Effects of DNA repair gene polymorphisms on DNA damage in human lymphocytes induced by a vinyl chloride metabolite in vitro.

BACKGROUND: Epidemiologic studies suggest that variability in DNA damage from vinyl chloride monomer (VCM) may be partially mediated by genetic polymorphisms in DNA repair. This study aimed to corroborate these observations with controlled experiments in vitro using cell lines from individuals with differing DNA repair genotypes to determine damage following VCM metabolite exposure. METHODS: Matched pairs of lymphoblast cell lines (homozygous wild-type versus homozygous variant for either XRCC1 399 or XPD 751 polymorphism) were exposed to chloroacetaldehyde and analyzed by the cytokinesis-block micronucleus assay. RESULTS: All cell lines demonstrated a dose-response of increasing micronuclei with increasing exposure, but for both XRCC1 and XPD, the polymorphic cells peaked at higher micronucleus frequencies and declined at a slower rate to baseline than the wild-type cells. CONCLUSION: This supports the findings that XRCC1 and XPD polymorphisms may result in deficient DNA repair of VCM-induced genetic damage.

Polymorphisms in BER and NER pathway genes: effects on micronucleus frequencies among vinyl chloride-exposed workers in Northern China.

In this study, a group of 317 workers occupationally exposed to vinyl chloride monomer and 166 normal, unexposed referents in Shandong province (Northern China) were examined for chromosomal damage in peripheral lymphocytes using the cytokinesis-blocked micronucleus (CB-MN) assay. The exposure group (3.47±2.65)‰ showed higher micronucleus frequency than the unexposed workers (2.51±1.96)‰ (P<0.01). We explored the relationship between genetic polymorphisms of XRCC1 (-77C/T, Arg194Trp, Arg280His, Arg399Gln), APE1 Asp148Glu, XPA Ala23Gly, XPC.PAT, XPC Ala499Val, XPC Lys939Gln, XPF 5'-UTR T2063A, XPG Exon15 G-C, ERCC13'-UTR C8092A and susceptibility of chromosomal damage in all the subjects. It was found that XRCC1 -77, XRCC1 280, APE1148, XPC.PAT, XPG Exon15 G-C, and ERCC13'-UTR C8092A polymorphisms showed no significant associations with micronucleus frequency in unexposed workers. However, among the exposed workers individuals with XRCC1 (-77C/T, Arg194Trp, Arg280His, Arg399Gln) polymorphisms had a significantly higher micronucleus frequency as seen in mean frequency ratios (FR) compared with their homozygous wild-type genotypes (FR=1.21, 95% CI: 1.05-1.39; P<0.01); (FR=1.14, 95% CI: 1.00-1.38; P<0.05) and (FR=1.26, 95% CI: 1.11-1.44; P<0.01); (FR=1.23, 95% CI: 1.08-1.46; P<0.01). Four SNP sites in the nucleotide excision repair (NER) pathway were associated with susceptibility for MN frequency in either unexposed or exposed workers. Further, we observed the gene-MN association changed with exposure for XRCC1 (-77C/T, Arg194Trp, Arg280His, Arg399Gln), XPA Ala23Gly, XPC Ala499Val, XPC Lys939Gln, XPF 5'-UTR T2063A. Moreover, Individuals carrying the XPC (PAT)-(499)-(939) diplotype, PAT-CG/PAT-TG, had a higher MN frequency, compared with individuals carrying the wild-type PAT-CA/PAT-CA.

C-Terminal p53 Palindromic Tetrapeptide Restores Full Apoptotic Function to Mutant p53 Cancer Cells In Vitro and In Vivo.

