[Effects of Traffic-related Air Pollution Exposure on DNA Methylation].

The goal of the present study was to explore the effects of traffic-related air pollution exposure on DNA methylation. Into five groups of 6, 30 healthy Wistar rats were randomly divided. Three groups of rats were then exposed to traffic-related air pollution at high (tunnel), moderate (crossroad), and low (control) pollution levels for 7 d, whereas the two other groups were exposed in the tunnel for 14 d/28 d. The levels of PM10 and NO2 were measured during the exposure. The study was performed in spring and autumn, and lung tissue and blood were collected after the exposure. Promoter methylation levels of p 53 , MGMT, and MAGE-A 4 were quantified via pyrosequencing. The levels of PM10 and NO2 in the crossroad and tunnel groups were significantly higher than those in the control group. After 7 d exposure in autumn, promoter methylation levels of p 53 and MGMT in lung tissue significantly decreased, and the methylation status continued to decrease with increasing exposure time; MAGE-A 4 was highly methylated and showed no difference among the three groups. DNA methylation in lung tissue was more likely to be changed compared with that in blood during 7 d exposure. As the exposure time increased, DNA methylation changes between blood and lung tissue started to coincide. In lung tissue, PM10 exposure was significantly associated with decreased p 53 promoter methylation (r=-0.347, P=0.038) and NO2 exposure was significantly associated with decreased promoter methylation of p 53, MGMT, and MAGE-A 4 (r=-0.482, -0.444, and -0.346, respectively; P< 0.05). In blood, PM10 and NO2 were significantly and positively associated with MAGE-A 4 promoter methylation (r=0.395 and 0.431, respectively; P< 0.05). Traffic-related air pollution exposure may induce promoter hypomethylation of p 53 and MGMT.

Aberrant promoter methylation of cell adhesion-related genes associated with clinicopathologic features in non-small cell lung cancer in China.

PURPOSE: The aim of this study was to investigate the methylation status of three cell adhesion-related genes including CDH1, TSLC1 and TIMP3 in non-small cell lung cancer and explore its association with clinicopathologic features and various environmental risk factors. METHODS: We detected the aberrant methylation presence of these genes by methylation-specific polymerase chain reaction and analyzed the potential correlations with multivariate logistic regression model as well as stepwise logistic regression. RESULTS: For CDH1, promoter methylation was less frequent in adenosquamous carcinomas than adenocarcinomas (OR=0.35, 95%CI=0.13-0.96); pickled food increased the methylation frequency (OR=2.23, 95%CI=1.09-4.54) while light smoking and fruit intake decreased that (OR=0.43, 95%CI=0.19-0.97; OR=0.37, 95%CI=0.15-0.95). For TSLC1, males and toxin exposure increased methylation frequency (OR=6.25, 95%CI=1.05-37.13; OR=2.42, 95%CI=1.01-5.77) while light smoking and radiation exposure decreased that (OR=0.14, 95%CI=0.03-0.60; OR=0.17, 95%CI=0.04-0.87). For TIMP3, males showed lower methylation frequency than females (OR=0.18, 95%CI=0.04-0.88) while central lung cancer, heavy smoking and radiation exposure presented higher aberrant DNA methylation status (OR=2.19, 95%CI=1.07-4.52; OR=6.99, 95%CI=1.32-37.14; OR=2.30, 95%CI=1.04-5.08). CONCLUSIONS: Aberrant promoter methylation of cell adhesion-related tumor suppressor genes in lung cancer displayed varieties of gene-specific correlations with clinicopathologic features and various environmental risk factors.

[Lung cancer and its epigenetics association with chronic obstructive pulmonary disease].

Lung cancer and chronic obstructive pulmonary disease (COPD) are the leading causes of morbidity and mortality worldwide. Development of lung cancer involves both genetic and environment factors. In addition to genetic alterations, epigenetic mechanism is closely involved in pathogenesis of lung cancer. Characterized by an abnormal persistent inflammatory response to noxious environmental stimulation, COPD has shown to increase the susceptibility for lung tumorigenesis in previous research. Current research on epigenetics of lung cancer and COPD has focused on aberrant DNA methylation, histone acetylation and non-coding RNAs regulation. The aberrant DNA methylation associated with lung cancer and COPD has included overexpression of DNA methyltransferase, global DNA hypomethylation and DNA hypermethylation in promoter regions, while histone acetylation and histone methylation are the major changes for histone modification, in which histone acetyltransferases, histone deacetylases, histone methyltransferases and histone demethylases play the most important roles. RNA interference and microRNAs are both hot topics of research on non-coding RNAs regulation. Understanding of concurrent epigenetic alterations in the pathogenesis of lung cancer and COPD may facilitate identification of specific therapeutic targets and development of effective treatment.

