The Relationships Between Blood Pb Levels and Blood Pressure Among Lead-Exposed Workers in China: A Repeated-Measure Study.

OBJECTIVES: To explore the differences in the increase of systolic blood pressure (SBP) and diastolic blood pressure (DBP) in 3 consecutive years among lead (Pb) workers. METHODS: Four hundred forty-eight Pb workers were enrolled in this repeated-measure study. Blood Pb, SBP, and DBP were measured in 2015 to 2017. Repeated measure of analysis of variance was used to compare the differences in the increase of SBP and DBP. RESULTS: The mean SBP values were 124.0/125.5/126.9 mm Hg, and the mean DBP values were 75.4/77.4/77.8 mm Hg from 2015 to 2017. The differences in the increase of SBP and DBP were 2.94/2.42 mm Hg during the 3-year period. The average annual increase of SBP or DBP showed an upward trend in different Pb dose groups ( F = 4.904, P = 0.002; F = 3.612, P = 0.013). CONCLUSIONS: Lead exposure caused average annual increases in SBP and DBP with 0.98 and 0.81 mm Hg, which provided basic data for health surveillance.

Biochar-Dual Oxidant Composite Particles Alleviate the Oxidative Stress of Phenolic Acid on Tomato Seed Germination.

Phenolic acid is a well-known allelochemical, but also a pollutant in soil and water impeding crop production. Biochar is a multifunctional material widely used to mitigate the phenolic acids allelopathic effect. However, phenolic acid absorbed by biochar can still be released. In order to improve the removal efficiency of phenolic acids by biochar, the biochar-dual oxidant (BDO) composite particles were synthesized in this study, and the underlying mechanism of the BDO particles in ameliorating p-coumaric acid (p-CA) oxidative damage to tomato seed germination was revealed. Upon p-CA treatment, the BDO composite particles application increased the radical length, radical surface area, and germination index by 95.0%, 52.8%, and 114.6%, respectively. Compared to using biochar or oxidants alone, the BDO particles addition resulted in a higher removal rate of p-CA and produced more O2•-, HO•, SO4•- and 1O2 radicals via autocatalytic action, suggesting that BDO particles removed phenolic acid by both adsorption and free radical oxidation. The addition of BDO particles maintained the levels of the antioxidant enzyme activity close to the control, and reduced the malondialdehyde and H2O2 by 49.7% and 49.5%, compared to the p-CA treatment. Integrative metabolomic and transcriptomic analyses revealed that 14 key metabolites and 62 genes were involved in phenylalanine and linoleic acid metabolism, which increased dramatically under p-CA stress but down-regulated with the addition of BDO particles. This study proved that the use of BDO composite particles could alleviate the oxidative stress of phenolic acid on tomato seeds. The findings will provide unprecedented insights into the application and mechanism of such composite particles as continuous cropping soil conditioners.

Associations between DNA methylation and genotoxicity among lead-exposed workers in China.

Studies have shown that lead (Pb) exposure caused genotoxicity, however, the underlying mechanisms remain unclear. A mechanism may be via DNA methylation which is one of the most widely studied epigenetic regulations for cellular activities. Whether this is involved in Pb-induced genotoxicity has rarely been studied. Our study aimed to examine whether DNA methylation was associated with Pb exposure and genotoxicity, and to explore its potential mediating roles. A total of 250 Pb-exposed workers were enrolled. Blood lead levels (BLLs) and genotoxic biomarkers (Micronuclei and Comet) were analyzed. Methylation levels at CpG sites of LINE1 and Alu and promoter region of P53, BRCA1, TRIM36 and OGG1 were measured by pyrosequencing. Generalized linear model (GLM) combined with restricted cubic splines (RCS) were used to analyze relationships between Pb exposure, DNA methylation and genotoxicity. Mediation effect was used to explore mediating roles of DNA methylation. The distribution of BLLs was right-skewed and showed wide ranges from 23.7 to 636.2 µg/L with median (P25, P75) being 218.4 (106.1, 313.9) µg/L among all workers. Micronuclei frequencies showed Poisson distribution [1.94 ± 1.88‰] and Comet tail intensity showed normal distribution [1.69 ± 0.93%]. GLM combined with RCS showed that Alu methylation was negatively associated with BLLs, while P53 and OGG1 methylation were positively associated with BLLs. Micronuclei were negatively associated with Alu and TRIM36 methylation but positively with P53 methylation. Comet was positively associated with P53 and BRCA1 methylation. Mediation effect showed that Alu methylation mediated 7% effects on association between Pb exposure and micronuclei, whereas, P53 methylation mediated 14% and BRCA1 mediated 9% effects on association between Pb exposure and Comet. Our data show that Pb exposure induced changes of global and gene-specific DNA methylation which mediated Pb-induced genotoxicity.

