GRIN2B on developmental processes underlying learning and memory as well as other neurological functions in children and demonstrate, further, modification of Pb effects on these processes by specific variants of both GRIN2A and GRIN2B genes. These observations highlight the importance of genetic factors in defining susceptibility to Pb neurotoxicity and may have important public health implications for future strategies aimed at protecting children and adolescents from potential health risks associated with low-level Pb exposure.

Genetic polymorphisms affecting susceptibility to mercury neurotoxicity in children: summary findings from the Casa Pia Children's Amalgam clinical trial.

Mercury (Hg) is neurotoxic, and children may be particularly susceptible to this effect. A current major challenge is identification of children who may be uniquely susceptible to Hg toxicity because of genetic predisposition. We examined the possibility that common genetic variants that are known to affect neurologic functions or Hg handling in adults would modify the adverse neurobehavioral effects of Hg exposure in children. Three hundred thirty subjects who participated as children in the recently completed Casa Pia Clinical Trial of Dental Amalgams in Children were genotyped for 27 variants of 13 genes that are reported to affect neurologic functions and/or Hg disposition in adults. Urinary Hg concentrations, reflecting Hg exposure from any source, served as the Hg exposure index. Regression modeling strategies were employed to evaluate potential associations between allelic status for individual genes or combinations of genes, Hg exposure, and neurobehavioral test outcomes assessed at baseline and for 7 subsequent years during the clinical trial. Among boys, significant modification of Hg effects on neurobehavioral outcomes over a broad range of neurologic domains was observed with variant genotypes for 4 of 13 genes evaluated. Modification of Hg effects on a more limited number of neurobehavioral outcomes was also observed for variants of another 8 genes. Cluster analyses suggested some genes interacting in common processes to affect Hg neurotoxicity. In contrast, significant modification of Hg effects on neurobehavioral functions among girls with the same genotypes was substantially more limited. These observations suggest increased susceptibility to the adverse neurobehavioral effects of Hg among children, particularly boys, with genetic variants that are relatively common to the general human population. These findings advance public health goals to identify factors underlying susceptibility to Hg toxicity and may contribute to strategies for preventing adverse health risks associated with Hg exposure.

Genetic polymorphisms of catechol-O-methyltransferase modify the neurobehavioral effects of mercury in children.

Mercury (Hg) is neurotoxic and children may be particularly susceptible to this effect. A current major challenge is identification of children who may be uniquely susceptible to Hg toxicity because of genetic disposition. This study examined the hypothesis that genetic variants of catechol-O-methyltransferase (COMT) that are reported to alter neurobehavioral functions that are also affected by Hg in adults might modify the adverse neurobehavioral effects of Hg exposure in children. Five hundred and seven children, 8-12 yr of age at baseline, participated in a clinical trial to evaluate the neurobehavioral effects of Hg from dental amalgam tooth fillings. Subjects were evaluated at baseline and at seven subsequent annual intervals for neurobehavioral performance and urinary Hg levels. Following the clinical trial, genotyping assays were performed for single-nucleotide polymorphisms (SNPs) of COMT rs4680, rs4633, rs4818, and rs6269 on biological samples provided by 330 of the trial participants. Regression-modeling strategies were employed to evaluate associations between allelic status, Hg exposure, and neurobehavioral test outcomes. Similar analysis was performed using haplotypes of COMT SNPs. Among girls, few interactions for Hg exposure and COMT variants were found. In contrast, among boys, numerous gene-Hg interactions were observed between individual COMT SNPs, as well as with a common COMT haplotype affecting multiple domains of neurobehavioral function. These findings suggest increased susceptibility to the adverse neurobehavioral effects of Hg among children with common genetic variants of COMT, and may have important implications for strategies aimed at protecting children from the potential health risks associated with Hg exposure.

Modification of neurobehavioral effects of mercury by genetic polymorphisms of metallothionein in children.