We previously demonstrated that a synthetic monomer peptide derived from the C-terminus of p53 (aa 361−382) induced preferential apoptosis in mutant p53 malignant cells, but not normal cells. The major problem with the peptide was its short half-life (half-life < 10 min.) due to a random coil topology found in 3D proton NMR spectroscopy studies. To induce secondary/tertiary structures to produce more stability, we developed a peptide modelled after the tetrameric structure of p53 essential for activation of target genes. Starting with the above monomer peptide (aa 361−382), we added the nuclear localization sequence of p53 (aa 353−360) and the end of the C-terminal sequence (aa 383−393), resulting in a monomer spanning aa 353−393. Four monomers were linked by glycine to maximize flexibility and in a palindromic order that mimics p53 tetramer formation with four orthogonal alpha helices, which is required for p53 transactivation of target genes. This is now known as the 4 repeat-palindromic-p53 peptide or (4R-Pal-p53p). We explored two methods for testing the activity of the palindromic tetrapeptide: (1) exogenous peptide with a truncated antennapedia carrier (Ant) and (2) a doxycycline (Dox) inducer for endogenous expression. The exogenous peptide, 4R-Pal-p53p-Ant, contained a His tag at the N-terminal and a truncated 17aa Ant at the C-terminal. Exposure of human breast cancer MB-468 cells and human skin squamous cell cancer cells (both with mutant p53, 273 Arg->His) with purified peptide at 7 µM and 15 µM produced 52% and 75%, cell death, respectively. Comparatively, the monomeric p53 C-terminal peptide-Ant (aa 361−382, termed p53p-Ant), at 15 µM and 30 µM induced 15% and 24% cell death, respectively. Compared to the p53p-Ant, the exogenous 4R-pal-p53p-Ant was over five-fold more potent for inducing apoptosis at an equimolar concentration (15 µM). Endogenous 4R-Pal-p53p expression (without Ant), induced by Dox, resulted in 43% cell death in an engineered MB468 breast cancer stable cell line, while endogenous p53 C-terminal monomeric peptide expression produced no cell death due to rapid peptide degradation. The mechanism of apoptosis from 4R-Pal-p53p involved the extrinsic and intrinsic pathways (FAS, caspase-8, Bax, PUMA) for apoptosis, as well as increasing reactive oxygen species (ROS). All three death pathways were induced from transcriptional/translational activation of pro-apoptotic genes. Additionally, mRNA of p53 target genes (Bax and Fas) increased 14-fold and 18-fold, respectively, implying that the 4R-Pal-p53p restored full apoptotic potential to mutant p53. Monomeric p53p only increased Fas expression without a transcriptional or translational increase in Fas, and other genes and human marrow stem cell studies revealed no toxicity to normal stem cells for granulocytes, erythrocytes, monocytes, and macrophages (CFU-GEMM). Additionally, the peptide specifically targeted pre-malignant and malignant cells with mutant p53 and was not toxic to normal cells with basal levels of WT p53.

Gene Therapy with p14/tBID Induces Selective and Synergistic Apoptosis in Mutant Ras and Mutant p53 Cancer Cells In Vitro and In Vivo.

Any gene therapy for cancer will be predicated upon its selectivity against cancer cells and non-toxicity to normal cells. Therefore, safeguards are needed to prevent its activation in normal cells. We designed a minimal p14ARF promoter with upstream Ap1 and E2F enhancer elements and a downstream MDR1 inhibitory element, TATA box, and a transcription initiation site (hereafter p14ARFmin). The modified p14ARFmin promoter was linked to bicistronic P14 and truncated BID (tBID) genes, which led to synergistic apoptosis via the intrinsic and extrinsic pathways of apoptosis when expressed. The promoter was designed to be preferentially activated by mutant Ras and completely inhibited by wild-type p53 so that only cells with both mutant Ras and mutant p53 would activate the construct. In comparison to most p53 gene therapies, this construct has selective advantages: (1) p53-based gene therapies with a constitutive CMV promoter cannot differentiate between normal cells and cancer cells, and can be toxic to normal cells; (2) our construct does not induce p21WAF/CIPI in contrast to other p53-based gene therapies, which can induce cell cycle arrest leading to increased chemotherapy resistance; (3) the modified construct (p14ARFmin-p14-tBID) demonstrates bidirectional control of its promoter, which is completely repressed by wild-type p53 and activated only in cells with both RAS and P53 mutations; and (4) a novel combination of genes (p14 and tBID) can synergistically induce potent intrinsic and extrinsic apoptosis in cancer cells.