Research on germplasm diversity of Amomum villosum. Lour in genuine producing area.

BACKGROUND: Genuine Chinese medicine is produced from medicinal plant cultivated in a specific region and is of better quality and efficacy, more consistently qualified and famous than that from the same medicinal plant cultivated in other regions. The cultivating region of genuine medicinal plant is known as the genuine producing area. Yangchun City, which is in Guangdong Province of China, is a genuine producing area for the famous Chinese medicine Amomi Fructus (also called Sharen). Amomi Fructus is the ripe and dry fruit of the Zingiberaceae plant A. villosum Lour.. A. villosum was introduced from the Persian Gulf region and has been cultivated in China for over 1000 years. Until now there are no reports on screening for good germplasm of A. villosum. METHODS: The contents of volatile oil and bornyl acetate of Amomi Fructus from 14 populations were determined with GC method, and the relative contents of the main chemical components in the volatile oils were determined with GC-MS method. Evaluation and variance analysis of the comprehensive quality of the 14 samples were conducted by means of a multi-indicator entropy-weight TOPSIS model (Technique for Order Preference by Similarity to an Ideal Solution) combined with OPLS-DA (Orthogonal Partial Least Squares Discrimination Analysis) and HCA (Hierarchical Clustering Analysis). The ISSR (Inter-Simple Sequence Repeat) molecular marker technique and the UPGMA (unweighted pair-group method with arithmetic means) were employed to analyze the genetic relationship among A. villosum populations. RESULTS: The contents of volatile oil and bornyl acetate differed significantly among the different populations, but the main chemical component in the volatile oil was the same in all the samples, which was bornyl acetate. OPLS-DA results showed that 9 indicators were the main factors influencing the quality differences among the 14 populations. The entropy-weight TOPSIS results showed that there were significant differences in the comprehensive qualities of the 12 populations from the genuine producing area. The best quality of fruit was found in the genuine producing area of Chunwan Town; the qualities of 33% of genuine fruits were lower than that of non-genuine fruits. Twenty-three DNA fragments were obtained by ISSR-PCR amplification using four ISSR primers, eleven of which were polymorphic loci, which accounted for 47.8%. The similarity coefficients (GS) of different populations of A. villosum ranged from 0.6087 to 0.9565. CONCLUSION: There are significant differences among different populations of A. villosum in terms of the kinds of major chemical components and their contents, comprehensive quality and genetic diversity. The germplasm resources of A. villosum are rich in the genuine producing area. It means superior germplasm could be selected in the area. The comprehensive quality of the fruit of A. villosum from the non-genuine producing area is better than some of that from genuine producing area, proving that the non-genuine producing area can also produce Amomi Fructus with excellent quality.

Occupational lead exposure on genome-wide DNA methylation and DNA damage.