Mercury (Hg) is neurotoxic, and children may be particularly susceptible to this effect. A current major challenge is the identification of children who may be uniquely susceptible to Hg toxicity because of genetic disposition. We examined the hypothesis that genetic variants of metallothionein (MT) that are reported to affect Hg toxicokinetics in adults would modify the neurotoxic effects of Hg in children. Five hundred seven children, 8-12 years of age at baseline, participated in a clinical trial to evaluate the neurobehavioral effects of Hg from dental amalgam tooth fillings. Subjects were evaluated at baseline and at 7 subsequent annual intervals for neurobehavioral performance and urinary Hg levels. Following the completion of the clinical trial, we performed genotyping assays for variants of MT isoforms MT1M (rs2270837) and MT2A (rs10636) on biological samples provided by 330 of the trial participants. Regression modeling strategies were employed to evaluate associations between allelic status, Hg exposure, and neurobehavioral test outcomes. Among girls, few significant interactions or independent main effects for Hg exposure and either of the MT gene variants were observed. In contrast, among boys, numerous significant interaction effects between variants of MT1M and MT2A, alone and combined, with Hg exposure were observed spanning multiple domains of neurobehavioral function. All dose-response associations between Hg exposure and test performance were restricted to boys and were in the direction of impaired performance. These findings suggest increased susceptibility to the adverse neurobehavioral effects of Hg among children with relatively common genetic variants of MT, and may have important public health implications for future strategies aimed at protecting children and adolescents from the potential health risks associated with Hg exposure. We note that because urinary Hg reflects a composite exposure index that cannot be attributed to a specific source, these findings do not support an association between Hg in dental amalgams specifically and the adverse neurobehavioral outcomes observed.

Modification of neurobehavioral effects of mercury by a genetic polymorphism of coproporphyrinogen oxidase in children.

Mercury (Hg) is neurotoxic, and children may be particularly susceptible to this effect. A current major challenge is the identification of children who may be uniquely susceptible to Hg toxicity because of genetic disposition. We examined the hypothesis that CPOX4, a genetic variant of the heme pathway enzyme coproporphyrinogen oxidase (CPOX) that affects susceptibility to mercury toxicity in adults, also modifies the neurotoxic effects of Hg in children. Five hundred seven children, 8-12 years of age at baseline, participated in a clinical trial to evaluate the neurobehavioral effects of Hg from dental amalgam tooth fillings in children. Subjects were evaluated at baseline and at 7 subsequent annual intervals for neurobehavioral performance and urinary mercury levels. Following the completion of the clinical trial, genotyping assays for CPOX4 allelic status were performed on biological samples provided by 330 of the trial participants. Regression modeling strategies were employed to evaluate associations between CPOX4 status, Hg exposure, and neurobehavioral test outcomes. Among girls, few significant CPOX4-Hg interactions or independent main effects for Hg or CPOX4 were observed. In contrast, among boys, numerous significant interaction effects between CPOX4 and Hg were observed spanning all 5 domains of neurobehavioral performance. All underlying dose-response associations between Hg exposure and test performance were restricted to boys with the CPOX4 variant, and all of these associations were in the expected direction where increased exposure to Hg decreased performance. These findings are the first to demonstrate genetic susceptibility to the adverse neurobehavioral effects of Hg exposure in children. The paucity of responses among same-age girls with comparable Hg exposure provides evidence of sexual dimorphism in genetic susceptibility to the adverse neurobehavioral effects of Hg in children and adolescents.

Disordered porphyrin metabolism: a potential biological marker for autism risk assessment.

