LDL receptor-related protein 5 rs648438 polymorphism is associated with the risk of skeletal fluorosis.

To investigate the potential association between LRP5 rs648438 polymorphism and the risk of skeletal fluorosis (SF) was evaluated in a cross-sectional case-control study conducted in Shanxi, China, in 2019. A total of 973 individuals were enrolled in this study, in which cases and controls were 346 and 627, respectively. SF was diagnosed according to the standard WS/192-2008 (China). The LRP5 rs648438 was detected by the multiple PCR and sequencing. LRP5 rs648438 was found to follow a dominant genetic model using a web-based SNP-STATS software. Logistic regression analysis found that the TC/CC genotype of LRP5 rs648438 might be a protective factor for SF. When stratified by gender, this protective effect of TC/CC genotype in rs648438 was pronounced in males. There was an interaction between gender and rs648438 on risk of SF. Our study suggested that TC/CC genotype of rs648438 might be a protective factor for water-drinking-type skeletal fluorosis, especially in male participants.

Research for type 2 diabetes mellitus in endemic arsenism areas in central China: role of low level of arsenic exposure and KEAP1 rs11545829 polymorphism.

Type 2 diabetes mellitus (T2DM) is one of the major public health problems worldwide; both genetic and environmental factors are its risk factors. Arsenic, an environmental pollutant, might be a risk factor for T2DM, but the association of low-to-moderate level arsenic exposure with the risk of T2DM is still inconsistent. Single nucleotide polymorphisms (SNPs) can affect the development of T2DM, but the study on KEAP1 rs11545829 (G>A) SNP is few. In this paper, we explored the effect of KEAP1 rs11545829 (G>A) SNP and low-to-moderate level arsenic exposure on risk of T2DM in a cross-sectional case-control study conducted in Shanxi, China. Total of 938 participants, including 318 T2DM cases and 618 controls, were enrolled. Blood glycosylated haemoglobin (HbA1c) was detected by Automatic Biochemical Analyzer, and participants with HbA1c≧6.5% were diagnosed as T2DM. Urinary total arsenic (tAs, mg/L), as the indicator of arsenic exposure, was detected by liquid chromatography-atomic fluorescence spectrometry (LC-AFS). Genomic DNA was extracted and the genotypes of KEAP1 rs11545829 SNP were examined by multiplex polymerase chain reaction (PCR). The urinary tAs concentration in recruited participants was 0.075 (0.03-0.15) mg/L, and was associated with an increased risk of T2DM (OR = 8.45, 95% CI 2.63-27.17); rs11545829 mutation homozygote AA genotype had a protective effect on risk of T2DM (OR = 0.42, 95 % CI 0.25-0.73). Although this protective effect of AA genotype was found in participants with higher urinary tAs level (>0.032 mg/L) (OR = 0.48, 95% CI 0.26-0.86), there was no interaction effect for arsenic exposure and rs11545829 SNP on risk of T2DM. In addition, BMI modified the association between rs11545829 SNP and the risk of T2DM (RERI = -1.11, 95% CI -2.18-0.04). The present study suggest that low-to-moderate level arsenic exposure may be a risk factor, while KEAP1 rs11545829 SNP mutation homozygote AA genotype may be a protective factor for risk of T2DM, especially for T2DM patients with urinary tAs level>0.032 mg/L.

Association between Bone Morphogenetic Protein 2 Gene Polymorphisms and Skeletal Fluorosis of The Brick-tea Type Fluorosis in Tibetans and Kazakhs, China.

To investigate the potential association between BMP2 single nucleotide polymorphisms (SNPs) and brick-tea-type skeletal fluorosis risk in cross-sectional case-control study conducted in Sinkiang and Qinghai, China, a total of 598 individuals, including 308 Tibetans and 290 Kazakhs, were enrolled. Using the standard WS/192-2008 (China), 221 skeletal fluorosis cases were diagnosed, including 123 Tibetans and 98 Kazakhs. Logistic regressions 2 analysis did not find the association between SNPs (Rs235764, Rs235739 and Rs996544) and skeletal fluorosis. Genetic models, linkage disequilibrium (LD) and haplotype analysis were not found to be associated with risk of skeletal fluorosis after adjustment by age and sex (P>0.05).Our data suggested that Rs 235764, Rs 235739 and Rs 996544 were not linked susceptibility for skeletal fluorosis in our cross-sectional case-control study.