Lead (Pb) exposure can induce DNA damage and alter DNA methylation but their inter-relationships have not been adequately determined. Our overall aims were to explore such relationships and to evaluate underlying epigenetic mechanisms of Pb-induced genotoxicity in Chinese workers. Blood Pb levels (BLLs) were determined and used as individual's Pb-exposure dose and the Comet assay (i.e., % tail DNA) was conducted to evaluate DNA damage. In the screening assay, 850 K BeadChip sequencing was performed on peripheral blood from 10 controls (BLLs ≤100 µg/L) and 20 exposed workers (i.e., 10 DNA-damaged and 10 DNA-undamaged workers). Using the technique, differentially methylated positions (DMPs) between the controls and the exposed workers were identified. In addition, DMPs were identified between the DNA-undamaged and DNA-damaged workers (% tail DNA >2.14%). In our validation assay, methylation levels of four candidate genes were measured by pyrosequencing in an independent sample set (n = 305), including RRAGC (Ras related GTP binding C), USP1 (Ubiquitin specific protease 1), COPS7B (COP9 signalosome subunit 7 B) and CHEK1 (Checkpoint kinase 1). The result of comparisons between the controls and the Pb-exposed workers show that DMPs were significantly enriched in genes related to nerve conduction and cell cycle. Between DNA-damaged group and DNA-undamaged group, differentially methylated genes were enriched in the pathways related to cell cycle and DNA integrity checkpoints. Additionally, methylation levels of RRAGC and USP1 were negatively associated with BLLs (P < 0.05), and the former mediated 19.40% of the effect of Pb on the % tail DNA. These findings collectively indicated that Pb-induced DNA damage was closely related to methylation of genes in cell cycle regulation, and methylation levels of RRAGC were involved in Pb-induced genotoxicity.

Global and gene-specific promoter methylation, and micronuclei induction in lead-exposed workers: A cross-sectional study.

Perturbation of epigenetic regulation is a well-established mechanism for cancer but its role for lead (Pb)-associated toxicity has not been adequately investigated. We aimed to investigate whether occupational Pb exposure is associated with micronuclei (MN) frequency and to further explored the mediating roles of epigenetic gene regulation. All the Pb-exposed workers recruited from a Chinese acid battery factory, blood lead levels (BLLs) and MN frequency in lymphocytes were measured. In addition, methylation levels of seven genes (Line-1, RASSF1A, RUNX3, p16, CYP26C1, hMLH1, p15) were examined among 230 workers. Robust Poisson regression model was used to investigate the association between BLLs and MN frequency. Mediation analysis was used to explore the mediating role of specific DNA methylation. Among total 677 participants, 71% were male, median BLLs was 229.1 µg/L (P25  = 155.5, P75  = 319.3; ranged from 8.9 to 647.7 µg/L), mean MN frequency was 2.5‰ (SD = 1.8‰; ranged from 0 to 9‰). Results from base model, adjusted for age, sex, and body mass index, showed that MN frequency would increase 1.38 (95%confidential interval: 1.34, 1.43) per 100 µg/L increment in BLLs. Using categorized exposure variable analyses, a BLLs dose-response increase in MN frequency was observed: 2.74 (2.13, 3.51), 3.43 (2.73, 4.32), 4.41 (3.89, 5.01) to 6.86 (6.02, 7.81). Mediation analysis indicated that Line-1 methylation significantly mediated 3.6% of the association of BLLs with MN frequency. Occupational Pb exposure induces MN frequency in a dose-response relationship. Part of this association was mediated by Line-1 promotor methylation.

Early occupational exposure to lead on neutrophil-to-lymphocyte ratio and genotoxicity.

BACKGROUND: Lead (Pb) is known to induce detrimental health effects in exposed populations, including hematotoxicity and genotoxicity. Complete blood count (CBC) is a cost-effective and easy way to determine toxicity, and variations in proportion of different types of leukocytes: neutrophil-to-lymphocyte ratio (NLR) and lymphocyte-to-monocyte ratio (LMR) are further evidence of hematotoxicity. However, few studies have been conducted to systematically evaluate effects of occupational Pb exposure on NLR and LMR, and their associations with genotoxicity. OBJECTIVES: Our study was aimed to systematically assess the effects of current occupational Pb exposure on NLR and LMR, and their associations with genotoxicity. METHODS: Our investigation was performed on 1176 workers from a newly built battery factory in North China. The workers had just entered their current job position in recent years and most of them had no previous history of occupational exposure to Pb. Blood lead levels (BLLs) and leukocytes indices were detected for all participants. Cytokinesis-blocked micronucleus assay (MN; n = 675) and alkaline comet assay (% tail DNA; n = 869) were used to assess genotoxicity. Multivariate linear and Poisson regression analyses were conducted to examine associations between leukocytes indices, genotoxic biomarkers and BLLs with adjustment for covariates. Spearman correlation and mediation analyses were used to investigate relationships between NLR and genotoxicity. RESULTS: Among all the exposed workers, NLR increased with increasing BLLs. However, WBC and LMR did not change significantly. Significant and dose-dependent increases in both MN frequencies and % tail DNA were observed among groups with different exposure doses. Compared with the normal NLR group (1.48 ≤ NLR < 4.58), the high NLR group (NLR ≥ 4.58) had higher % tail DNA. In addition, there was a significant and positive association between NLR and % tail DNA among all the workers, and % tail DNA mediated 15% of the effect of Pb on increasing NLR. CONCLUSION: Our large-scale population study shows that Pb exposure increased NLR and induced genotoxicity. There was an association between elevated NLR and DNA damage. In addition, the mediation effect of % tail DNA on the relationship between BLLs and NLR provided mechanistic evidence that certain mechanisms, e.g. inflammation, may be involved in elevation of NLR from Pb exposure. Therefore, NLR may be a convenient and sensitive biomarker for indication of Pb toxicity. Further studies are needed to validate the proposed mechanism and NLR as a biomarker.