Autism (AUT) is a complex neurodevelopmental disorder that, together with Asperger's syndrome and Pervasive Developmental Disorder-Not Otherwise Specified (PDD-NOS), comprises the expanded classification of autistic spectrum disorder (ASD). The heterogeneity of ASD underlies the need to identify biomarkers or clinical features that can be employed to identify meaningful subtypes of ASD, define specific etiologies, and inform intervention and treatment options. Previous studies have shown that disordered porphyrin metabolism, manifested principally as significantly elevated urinary concentrations of pentacarboxyl (penta) and coproporphyrins, is commonly observed among some children with ASD. Here, we extend these observations by specifically evaluating penta and coproporphyrins as biological indicators of ASD among 76 male children comprising 30 with validated AUT, 14 with PDD-NOS, and 32 neurotypical (NT) controls. ASD children (AUT and PDD-NOS) had higher mean urinary penta (P < 0.006) and copro (P < 0.006) concentrations compared with same-aged NT children, each characterized by a number of extreme values. Using Receiver Operating Characteristic curve analysis, we evaluated the sensitivity and specificity of penta, copro, and their combined Z-scores in ASD detection. The penta sensitivity was 30% for AUT and 36% for PDD-NOS, with 94% specificity. The copro sensitivity was 33% and 14%, respectively, with 94% specificity. The combined Z-score measure had 33% and 21% sensitivity for AUT and PDD-NOS, respectively, with 100% specificity. These findings demonstrate that porphyrin measures are strong predictors of both AUT and PDD-NOS, and support the potential clinical utility of urinary porphyrin measures for identifying a subgroup of ASD subjects in whom disordered porphyrin metabolism may be a salient characteristic.

Urinary porphyrin excretion in neurotypical and autistic children.

BACKGROUND: Increased urinary concentrations of pentacarboxyl-, precopro- and copro-porphyrins have been associated with prolonged mercury (Hg) exposure in adults, and comparable increases have been attributed to Hg exposure in children with autism (AU). OBJECTIVES: This study was designed to measure and compare urinary porphyrin concentrations in neurotypical (NT) children and same-age children with autism, and to examine the association between porphyrin levels and past or current Hg exposure in children with autism. METHODS: This exploratory study enrolled 278 children 2-12 years of age. We evaluated three groups: AU, pervasive developmental disorder-not otherwise specified (PDD-NOS), and NT. Mothers/caregivers provided information at enrollment regarding medical, dental, and dietary exposures. Urine samples from all children were acquired for analyses of porphyrin, creatinine, and Hg. Differences between groups for mean porphyrin and Hg levels were evaluated. Logistic regression analysis was conducted to determine whether porphyrin levels were associated with increased risk of autism. RESULTS: Mean urinary porphyrin concentrations are naturally high in young children and decline by as much as 2.5-fold between 2 and 12 years of age. Elevated copro- (p < 0.009), hexacarboxyl- (p < 0.01) and pentacarboxyl- (p < 0.001) porphyrin concentrations were significantly associated with AU but not with PDD-NOS. No differences were found between NT and AU in urinary Hg levels or in past Hg exposure as determined by fish consumption, number of dental amalgam fillings, or vaccines received. CONCLUSIONS: These findings identify disordered porphyrin metabolism as a salient characteristic of autism. Hg exposures were comparable between diagnostic groups, and a porphyrin pattern consistent with that seen in Hg-exposed adults was not apparent.

The association between serotonin transporter gene promotor polymorphism (5-HTTLPR) and elemental mercury exposure on mood and behavior in humans.

A functional polymorphism in the serotonin transporter (5-HTT) gene-linked polymorphic region (5-HTTLPR) is reported to affect mood and behavior in humans. In this study, the effects of 5-HTTLPR polymorphism on neurobehavioral and mood domains that are known to be affected by elemental mercury (Hg degrees ) exposure in human subjects were examined. The Behavioral Evaluation for Epidemiologic Studies (BEES) test battery was administered concurrently with urine and buccal-cell collections for 164 male dentists (DD) and 101 female dental assistants (DA) with occupational exposure to Hg degrees for an average of 19 and 10 yr, respectively. Geometric mean urinary mercury (Hg) levels in DD and DA were 2.52 (2.22) microg/L and 1.98 (1.98) microg/L, respectively. Corresponding indices of chronic occupational Hg degrees exposure, weighted for historical exposure, were 1212 (1877) and 316 (429). 5-HTTLPR status was 40% and 20% wild type, 40% and 56% single allelic substitution, and 20% and 24% double allelic substitution for the two genders. DD and DA were evaluated separately. Regression analyses controlled for age, premorbid intelligence, frequency of alcohol per week, and education. 5-HTTLPR polymorphism was associated with 5 behavioral measures in DD and with 12 behavioral measures in DA. Mood scores were more consistently associated with the variant in both groups. The strongest evidence for an additive effect for urinary Hg and 5-HTTLPR polymorphism in both groups was for tests of Finger Tap(Alternate) and Hand Steadiness(Factor1). Other significant additive effects that were less consistent across groups were also observed. These results add to the growing evidence of genetic determinants of mood and behavior that potentially increase susceptibility to Hg toxicity in humans.