Association between fluoride exposure and kidney function in adults: A cross-sectional study based on endemic fluorosis area in China.

BACKGROUND: The kidney toxicity of fluoride exposure has been demonstrated in animal studies, and a few studies have reported kidney function injury in children with fluoride exposure. However, epidemiological information for the effects of long-term fluoride exposure on adult kidney function remains limited. METHODS: We conducted a cross-sectional investigation in Wenshui County, Shanxi Province to examine the association between fluoride exposure and kidney function in adults, and a total of 1070 adults were included in our study. Urinary fluoride concentrations were measured using the national standardized ion selective electrode method. And markers of kidney function injury (urinary NAG, serum RBP, serum Urea, serum C3, serum UA and serum αl-MG) were measured using automatic biochemical analyzer. Multivariate linear regression analysis and binary logistic regression model were used to assess the relationship between urinary fluoride and markers of kidney function injury. RESULTS: Urinary fluoride was positively correlated with urinary NAG and serum Urea, negatively correlated with serum C3. In multivariate linear regression models, every 1 mg/L increment of urinary fluoride was associated with 1.583 U/L increase in urinary NAG, 0.199 mmol/L increase in serum Urea, 0.037 g/L decrease in serum C3 after adjusting for potential confounding factors. In the binary logistic regression model, higher levels of urinary fluoride were associated with an increased risk of kidney function injury. Determination of kidney function based on urinary NAG, every 1 mg/L increment in the urinary fluoride concentrations was associated with significant increases of 22.8% in the risk of kidney function injury after adjusting for potential confounding factors. Sensitivity analysis for the association between urinary fluoride concentrations and markers of kidney function (urinary NAG, serum Urea, and serum C3) by adjusting for the covariates, it is consistent with the primary analysis. CONCLUSIONS: Our study suggests that long-term fluoride exposure is associated with kidney function in adults, and urinary NAG is a sensitive and robust marker of kidney dysfunction caused by fluoride exposure, which could be considered for the identification of early kidney injury in endemic fluorosis areas.

Association between ALOX15 gene polymorphism and brick-tea type skeletal fluorosis in Tibetans, Kazaks and Han, China.

To evaluate the association between ALOX15 gene polymorphism and skeletal fluorosis (SF), a case-control study was conducted. A total of 1023 individuals, including 308 Tibetans, 290 Kazaks and 425 Han, were enrolled in this study, in which cases and controls were 278 and 745, respectively. SF was diagnosed by X-ray absorptiometry. SNPs were genotyped using the Sequenom Mass ARRAY system. The genotypes of ALOX15 rs7220870, rs2664593 and rs1107852 were not associated with the risk of SF. After reconstructing the haplotype of rs7220870 and rs11078528, the risk effect of haplotype CA was found in Han participants aged ≤45 years or with moderate fluoride intake. Diplotype of CC/CC had a protective effect on SF risk in Han participants; whereas, CA/CC diplotype showed a risk effect on SF risk in participants aged ≥65; Our results provide the first evidence of an association between ALOX15 gene polymorphism and SF risk in Han participants.Abbreviation: SF: Skeletal fluorosis; SNP: Single Nucleotide polymorphism.

Fluoride Exposure from Drinking Water Increases the Risk of Stroke: An Ecological Study in Changwu Town, China.

