Modifying effect of COMT gene polymorphism and a predictive role for proteomics analysis in children’s intelligence in endemic fluorosis area in Tianjin, China.

Abstract

Cumulative fluoride exposure has adverse influences on children’s intelligence quotient (IQ). In addition, Catechol-O-methyltransferase (COMT) gene Val158Met polymorphism (rs4680) is associated with cognitive performance. This study aimed to evaluate the associations of COMT polymorphism and alterations of protein profiles with children’s intelligence in endemic fluorosis area. We recruited 180 schoolchildren (10-12 years old) from high fluoride exposure (1.40 mg/L) and control areas (0.63 mg/L) in Tianjin City, China. The children’s IQ, fluoride contents in drinking water (W-F), serum (S-F) and urine (U-F); serum thyroid hormone levels, COMT Val158Met polymorphism, and plasma proteomic profiling were determined. Significant high levels of W-F, S-F, U-F, along with poor IQ scores were observed in the high fluoride exposure group compared with those in control (all P < 0.05). S-F and U-F were inversely related with IQ (r_s = – 0.47, P < 0.01; r_s= – 0.45, P = 0.002). Importantly, higher fluoride exposure was associated with steeper cognitive decline among children with the reference allele Val compared to those homozygous or heterozygous for the variant allele Met (95% CI, -16.80 to 2.55; P interaction < 0.01). Additionally, 5 up-regulated protein spots related to cell immunity and metabolism were detected in children with high fluoride exposure compared to the control. In conclusion, fluoride exposure was adversely associated with children’s intelligence, whereas the COMT polymorphism may increase the susceptibility to the deficits in IQ due to fluoride exposure. Moreover, the proteomic analysis can provide certain basis for identifying the early biological markers of fluorosis among children.

EXCERPTS:

Children’s IQ scores.

A Combined Raven’s Test for Rural China (CRT-RC) was taken to evaluate the IQ of each child (Wang, 2007). All tests were administered at school by a trained examiner who was masked to participants’ drinking water fluoride levels. The 7 categories of this test scores are as follows: 69 retarded (low);
70–79 borderline (below average);
80–89 dull normal (low average);
90–109 normal (average);
110–119 high normal (high average);
120–129 superior (good); and
130 very superior (excellent).

RESULTS

Sociodemographic and Clinical Characteristics
Descriptive data of the children and fluoride contents are listed in Table 1. The average age of children was 11.35 years, and approximately one-third of the subjects were boys. Children in the 2 groups were well matched for age and gender. According to their age and exposure duration, children on average have been exposed since birth to the wells used by the household. SF and U-F were significantly higher in children exposed to a higher drinking W-F concentration. Interestingly, there is a pretty clear trend that U-F>W-F>S-F, both in the 2 groups. In regard to the thyroid hormones, we found that TSH in the high fluoride group was notably increased (P¼0.03), whereas T4 was somewhat decreased but did not reach statistical significance (P¼0.06). However, the individual values of thyroid hormones were all within the normal range. For children exposed to high fluoride drinking water, the mean IQ value was notably decreased, whereas the percentage of poor IQ (IQ scores fell below the normal range of 90) was markedly increased, when compared with those of control cohorts (both P<0.01).

For the COMT gene, 15.56% of children had the val/val genotype, 48.89% had the met/val genotype, and 35.56% had the met/met genotype, with variant allele frequency of 0.40. The alleles for gene polymorphism conformed to the Hardy-Weinberg equilibrium (control group: v2¼0.375, P>0.05; high fluoride group: v2¼1.777, P>0.05; total subjects: v2¼0.062, P>0.05), which indicated that samples selected were representative.

Associations among Exposure Markers, IQ, and Risk Factors

Data from both high fluoride and control regions were combined for analysis. As shown in Table 2, S-F and U-F were positively correlated, and both increased with fluoride concentration in drinking water. Among all the children, S-F and U-F were positively correlated with years of residence. In addition, IQ had a negative relationship to S-F and U-F (all P<0.05).

Associations between Fluoride Levels and IQ Scores

To further examine the dose-response relationship of S-F or U-F levels with IQ scores, the children were assigned by their fluoride concentrations in serum and urinary and divided into 9 groups of about 20 in each. The mean value of S-F concentration of the lowest group was 0.003 mg/l and of the highest group was 0.333 mg/l, whereas for U-F was 0.459 mg/l and 3.290 mg/l, respectively. For each group, the IQ difference from mean value was plotted against fluoride concentrations in serum and urinary (Fig. 1). From the first group to the ninth, mean IQ scores declined about 16.40 (S-F) and 14.40 (U-F) points, respectively. Besides, mean IQ scores of the last 5 groups were below the mean value of total IQ scores. Manifestly, there was an inverse dose-response relationship between S-F or U-F levels and IQ scores.

DISCUSSION

… The present study has several limitations. First, the crosssectional observational design does not allow us to determine temporal or causal associations between fluoride and cognition. Second, the study has a relatively small sample size, which limits the power to assess effects of gene-environment interactions on children’s IQ. Third, we did not evaluate the cognitive performance of children using other neurocognitive tests except for CRT-RC. Despite the study limitations, this is the first gene environment study investigating the potential impact of COMT single-nucleotide polymorphism (SNP) on the relationship between children’s cognitive performance and exposure to elemental fluoride.

CONCLUSION
In summary, our data suggest that the intelligence of children is affected by the COMT gene polymorphism and, in particular, this SNP plays a role in modifying the effect of fluoride exposure on cognition. Children with COMT reference allele had a higher risk for cognitive impairments after fluoride exposure. Additionally, proteomics analysis represents early specific markers of developmental fluoride neurotoxicity. Hence, our findings provide certain basis for clarifying the mechanisms and identifying molecular targets of pharmacological interventions for potential delayed therapy.
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