Urinary porphyrin excretion in children with mercury amalgam treatment: findings from the Casa Pia Children's Dental Amalgam Trial.

Increases in the urinary concentrations of pentacarboxyl- and coproporphyrins and the appearance of the atypical precoproporphyrin have been defined in relation to mercury (Hg) body burden in animal studies, and this change in the porphyrin excretion pattern has been described as a biomarker of occupational Hg exposure and toxicity in adult human subjects. In the present studies, urinary porphyrins were determined in relation to Hg exposure in children and adolescents, 8-18 yr of age, over the 7-yr course of a clinical trial designed to evaluate the neurobehavioral and renal effects of dental amalgam in children. Subjects were randomized to either dental amalgam or composite resin treatments. Urinary porphyrins and creatinine concentrations were measured at baseline and annually in all subjects. Results were evaluated using linear regression analysis. No significant differences between treatment groups (amalgam versus composite) were found when comparing all subjects for any of the porphyrins of interest. However, incipent amalgam treatment-specific increases were observed in the mean concentrations of penta-, precopro- and coproporphyrins especially when the analyses were restricted to younger subjects (8 to 9 yr old at baseline), and these increases were most apparent during yr 2 through 3 of follow-up, the period of highest mercury exposure from amalgam treatment. Based on the mean number of amalgam fillings received by children in this group (17.8), the renal Hg concentration associated with incipient increases in urinary porphyrins was estimated to be approximately 2.7 microg/g renal cortex. This value corresponds to an observed mean urinary Hg concentration of 3.2 microg/g creatinine, which is approximately fivefold less than that at which renal damage from Hg exposure is estimated to occur in children. These findings are consistent with growing evidence supporting the sensitivity of urinary porphyrins as a biological indicator of subclinical Hg exposure in children.

Urinary porphyrin excretion in normal children and adolescents.

BACKGROUND: Urinary porphyrins are diagnostic of various metabolic disorders and xenobiotic exposures, but comprehensive normative data for urinary porphyrin concentrations in children are currently unavailable. METHODS: Subjects were participants in a prospective, randomized, controlled clinical trial of dental materials safety, 8 to 12 y at inception, who were followed longitudinally for 7 y after baseline with an extensive battery of neurobehavioral, neurological, renal function and urinary porphyrin assessments. Porphyrins were quantified by HPLC. Linear regression analyses were used to measure associations of porphyrin levels with age and gender. RESULTS: Mean concentrations, 95% confidence intervals, and 10th, 50th, and 90th percentiles for all 5 typically excreted urinary porphyrins are presented by year of age and by gender. Unadjusted urinary concentrations (microg/l) of all 5 porphyrins remained relatively constant throughout the age range of 8-18 y for both males and females. In contrast, creatinine-adjusted urinary porphyrin concentrations (microg/g) declined significantly throughout this age range in both genders. Boys had significantly higher pentacarboxyl- and copro-porphyrin levels compared with girls both before and after creatinine adjustment. CONCLUSIONS: Normative longitudinal data provided herein may facilitate the clinical assessment of pediatric metabolic disorders and may be of particular relevance in evaluating porphyrin changes as a biological indicator of disease or xenobiotic exposures among children and adolescents.

Cloning, expression, and biochemical properties of CPOX4, a genetic variant of coproporphyrinogen oxidase that affects susceptibility to mercury toxicity in humans.