Proteomic detection of cancer in asbestosis patients using SELDI-TOF discovered serum protein biomarkers.

OBJECTIVES: To identify biomarkers for cancer in asbestosis patients. METHODS: SELDI-TOF and CART were used to identify serum biomarker profiles in 35 asbestosis patients who subsequently developed cancer and 35 did not develop cancer. RESULTS: Three polypeptide peaks (5707.01, 6598.10, and 20,780.70 Da) could predict the development of cancer with 87% sensitivity and 70% specificity. The first two peaks were identified as KIF18A and KIF5A, respectively, and are part of the Kinesin Superfamily of proteins. CONCLUSIONS: We identified two Kinesin proteins that can be potentially used as blood biomarkers to identify asbestosis patients at risk of developing lung cancer.

Genetic polymorphisms of XRCC1, HOGG1 and MGMT and micronucleus occurrence in Chinese vinyl chloride-exposed workers.

In this study, a group of 313 workers occupationally exposed to vinyl chloride monomer (VCM) and 141 normal unexposed referents were examined for chromosomal damage using the cytokinesis-blocked micronucleus (CBMN) assay in peripheral lymphocytes. We explored the relationship between genetic polymorphisms of XRCC1 (Arg194Trp, Arg280His and Arg399Gln), MGMT(Leu84Phe) and hOGG1 (Ser326Cys) and susceptibility of chromosomal damage induced by VCM. Polymerase chain reaction-restriction fragment length polymorphism techniques were used to detect polymorphisms in XRCC1, hOGG1 and MGMT. It was found that the micronuclei (MN) frequency of exposed workers (4.86 +/- 2.80) per thousand was higher than that of the control group (1.22 +/- 1.24) per thousand (P < 0.01). Increased susceptibility to chromosomal damage as evidenced by higher MN frequency was found in workers with hOGG1 326 Ser/Cys genotype [frequency ratio (FR) = 1.21, 95% confidence interval (CI): 1.02-1.46; P < 0.05], XRCC1 194 Arg/Trp (FR = 1.12, 95% CI: 1.00-1.25; P < 0.05) and XRCC1 280 Arg/His and His/His genotypes (FR = 1.12, 95% CI 1.00-1.26, P < 0.05). Moreover, among susceptibility diplotypes, CGA/CAG carriers had more risk of MN frequency compared with individuals with wild-type CGG/CGG (FR = 1.67, 95% CI: 1.19-2.23; P < 0.05). MN frequency also increased significantly with age in the exposed group (FR = 1.13, 95% CI: 1.00-1.28; P < 0.05). Thus, CB-MN was a sensitive index of early damage among VCM-exposed workers. Genotype XRCC1 Arg194Trp, Arg280His, hOGG1 Ser326Cys, diplotype CGA/CAG and higher age may have an impact on the chromosome damage induced by VCM.

A prospective study of respiratory symptoms associated with chronic arsenic exposure in Bangladesh: findings from the Health Effects of Arsenic Longitudinal Study (HEALS).

BACKGROUND AND AIMS: A prospective cohort study was conducted to evaluate the effect of arsenic (As) exposure from drinking water on respiratory symptoms using data from the Health Effects of Arsenic Exposure Longitudinal Study (HEALS), a large prospective cohort study established in Ariahazar, Bangladesh in 2000-2002. A total of 7.31, 9.95 and 2.03% of the 11 746 participants completing 4 years of active follow-up reported having a chronic cough, breathing problem or blood in their sputum, respectively, as assessed by trained physicians. METHODS: Cox regression models were used to estimate HRs for respiratory symptoms during the follow-up period in relation to levels of chronic As exposure assessed at baseline, adjusting for age, gender, smoking, body mass index, education and arsenic-related skin lesion status. RESULTS: Significant positive associations were found between As exposure and respiratory symptoms. As compared with those with the lowest quintile of water As level (178 microg/l), respectively. Similarly, the corresponding HRs in relation to the second to fifth quintiles of urinary arsenic were 1.10 (95% CI 0.94 to 1.27), 1.11 (95% CI 0.95 to 1.29), 1.29 (95% CI 1.11 to 1.49) and 1.35 (95% CI 1.16 to 1.56), respectively. These associations did not differ appreciably by cigarette smoking status. CONCLUSIONS: This prospective cohort study found a dose-response relationship between As exposure and clinical symptoms of respiratory diseases in Bangladesh. In particular, these adverse respiratory effects of As were clearly evident in the low to moderate dose range, suggesting that a large proportion of the country's population may be at risk of developing serious lung diseases in the future.