BACKGROUND: Stroke is a major cause of death globally and the leading cause in China. Excessive fluoride exposure has been linked to cardiovascular conditions related to stroke risk factors such as hypertension, atherosclerosis, dyslipidemia, and cardiomyopathy. However, evidence supporting the association between fluoride exposure and stroke risk is limited. METHODS: We constructed an ecological study in Changwu Town, Heilongjiang Province, China, a typical endemic fluorosis area caused by excessive fluoride exposure from drinking water. We collected demographic data, stroke prevalence, and mortality information from 2017 to 2021. Fluoride exposure data were obtained from the national monitoring project on endemic fluorosis. Water fluoride concentrations were measured using the standardized methods. Trend changes in stroke rates were assessed using annual percentage change (APC). Differences in stroke rates among fluoride exposure groups were analyzed using chi-square tests. RESULTS: From 2017 to 2021, the all-ages and age-standardized stroke prevalence rates of permanent residents in Changwu Town increased year by year, while the all-ages and age-standardized mortality rates did not change significantly. The prevalence rates of stroke were significantly higher in endemic fluorosis areas compared to non-endemic areas (p < 0.001). Stratifying the population into tertile groups based on the water fluoride cumulative exposure index (WFCEI) revealed statistically significant differences in stroke prevalence rates (p < 0.001), showing a dose-response relationship with the WFCEI. However, the all-ages and age-standardized mortality rates of stroke were not found to be related to fluoride exposure. CONCLUSIONS: Long-term excessive fluoride exposure from drinking water may increase the risk of stroke prevalence, indicating fluoride overexposure as a potential risk factor for stroke.

Effect of Fluoride in Drinking Water on Fecal Microbial Community in Rats.

Intestinal nutrition has a close association with the onset and development of fluorosis. Intestinal microbes play a major role in intestinal nutrition. However, the effect of fluoride on intestinal microbes is still not fully understood. This study aimed to evaluate the dose-response of fluoride on fecal microbes as well as the link between fluorosis and fecal microbes. The results showed that fluoride did not significantly alter the diversity of fecal microbiota, but richness estimators (ACE and Chao) increased first, and then decreased with the increase of water fluoride. At the genus level, 150 mg/L fluoride significantly reduced the abundances of Roseburia and Clostridium sensu stricto, and 100 mg/L and 150 mg/L fluoride obviously increased the abundances of Unclassified Ruminococcaceaes and Unclassified Bdellovibrionales, respectively. The correlation analysis showed fluoride exposure had a negative association with Roseburia and Turicibacter and was positively associated with Pelagibacterium, Unclassified Ruminococcaceae, and Unclassified Bdellovibrionales. Dental fluorosis was negatively associated with Clostridium sensu stricto, Roseburia, Turicibacter, and Paenalcaligenes and had a positive association with Pelagibacterium, Unclassified Ruminococcaceae, and Unclassified Bdellovibrionales. In conclusion, this study firstly reports fluoride in drinking water has a remarkable biphasic effect on fecal microbiota in rats, and some bacteria are significantly associated with fluoride exposure and dental fluorosis. These results indicate the gut microbiota may play an important role in fluorosis, and some bacteria are likely to be developed as biomarkers for fluorosis.

miR-21-5p and canonical Wnt signaling pathway promote osteoblast function through a feed-forward loop induced by fluoride.

Long-term excessive exposure to fluoride from environmental sources can cause serious public health problems such as dental fluorosis and skeletal fluorosis. The aberrant activation of osteoblasts in the early stage is one of the critical steps during the pathogenesis of skeletal fluorosis and canonical Wnt signaling pathway participate in the progress. However, the specific mechanism that how canonical Wnt signaling pathway was mediated is not yet clear. In this study, we found that miR-21-5p induced the activation of canonical Wnt signaling pathway via targeting PTEN and DKK2 during fluoride induced osteoblasts activation and firstly demonstrated the forward loop between canonical Wnt signaling and miR-21-5p in the process. These findings suggested an important regulatory role of miR-21-5p on canonical Wnt signaling pathway during skeletal fluorosis and miR-21-5p might be a potential therapeutic target for skeletal fluorosis.

Upregulation of miR-200c-3p induced by NaF promotes endothelial apoptosis by activating Fas pathway.

Fluoride has been considered as a risk factor of cardiovascular disease due to its endothelial toxicology. However, the mechanism underlying the endothelial toxicity of fluoride has not been clearly illustrated. MiR-200c-3p was strongly linked with endothelial function and its level is increased in serum of fluorosis patients, but it is unclear the role of miR-200c-3p in the fluoride induced endothelial dysfunction. In this study, we confirmed that fluoride exposure induced the apoptosis of endothelial cells both in established rats model and cultured human umbilical vein endothelial cells (HUVECs). And miR-200c-3p was found to be upregulated in NaF treated HUVECs. Fluoride stimulation increased caspase-dependent apoptosis through miR-200c-3p upregulation, with repressing expression of its target gene Fas-associated phosphatase 1 (Fap-1), which functioned as Fas inhibitor. This resulted in activation of Fas-associated extrinsic apoptosis via interaction with increased Fas, Fadd, Cleaved Caspase-8 and Cleaved Caspase-3. The activation of Fas-associated extrinsic apoptosis was abrogated by miR-200c-3p inhibitor. Furthermore, the antiapoptotic effect of downregulated miR-200c-3p was restored by Fap-1 siRNA. These results suggested a determinant role of the miR-200c-3p/Fap-1 axis in fluoride induced endothelial apoptosis.