Coproporphyrinogen oxidase (CPOX) catalyzes the two-step decarboxylation of coproporphyrinogen-III to protoporphyrinogen-IX in the heme biosynthetic pathway. Previously we described a specific polymorphism (A814C) in exon 4 of the human CPOX gene (CPOX4) and demonstrated that CPOX4 is associated with both modified urinary porphyrin excretion and increased neurobehavioral deficits among human subjects with low-level mercury (Hg) exposure. Here, we sought to characterize the gene products of CPOX and CPOX4 with respect to biochemical and kinetic properties. Coproporphyrinogen-III was incubated with recombinantly expressed and purified human CPOX and CPOX4 enzymes at various substrate concentrations, with or without Hg(2+) present. Both CPOX and CPOX4 formed protoporphyrinogen-IX from coproporphyrinogen-III; however, the affinity of CPOX4 was twofold lower than that of CPOX (CPOX K(m) = 0.30 microM, V(max) = 0.52 pmol protoporphyrin-IX; CPOX4 K(m) = 0.54 microM, V(max) = 0.33 pmol protoporphyrin-IX). Hg(2+) specifically inhibited the second step of coproporphyrinogen-III decarboxylation (harderoporphyrinogen to protoporphyrinogen-IX) in a dose dependent manner. We also compared the catalytic activities of CPOX and CPOX4 in human liver samples. The specific activities of CPOX in mutant livers were significantly lower (40-50%) than those of either wild-type or heterozygous. Additionally, enzymes from mutant, heterozygous and wild-type livers were comparably inhibited by Hg(2+) (10 microM), decreasing CPOX4 activity to 25% that of the wild-type enzyme. These findings suggest that CPOX4 may predispose to impaired heme biosynthesis, which is limited further by Hg exposure. These effects may underlie increased susceptibility to neurological deficits previously observed in Hg-exposed humans with CPOX4.

Catechol O-methyltransferase (COMT) VAL158MET functional polymorphism, dental mercury exposure, and self-reported symptoms and mood.

Associations were evaluated between a functional single nucleotide polymorphism (Val158Met) in the gene encoding the catecholamine catabolic enzyme catechol O-methyltransferase (COMT), dental mercury exposure, and self-reported symptoms and mood among 183 male dentists and 213 female dental assistants. Self-reported symptoms, mood, and detailed work histories were obtained by computerized questionnaire. Spot urine samples were collected and analyzed for mercury concentrations to evaluate recent exposures, whereas a chronic mercury exposure index for all subjects was created from the work histories. COMT polymorphism status was determined using a polymerase chain reaction (PCR)-based assay. Scores for current, recent, and chronic self-reported symptom groups and six self-reported mood factors were evaluated with respect to recent and chronic mercury exposure and COMT polymorphism status. Multiple regression analysis controlled for age, socioeconomic status, tobacco and alcohol use, self-reported health problems, and medications. Separate evaluations were conducted for dentists and dental assistants. No consistent patterns of association between either urinary mercury concentration or the chronic index of mercury exposure and any category of symptoms were observed. However, consistent and significant associations were found between increased symptoms and the COMT polymorphism involving the double allelic substitution (full mutation) compared to subjects with no substitutions. Associations with mood were limited to polymorphism status among female dental assistants, and were observed for four of six mood factors and overall mood score. These findings extend evidence of genetic factors potentially affecting human susceptibility to the toxic effects of mercury and other environmental chemicals.

The association between serotonin transporter gene promoter polymorphism (5-HTTLPR), self-reported symptoms, and dental mercury exposure.