[Effects of 5-Aza-2-deoxycytidine on DNA methylation of anti-oncogenes in non-small cell lung cancer cells].

OBJECTIVE: To observe the expression of SFRP1 gene methylation in non-small cell lung cancer (NSCLC), and study the effect of 5-Aza-2-deoxycytidine (5-Aza-CdR) on DNA methylation and expression of SFRP1, p16 and MGMT genes in the human lung cancer cell line SPC-A-1 cells. METHODS: SP immunohistochemistry and methylation-specific PCR were used to detect the SFRP1 methylation in 60 NSCLC cases, and 21 cases of benign lung diseases were used as control group. SPC-A-1 cells were cultured and treated with 5-Aza-CdR. The promoter methylation status of SFRP1, p16 and MGMT genes were detected by methylation-specific polymerase (MSP) chain reaction, and mRNAs were detected by real-time PCR. RESULTS: The positive rate of SFRP1 gene methylation in NSCLC was significantly higher than that in normal lung tissue (58.3% vs. 14.3%; χ(2) = 12.118, P = 0.001). SFRP1 gene methylation was closely correlated with lymph node metastasis and degree of differentiation in NSCLC (P < 0.05). SFRP1 protein expression was correlated with clinical stage, degree of differentiation and lymph node metastasis in NSCLC (P < 0.05). The positive expression of SFRP1 protein in 30 cases of NSCLC tissue containing SFRP1 gene methylation was significantly higher than that in non-methylated NSCLC (68.6% vs. 24.0%; χ(2) = 9.613, P = 0.002). SFRP1 gene methylation was closely correlated with SFRP1 gene protein expression in NSCLC (P < 0.05). Negative expression of SFRP1 protein was correlated with the differentiation, clinical stage, and lymph node metastasis in NSCLC (all P < 0.05). Without 5-Aza-CdR treatment, the expressions of methylation of SFRP1, p16 and MGMT genes and their mRNA were low. After 5-Aza-CdR treatment at different concentrations, their expressions were significantly elevated (all P < 0.05). CONCLUSIONS: SFRP1 gene methylation is closely associated with carcinogenesis and development of NSCLC. 5-Aza-CdR may reverse the methylation of SFRP1, p16 and MGMT genes, and facilitate the re-expression of the anti-oncogenes.

[Correlation between RARbeta gene promoter methylation and P53 gene mutations in non-small cell lung cancer].

OBJECTIVE: To investigate the correlation between RARbeta gene promoter methylation and P53 gene mutations in non-small cell lung cancer (NSCLC). METHODS: Promoter methylation of RARbeta and P53 mutations of exons 5 through 9 in 198 resected primary NSCLC tissues were determined by methylation-specific PCR and direct sequencing. RESULTS: RARbeta gene promoter methylation and P53 mutation were detected in 58.1% and 36.4% of tumors, respectively. Both were higher in males than in females and in smokers than in nonsmokers. A higher prevalence of RARbeta promoter methylation was found in patients with advanced stage tumors than those with TNM stage I. P53 gene mutations were more frequent in squamous cell carcinoma and adeno-squamous carcinoma than adenocarcinoma. All such differences were statistically significant (P< 0.05). Frequencies of P53 mutations, including G:C>T:A mutations, transversions and missense mutations were significantly higher in tumors with RARbeta methylation than in those without (P< 0.05). A significantly higher prevalence of RARbeta methylation was found in tumors with only G:C>T:A mutation in P53 gene than those without P53 mutations (P< 0.05). This difference (OR=3.737, 95%CI: 1.414-9.873) was still statistically significant (P< 0.05) in smokers (OR=4.020, 95%CI: 1.263-12.800), squamous cell carcinomas (OR=5.480, 95%CI: 1.400-21.446) or patients with advanced tumors (OR=3.446, 95%CI: 1.054-11.267) after adjusting for age and sex. CONCLUSION: RARbeta methylation is associated with G:C>T:A mutations in P53 gene in NSCLC.

[Effects of CYP1A1 and GSTM1 gene polymorphisms and BPDE-DNA adducts on lung cancer].