Development of a benchmark dose for lead-exposure based on its induction of micronuclei, telomere length changes and hematological toxicity.

BACKGROUND: Excessive lead exposure is associated with adverse health effects. However, there is a lack of systematic investigation using large populations to ascertain acceptable exposure limits. OBJECTIVES: Our study was aimed to identify human exposure-response relationships between lead exposure and health-related outcomes, and to determine a benchmark dose (BMD). METHODS: A total of 1896 participants from a lead-acid battery plant were recruited. Blood lead levels (BLLs) were detected for all participants. Hematological parameters (n = 1896), micronuclei (MN) frequencies (n = 934), and relative telomere length (rTL) (n = 757) were also determined. Multivariate linear/Poisson regression analyses were performed to examine associations between BLLs and these health outcomes. Restricted cubic splines were used to identify dose-response relationships. Three BMD approaches were used to calculate BMD and its 95% lower confidence limit (BMDL). RESULTS: Among all participants, BLLs show a right-skewed distribution (median, 185.40 µg/L; 25th - 75th percentile, 104.63-271.70 µg/L). There existed significant differences for red blood cell (RBC), hemoglobin (Hb), MN and rTL among different BLL dose groups. After adjusting for possible confounders, all indicators were significantly associated with BLLs. Restricted cubic splines show that there were linear dose-response relationships for RBC and Hb with BLLs, while non-linear for MN and rTL. Results from the three BMD approaches indicate that the dichotomous models were better than continuous models to calculate BMD and BMDL of BLLs. The conservative BMDL obtained from RBC data was 135 for total, 104 for male and 175 µg/L for female. The corresponding BMDL obtained from Hb data was 105 for total, 116 for male and 70 µg/L for female. As for MN data, the BMDL estimate was 66 for total, 69 for male and 64 µg/L for female. Finally, the BMDL from rTL data was 35 for total, 32 for male and 43 µg/L for female. CONCLUSIONS: Our data show significant dose-response relationships between lead exposure and expressions of hematological toxicity and genotoxicity. The new BMDLs of 135 and 105 µg/L based on RBC and Hb, and even more strict level of 66 and 35 µg/L based on MN and rTL are lower than current exposure limits in China. Therefore, the four values can be considered as novel exposure limits. In addition, sex effect should be taken into account when setting occupational health standard. Considering that different biomarkers have different sensitivities, better understanding their relationships will certainly improve the current emphasis on precision health risk assessment.

Associations of blood lead levels with multiple genotoxic biomarkers among workers in China: A population-based study.

Carcinogenic effects from low doses of lead (Pb) exposure to populations have been suspected but not concluded. Therefore, a large-scale cross-sectional study was conducted by us to investigate genotoxic effects from Pb exposure during 2016-2018 in North China. Blood lead levels (BLLs) and cumulative blood lead levels (CBLLs) were measured. Multiple relevant biomarkers were used to assess genotoxicity of Pb: mitochondrial DNA copy number (mtDNAcn, n = 871), Comet Tail Intensity (n = 872), γ-H2AX (n = 345), relative telomere length (rTL, n = 757), micronuclei (MN, n = 934) and phosphatidylinositol glycan class A mutation (PIG-A, n = 362). The BLL data show right-skewed distribution, with increase of the median (P25, P75) from 17.4 (8.9, 26.4) µg/dl in 2016 to 18.5 (10.5, 27.2) µg/dl in 2017, and to 20.8 (11.3, 31.0) µg/dl in 2018. Multivariate regression analyses show that mtDNAcn was non-linearly associated with BLLs or CBLLs, i.e. decreased at low levels but increased at the higher levels. Comet and Micronuclei data show positive dose-response relationships with BLLs as well as CBLLs. γ-H2AX data show an overall increased trend with BLLs while rTL data show a shortening trend. No associations were found for PIG-A mutation with Pb exposure. Our findings indicate that current low dose exposure to Pb can still cause health hazards to occupational populations, and the mechanism may be via the induction of DNA & chromosome damage rather than via the mutagenesis pathway.