The associations between a polymorphism of the serotonin transporter gene (5-HTTLPR), dental mercury exposure, and self-reported symptoms were evaluated among 157 male dentists and 84 female dental assistants. Self-reported symptoms and detailed work histories were obtained by computerized questionnaire. Spot urine samples were collected and analyzed for mercury concentrations to evaluate recent exposures, whereas a chronic mercury exposure index was created from the work histories. 5-HTTLPR polymorphism status was determined using a polymerase chain reaction (PCR)-based assay. Scores for current, recent, and chronic self-reported symptom groups were evaluated with respect to recent and chronic mercury exposure and 5-HTTLPR polymorphism status. Multiple regression analysis controlled for age, socioeconomic status, tobacco and alcohol use, self-reported health problems, and medications. Analyses were restricted to Caucasian subjects due to the highly skewed distribution of the 5-HTTLPR polymorphism. Separate evaluations were conducted for dentists and dental assistants. In contrast to previous reports, no consistent associations were found between either urinary mercury concentration or the chronic index of mercury exposure and any category of symptoms. However, both significant and consistent associations were observed between increased symptoms and the 5-HTTLPR polymorphism involving two copies of the short or "s" allele (full mutation), but not with the polymorphism involving only one copy (heterozygous), demonstrating a gene-dose relationship for symptom reporting. These findings suggest that within this restricted population increased symptoms of depression, anxiety, and memory are associated with the 5-HTTLPR polymorphism among both males and females.

Biomarkers of kidney integrity in children and adolescents with dental amalgam mercury exposure: findings from the Casa Pia children's amalgam trial.

Mercury is toxic to the kidney, and dental amalgam is a source of mercury exposure. Few studies have evaluated the effects of dental amalgam on kidney function in a longitudinal context in children. Here, we evaluated urinary concentrations of glutathione S-transferases (GSTs) alpha and pi as biomarkers of renal proximal and distal tubular integrity, respectively, and albumin as a biomarker of glomerular integrity in children and adolescents 8-18 years of age over a 7-year course of dental amalgam treatment. Five hundred seven children, 8-12 years of age at baseline, participated in a clinical trial to evaluate the neurobehavioral and renal effects of dental amalgam in children. Subjects were randomized to either dental amalgam or resin composite treatments. Urinary GSTs alpha and pi, albumin, and creatinine concentrations were measured at baseline and annually in all subjects. Results were evaluated using linear regression analysis. GST-alpha concentrations were similar between treatment groups and in each sex and race (white vs. non-white) group in each follow-up year. GST-pi levels tended upward over the course of follow-up by four- to six-fold. This increase was seen in all groups irrespective of the treatment, race, or gender. Females had GST-pi levels approximately twice those of males at all ages. Albumin concentrations were constant throughout the follow-up period and did not differ by treatment, although females had 39% higher albumin levels than males. Additionally, we found no significant effects of amalgam treatment on the proportion of children with microalbuminuria (>30 mg/g creatinine). These findings are relevant within the context of children's health risk assessment as relates to the safety of mercury exposure from dental amalgam on kidney function. These data also provide normative values for sensitive indices of renal functional integrity that may serve in the evaluation of children and adolescents with renal disorders.

Neurological outcomes in children with and without amalgam-related mercury exposure: seven years of longitudinal observations in a randomized trial.

BACKGROUND: Although large-scale, randomized trials involving children have been completed and their results demonstrate an absence of neurobehavioral effects from clinical exposure to mercury amalgam, neurological findings from such studies have not been reported. METHODS: The authors conducted a randomized, prospective trial examining the safety of dental amalgam in which 507 children aged 8 through 12 years were assigned to treatment with either amalgam or resin-based composite. During seven years of follow-up, the authors performed annual clinical neurological examinations, including an evaluation of neurological hard signs (NHSs), presence of tremor and neurological soft signs (NSSs). RESULTS: The authors found no significant differences between treatment groups in any of the neurological measures. Groups did not differ with respect to the presence or absence of NHSs or tremor, nor the presence or absence or severity of NSSs at any point. As expected, NSS severity scores diminished with increasing age. CONCLUSIONS: Even at the levels of amalgam exposure in this study (a mean of 7.7-10.7 amalgam surfaces per subject across the seven years of follow-up), the authors conclude that exposure to mercury from dental amalgam does not adversely affect neurological status. CLINICAL IMPLICATIONS: The current evidence is that potential neurobehavioral or neurological effects from dental amalgam mercury exposure in children are inconsequential.

Longitudinal urinary creatinine excretion values among preadolescents and adolescents.