OBJECTIVE: To investigate the effect of CYP1A1 and GSTM1 genetic polymorphisms and BPDE-DNA adducts on lung tumorigenesis. METHODS: The case control study has included 200 cases of lung cancer and 200 controls. DNA was extracted from blood samples of all subjects. The genotype of both CYP1A1 and GSTM1 were detected with PCR-based restriction fragment length polymorphisms (PCR-RELP). BPDE-DNA adducts were detected with competitive ELISA. RESULTS: CYP1A1 mutant genotype and GSTM1 null genotype with smoke has increased the risk of lung cancer, with OR being 2.406(1.321-4.382), 2.755(1.470-5.163), respectively. The level of BPDE-DNA adducts in patients was greater than control, and the adduct level in ever smokers was higher than never smokers, the difference was statistically significant (P= 0.0252). GSTM1 null genotype individuals with BPDE-DNA level higher than 5 adducts/10(8) nucleotide have increased risk of lung cancer (OR= 1.988, 95%CI: 1.011-3.912). Compared with never smokers with CYP1A1 wild genotype, smokers with CYP1A1 mutation genotype had an increased risk of forming a higher level of DNA adducts (P= 0.0459). Smokers with GSTM1 null genotype formed more DNA adducts compared with never smokers with GSTM1 functional genotype (OR = 2.432, 95% CI: 1.072-4.517). CONCLUSION: GSTM1 null genotype with higher level DNA adducts may increase the risk of lung cancer. DNA adducts form easier in smokers with CYP1A1 mutation genotype and GSTM1 null genotype, which in turn may influence lung tumorigenesis.

Effect of CYP1A1 MSPI polymorphism on the relationship between TP53 mutation and CDKN2A hypermethylation in non-small cell lung cancer.

BACKGROUND AND AIMS: The molecular mechanisms of lung cancer susceptibility have not been fully understood. Although it has been described that germline polymorphisms are associated with either mutation or methylation of genes, the link between gene polymorphisms and gene-gene interactions has not been investigated. Therefore, we conducted this study to determine whether CYP1A1/GSTM1 polymorphisms can affect the relationship between TP53 mutation and CDKN2A hypermethylation in lung cancer. METHODS: This study included 196 primary non-small cell lung cancer (NSCLC) patients. CYP1A1 MSPI and GSTM1 polymorphisms were characterized through PCR-RFLP on DNA isolated from peripheral lymphocytes. TP53 mutations of exons 5 through 9 and CDKN2A promoter hypermethylation in both cancer tissues and corresponding normal tissues were analyzed by direct sequencing and methylation-specific PCR (MSP) respectively. RESULTS: TP53 mutation in the tumor was associated with squamous cell histology and CDKN2A methylation was associated with older age (≥60 years), heavy smoking (>30 pack-years), squamous cell histology and advanced stage (stage II-IV). After adjusting for age, sex, smoking degree, histology type and TNM stage, the correlation between TP53 mutation and CDKN2A methylation was significant in patients with CYP1A1 risk genotype (p = 0.038), but not in those with CYP1A1 homogeneity wild genotype (p = 0.151). CONCLUSIONS: This may suggest that TP53 mutation and CDKN2A methylation specifically interact to promote lung tumorigenesis in subjects with CYP1A1 risk genotype but not in those with CYP1A1 wild-type homozygotes, implying different pathways for the development of lung carcinoma with respect to CYP1A1 polymorphism.

[Current progress in research on epigenetic effects of air pollution].

Air pollution is associated with numerous diseases. In recent years,researches have increasingly showed that epigenetic modifications usually occur at the early stage of diseases, and make greater contributions to the occurrence and development of diseases compared to genetic abnormalities. Thus, researches on epigenetic effects of air pollution would serve for better understanding the interaction between air pollutants and genome in the pathogenesis of disease. Meanwhile, in order to reduce the exposure to air pollution and diminish the adverse effects related, further studies are needed to identify epigenetic biomarkers of air pollution so that we can take timely and effective measures in disease prevention.

[Relationship between promoter methylation of p16, DAPK and RAR beta genes and the clinical data of non-small cell lung cancer].