Relative telomere length and gene expression of shelterin complex proteins among vinyl chloride monomer-exposed workers in China.

Vinyl chloride monomer (VCM) is a confirmed carcinogen. The effects of VCM on telomeres and the gene expression of telomere complex proteins, shelterin, have not been well studied but could be of potential relevance to the carcinogenic mechanism of VCM and the health surveillance of VCM-exposed workers. A group of 241 VCM-exposed workers and 101 internal controls from the same plant in Shandong, China were recruited and quantitative polymerase chain reaction was preformed to measure relative telomere length (RTL) and gene expression of shelterin proteins. VCM cumulative exposure dose (CED) was estimated for the exposed workers. The differences in RTL and gene expression between groups were compared by Wald test fitted with robust regression. Shorter RTL was observed in VCM-exposed workers than in the controls (P < 0.001) and was related to CED of VCM. Shortened RTL was also significantly related to increasing age (P = 0.012) and high blood pressure (P = 0.056). Levels of gene expression of shelterin components in exposed workers were all lower than in controls except increased TIN2 expression, and the gene expression differences in TIN2 and POT1 among exposed and control groups were significant (P = 0.014 for TIN2 and P < 0.001 for POT1, respectively). VCM exposure is found associated with altered telomere length and gene expression of shelterin components. This provides new insights into the potential carcinogenic mechanisms of VCM and could be helpful for the health surveillance for VCM-exposed workers. Environ. Mol. Mutagen. 60:361-367, 2019. © 2018 Wiley Periodicals, Inc.

Basal condensation of Numb and Pon complex via phase transition during Drosophila neuroblast asymmetric division.

Uneven distribution and local concentration of protein complexes on distinct membrane cortices is a fundamental property in numerous biological processes, including Drosophila neuroblast (NB) asymmetric cell divisions and cell polarity in general. In NBs, the cell fate determinant Numb forms a basal crescent together with Pon and is segregated into the basal daughter cell to initiate its differentiation. Here we discover that Numb PTB domain, using two distinct binding surfaces, recognizes repeating motifs within Pon in a previously unrecognized mode. The multivalent Numb-Pon interaction leads to high binding specificity and liquid-liquid phase separation of the complex. Perturbations of the Numb/Pon complex phase transition impair the basal localization of Numb and its subsequent suppression of Notch signaling during NB asymmetric divisions. Such phase-transition-mediated protein condensations on distinct membrane cortices may be a general mechanism for various cell polarity regulatory complexes.

Hypermethylation of CpG islands is associated with increasing chromosomal damage in chinese lead-exposed workers.

Lead is a widely existing environmental pollutant with potential carcinogenicity. To investigate the association of blood lead level (B-Pb) with potential chromosomal damage and cancer, we analyzed micronucleus (MN) frequency of peripheral blood lymphocytes (PBLs) and the methylation status of six human tumor suppressor genes (TSGs) post lead exposure. In the study, 147 lead-exposed workers were divided into two groups according to their B-Pb P50 value, with other 50 lead-unexposed workers as a control group. The cytokinesis-blocked micronucleus (CBMN) assay was performed to detect chromosomal damage of PBLs of both lead-exposed and -unexposed workers. The methylation-specific polymerase chain reaction (MSP-PCR) was further used to examine the methylation status of six TSGs (GSTP1, hMLH1, MGMT, p14, p15, and p16). Results showed that MN frequencies of high B-Pb workers 8.1 ± 3.1‰ and low B-Pb workers 5.7 ± 2.3‰ were significantly higher than that of control group 2.8 ± 1.9‰ (P < 0.01), while the MN frequency of high B-Pb workers was also higher than that of the low B-Pb workers (P < 0.01). The MN frequency in PBLs of lead-exposed group with the methylated TSGs was significantly higher than that in PBLs with the unmethylated TSGs (P < 0.05). Notably, the CpG island methylator phenotype (CIMP) correlated with chromosome damage (P < 0.05). Additionally, workers with high B-Pb had higher chromosome damage than those with low B-Pb (P < 0.05). Taken altogether, the results suggest that lead-exposed workers with CIMP positive and high B-Pb have a higher risk of being vulnerable to tumorigenesis. Environ. Mol. Mutagen. 59:549-556, 2018. © 2018 Wiley Periodicals, Inc.