To present longitudinal urinary creatinine excretion data for developmentally normal children 9-17 years of age. Only extremely limited data have been presented of a longitudinal nature for this age group. Overall, 507 children who participated in a prospective, randomized, controlled trial of dental materials safety were followed longitudinally for 7 years with renal measures, including creatinine excretion. Urinary creatinine means, confidence intervals, and 10th, 50th, and 90th percentiles are presented by year of age for the entire group, by sex, and by race (whites, blacks). Urinary creatinine excretion increases with age for both sexes and for both races (P<0.0001). No significant sexual difference were observed, although a race difference occurs, with blacks showing higher excretion levels than whites (P=0.0003). We present longitudinal urinary creatinine excretion data for ages 9-17 in which creatinine excretion increases with age throughout the time period. No sexual differences are observed, although blacks excrete significantly higher levels of urinary creatinine than do whites.

The contribution of dental amalgam to urinary mercury excretion in children.

BACKGROUND: Urinary mercury concentrations are widely used as a measure of mercury exposure from dental amalgam fillings. No studies have evaluated the relationship of these measures in a longitudinal context in children. OBJECTIVE: We evaluated urinary mercury in children 8-18 years of age in relation to number of amalgam surfaces and time since placement over a 7-year course of amalgam treatment. METHODS: Five hundred seven children, 8-10 years of age at baseline, participated in a clinical trial to evaluate the neurobehavioral effects of dental amalgam in children. Subjects were randomized to either dental amalgam or resin composite treatments. Urinary mercury and creatinine concentrations were measured at baseline and annually on all participants. RESULTS: Treatment groups were comparable in baseline urinary mercury concentration (approximately 1.5 microg/L). Mean urinary mercury concentrations in the amalgam group increased to a peak of approximately 3.2 microg/L at year 2 and then declined to baseline levels by year 7 of follow-up. There was a strong, positive association between urinary mercury and both number of amalgam surfaces and time since placement. Girls had significantly higher mean urinary mercury concentrations than boys throughout the course of amalgam treatment. There were no differences by race in urinary mercury concentration associated with amalgam exposure. CONCLUSIONS: Urinary mercury concentrations are highly correlated with both number of amalgam fillings and time since placement in children. Girls excrete significantly higher concentrations of mercury in the urine than boys with comparable treatment, suggesting possible sex-related differences in mercury handling and susceptibility to mercury toxicity.

RhoA regulation of NF-kappaB activation is mediated by COX-2-dependent feedback inhibition of IKK in kidney epithelial cells.

Numerous studies have demonstrated a central role of renal tubular epithelial cells in the etiology of kidney injury and disease through the elaboration of inflammatory mediators. However, little is known about the cellular signaling mechanisms involved in this process. In this study we employed normal rat kidney epithelial (NRK52E) cells to identify a novel LPS-induced signaling pathway in which RhoA-mediated AP-1 activity promotes expression of cyclooxygenase-2 (COX-2) with consequent feedback inhibition of NF-kappaB activation through IKKbeta. Inhibition of RhoA signaling using either the RhoA kinase inhibitor Y-27632 or a dominant negative mutant of RhoA (RhoA-DN) dramatically extended the duration of p65-DNA binding, IkappaBalpha phosphorylation, and IKKbeta activity following LPS treatment. Prolongation of events associated with NF-kappaB activation was also observed in cells pretreated and/or cotransfected with the JNK inhibitor SP600125 or deletion mutants of MEKK1 (MEKK1-KD) or Jun (Jun-DN). Conversely, constitutive expression of RhoA prevented NF-kappaB activation by LPS, and this effect was reversed by cotransfection with MEKK1-KD. In addition, we found that the RhoA/AP-1 signaling axis plays a necessary role in COX-2 expression by LPS and that this effect is independent of NF-kappaB activation. Moreover, inhibition of COX-2 activity results in persistent p65-DNA binding, IkappaBalpha phosphorylation, and IKKbeta activity, similar to that observed after prevention of RhoA/AP-1 axis signaling. These findings suggest that COX-2 links the RhoA/AP-1 signaling cascade to NF-kappaB activation, thereby defining a novel integrated model for regulation of the inflammatory response of kidney epithelial cells to LPS and potentially other external stimuli.