ß-catenin mediates fluoride-induced aberrant osteoblasts activity and osteogenesis.

Excess fluoride in drinking water is an environmental issue of increasing worldwide concern, because of its adverse effect on human health. Skeletal fluorosis caused by chronic exposure to excessive fluoride is a metabolic bone disease characterized by accelerated bone turnover accompanied by aberrant activation of osteoblasts. It is not clear whether Wnt/ß-catenin signaling, an important signaling pathway regulating the function of osteoblasts, mediates the pathogenesis of skeletal fluorosis. A cross-sectional case-control study was conducted in Tongyu County, Jilin Province, China showed that fluoride stimulated the levels of OCN and OPG, resulting in accelerated bone turnover in patients with skeletal fluorosis. To investigate the influence of fluoride on Wnt/ß-catenin signaling pathway, 64 male BALB/c mice were allotted randomly to four groups and treated with deionized water containing 0, 55, 110 and 221 mg/L NaF for 3 months, respectively. The results demonstrated that fluoride significantly increased mouse cancellous bone formation and the protein expression of Wnt3a, phospho-GSK3ß (ser 9) and Runx2. Moreover, partial correlation analysis indicated that there was no significant correlation between fluoride exposure and Runx2 protein levels, after adjusting for ß-catenin, suggesting that ß-catenin might play a crucial role in fluoride-induced aberrant osteogenesis. In vivo, viability of SaoS2 cells was significantly facilitated by 4 mg/L NaF, and fluoride could induce the abnormal activation of Wnt/ß-catenin signaling, the expression of its target gene Runx2 and significantly increased Tcf/Lef reporter activity. Importantly, inhibition of ß-catenin suppressed fluoride-induced Runx2 protein expression and the osteogenic phenotypes. Taken together, the present study provided in vivo and in vitro evidence reveals a potential mechanism for fluoride-induced aberrant osteoblast activation and indicates that ß-catenin is the pivot molecule mediating viability and differentiation of osteoblasts and might be a therapeutic target for skeletal fluorosis.

sKlotho is associated with the severity of brick tea-type skeletal fluorosis in China.

The change of serum soluble Klotho (sKlotho) content is related to a variety of osteoarthropathy. However, its association with the severity of skeletal fluorosis (SF) is not clear. Here, the association of tea fluoride exposure with serum sKlotho levels and the severity of SF were investigated and further verified in a rat model of fluorosis. A cross sectional case control study was conducted in residents over 50 years old from brick-tea drinking areas in Qinghai and Xinjiang Provinces, China. Concentrations of fluoride in brick tea water and urine were determined by ion selective electrode method, and the levels of serum sKlotho were determined by ELISA method. Linear regression and ordered logistic regression models were constructed to examine the relationship among fluoride exposure, serum sKlotho levels and the severity of SF. The kidney and small intestine of Wistar rats were isolated for detection of Klotho by immunohistochemistry (IHC), and femoral artery blood was sampled to measure the serum levels of sKlotho. An increase of 1 mg/day in tea fluoride intake (TFI) was associated with a 12.070 pg/mL (95% CI: 0.452-23.689) increase in serum sKlotho levels and a 1.163-fold (95% CI: 1.007-1.342) increase in the severity of SF after adjusting for age, gender, and ethnicity. Serum sKlotho levels were also positively associated with the severity of SF (P < 0.05). The mediation analysis showed that serum sKlotho levels mediated 17.76% of the increase in the severity of SF caused by an increase of 1 mg/day of TFI. Moreover, a significant increase of serum sKlotho levels in fluoride-exposed groups was also seen in the rat model. The present study suggests that serum sKlotho may be a potential mediator of SF in brick tea-type fluorosis endemic areas.