Catalytic wet air oxidation of 2-chlorophenol over sewage sludge-derived carbon-based catalysts.

A sewage sludge derived carbon-supported iron oxide catalyst (FeSC) was prepared and used in the Catalytic Wet Air Oxidation (CWAO) of 2-chlorophenol (2-CP). The catalysts were characterized in terms of elemental composition, surface area, pHPZC, XRD and SEM. The performances of the FeSC catalyst in the CWAO of 2-CP was assessed in a batch reactor operated at 120°C under 0.9MPa oxygen partial pressure. Complete decomposition of 2-CP was achieved within 5h and 90% Total Organic Carbon (TOC) was removed after 24h of reaction. Quite a straight correlation was observed between the 2-CP conversion, the amount of iron leached in solution and the pH of the reaction mixture at a given reaction time, indicating a strong predominance of the homogeneous catalysis contribution. The iron leaching could be efficiently prevented when the pH of the solution was maintained at values higher than 4.5, while the catalytic activity was only slightly reduced. Upon four successive batch CWAO experiments, using the same FeSC catalyst recovered by filtration after pH adjustment, only a very minor catalyst deactivation was observed. Finally, based on all the identified intermediates, a simplified reaction pathway was proposed for the CWAO of 2-CP over the FeSC catalyst.

Separation and determination of degradation products of acid orange 7 by capillary electrophoresis/capacitively coupled contactless conductivity detector.

Capillary electrophoresis (CE) with capacitively coupled contactless conductivity detector (C(4)D) was developed to separate azo-dyestuff acid orange 7 (AO7) and its six degradation products. The analyzed products were sulfamic acid, oxalic acid, benzenesulfonic acid, 4-hydroxybenzene sulfonic acid, phthalic acid, and 4-aminobenzene sulfonic acid. In developing the method, types and concentrations of running buffers, injecting voltage and time, and applied voltage were tested to obtain optimum conditions to analyze target compounds. The separation was successfully achieved within 10 min using a fused-silica capillary under the following conditions: 20 mmol L(-1) acetate acid buffer, electrokinetic injection of -12 kV × 10 s, and applied voltage of -13 kV. The developed method was applied to analyze degradation products in situ during the reaction of AO7 with Fenton reagent (Fe(II)+H2O2 at pH 4.0).

Monoclonal antibodies against Actinobacillus pleuropneumoniae TonB2 protein expressed in Escherichia coli.

TonB is known to be a bacterial periplasmic protein that transduces proton from the inner membrane to the outer membrane receptor in complex with the ExbB and ExbD proteins. Actinobacillus pleuropneumoniae TonB2 protein is the second TonB protein that is important for iron acquisition and virulence. The TonB2 protein was verified to be immunogenic and could afford partial protection for animals from lethal infection. In the present study, the recombinant TonB2 (rTonB2) was overexpressed in Escherichia coli BL21(DE3) and purified. The rTonB2 was then used as antigen to immunize BALB/c mice for the production of monoclonal antibodies (MAb). Four clones of TonB2-specific MAb secretion hybridomas--2F2, 2G8, 3D2, and 6F10--were selected. The MAbs 2F2, 3D2, and 6F10 were classified as IgG1 isotype and 2G8 was of IgG2a isotype. Western blot and ELISA results indicated that MAbs had specific binding activity to rTonB2. The MAbs generated here will be used for further functional analyses of the TonB2 protein.