OBJECTIVE: To investigate the effects of promoter methylation of p16, death-associated protein kinase (DAPK) and retinoic acid receptor-beta (RAR beta) genes on clinical data in non-small cell lung cancers, and to study the effect of smoking on the risk of gene methylation. METHODS: The promoter methylation of p16, DAPK and RAR beta genes in 200 primary non-small cell lung cancers and the corresponding nonmalignant lung tissues were determined by methylation-specific PCR. RESULTS: Methylation in the tumor tissues was detected in 51.0% for p16, 60.0% for DAPK, and 58.0% for RAR beta gene, with significant differences (P < 0.05) when compared with those in the corresponding nonmalignant tissues(12.5%, 11.5% and 15.0%) respectively. p16 gene methylation in tumor tissue was associated with age significantly in unconditional logistic regression analysis (P < 0.01) and histologic type (P < 0.05). DAPK gene methylation in tumor tissue was associated significantly with age (P < 0.05), gender (P < 0.05) and clinical type (P < 0.05). RAR beta gene methylation in tumor tissue was associated with clinical type (P < 0.05) and tumor stage (P < 0.05) significantly. The interaction odds ratio (OR) for the gene-gene interaction in tumor tissue between p16 and DAPK was 1.987 (95%CI:1.055-3.743). The results of the gene-smoking analyses revealed that a relationship existed between cigarette smoking and p16 gene methylation (OR = 3.139, 95%CI: 1.046-9.419), the OR for the relationship of DAPK gene methylation and cigarette smoking was 3.585(95%CI: 1.270-10.123) in tumor tissue. The RAR beta gene methylation did not differ based on the smoking status of patients in tumor tissue. CONCLUSION: The p16, DAPK and RAR beta genes methylation are strongly associated with clinical data of non-small cell lung cancer, and methylation of p16 and DAPK genes are associated with tobacco smoking.

[Study on genotoxicity of aldicarb and methomyl].

Genotoxicity of aldicarb and methomyl was explored. The aldicarb and methomyl were diluted by the deionized water respectively, and then five concentrations of aldicarb were generated as 0.002, 0.02, 0.2, 2, 20 microg/L, methomyl as 0.02, 0.2, 2, 20, 200 microg/L. The micronucleus of carp erythrocyte was counted by micronucleus test. The mutation of bacteria was assessed by Ames test. The DNA damage of human lymphocytes was tested by comet assay. The genotoxicity of aldicarb and methomyl was estimated by the three toxicology tests mentioned above. The results showed that, in the micronucleus test, both any concentration of two pesticides were not able to induce higher frequency of micronucleus in carp erythrocyte (p > 0.05). Under condition of metabolic inactivation, although the number of colony with back mutation in any concentration of two pesticides did not exceed the double number of those with spontaneous mutation, the revertants of TA97 strains in the aldicarb 2-20 microg/L and the methomyl 20-200 microg/L were (129.17 +/- 17.00), (129.50 +/- 18.28), (109.83 +/- 10.80) and (114.17 +/- 9.37) entries/plate, respectively, they were significantly greater than those in spontaneous mutation (p < 0.05, p < 0.01). In the methomyl 200 microg/L group, the revertants of TA100 and TA102 strains were (147.83 +/- 23.29) and (275.83 +/- 20.63) entries/plate, respectively, they are significantly higher than that of the control group under condition of metabolic activation (p < 0.05). In comet assay, both the high concentration groups of aldicarb and methomyl resulted in different degrees of DNA damage of human peripheral blood lymphocytes. Compared with deionized water group, all of three indexes of comet assay in the aldicarb 20 microg/L groups and the methomyl 200 microg/L groups were significantly higher (p < 0.01). Despite that both aldicarb and methomyl did not results in damaging chromosome carp erythrocyte and producing apparent mutagenicity, the effect of mutagenicity and DNA damage in human lymphocytes were observed in high concentration groups of both aldicarb and methomyl. Water polluted by aldicarb and methomyl may have the potential adverse effects on the environment and human health.

[Influence of alcohol consumption on effectiveness of antihypertensive in male patients with essential hypertension].

OBJECTIVE: To explore the influence of alcohol consumption during ACEI (Benazepril) therapy on effectiveness of antihypertensive in male patients with essential hypertension. METHODS: A prospective cohort study was made and multiple linear regression and multiple logistic regression models were applied to data analysis. RESULTS: After 15 days ACEI therapy, the decrease in systolic blood pressure (deltaSBP) and diastolic blood pressure (deltaDBP) in alcohol-drinking groups was clearly smaller than that of non-drinking group. An inverse dose-response relation between alcohol drinking and (deltaSBP) as well as (deltaDBP) was observed either. To the non-alcohol-drinking group, subjects with alcohol-drinking more than 50 ml per day showed 5.26 mmHg (0.70 kPa) (P = 0.0116) and 3.32 mmHg (0.44kPa) (P = 0.0349) decreased in NSBP and DDBP, respectively. Logistic regression analysis demonstrated that the alcohol-drinking group's effect rate of antihypertensive were 45% lower (P = 0.0493) in SBP drop and 76% lower (P = 0.4750) in DBP drop respectively compared with non-and alcohol-drinking groups. CONCLUSION: Alcohol drinking during ACEI therapy can lower the effectiveness of antihypertensive in male patients with essential hypertension.