Protein and amino acid intakes in a rural area of Bangladesh.

BACKGROUND: Few studies have described protein and amino acid intakes in rural Bangladesh, a country with considerable undernutrition. OBJECTIVE: The purpose of this population-based study was to assess and describe protein and amino acid intakes in Araihazar, Bangladesh. METHODS: The study participants were 11,170 adult men and women who participated in the Health Effects of Arsenic Longitudinal Study (HEALS), which had a 98% participation rate. Dietary exposures were assessed by a food-frequency questionnaire that had been designed and validated for the HEALS study population. RESULTS: The mean body mass index (BMI) was 19.7 among all participants, and 34.9% of women and 44.4% of men had a BMI below 18.5. The average caloric intake was 2142 and 2394 kcal/day among women and men, respectively, and the mean protein intake was 67.5 and 78.2 g/day. The largest sources of protein were from rice and fish. Greater protein intake was related to younger age and several socioeconomic measures, including more years of education, land and television ownership, and employment in business, farming, or as a laborer (for men) or as a homemaker (for women). CONCLUSIONS: This study found a high prevalence of underweight among study participants. Nonetheless, most participants had adequate protein intake according to Food and Agriculture Organization standards for body weight.

Dietary intake of methionine, cysteine, and protein and urinary arsenic excretion in Bangladesh.

BACKGROUND: In Bangladesh, millions of people are exposed to arsenic in drinking water; arsenic is associated with increased risk of cancer. Once ingested, arsenic is metabolized via methylation and excreted in urine. Knowledge about nutritional factors affecting individual variation in methylation is limited. OBJECTIVES: The purpose of this study was to examine associations between intakes of protein, methionine, and cysteine total urinary arsenic in a large population-based sample. METHODS: The study subjects were 10,402 disease-free residents of Araihazar, Bangladesh, who participated in the Health Effects of Arsenic Longitudinal Study (HEALS). Food intakes were assessed using a validated food frequency questionnaire developed for the study population. Nutrient composition was determined by using the U.S. Department of Agriculture National Nutrient Database for Standard Reference. Generalized estimating equations were used to examine association between total urinary arsenic across quintiles of nutrient intakes while controlling for arsenic exposure from drinking water and other predictors of urinary arsenic. RESULTS: Greater intakes of protein, methionine, and cysteine were associated with 10-15% greater total urinary arsenic excretion, after controlling for total energy intake, body weight, sex, age, tobacco use, and intake of some other nutrients. CONCLUSIONS: Given previously reported risks between lower rates of arsenic excretion and increased rates of cancer, these findings support the role of nutrition in preventing arsenic-related disease.

Effect of the XRCC1 codon 399 polymorphism on the repair of vinyl chloride metabolite-induced DNA damage.

BACKGROUND: Recent epidemiologic evidence suggests that the common polymorphism at amino acid residue 399 of the x-ray cross complementing-1 (XRCC1) protein, a key component of the base excision repair (BER) pathway for DNA damage, plays a significant role in the genetic variability of individuals in terms of the mutagenic damage they experience following exposure to the carcinogen vinyl chloride (VC). The aim of this study was to provide support for the biological plausibility of these epidemiologic observations with experimental data derived from cell lines in culture from individuals who were either homozygous wild-type or homozygous variant for this XRCC1 polymorphism following exposure to chloroethylene oxide (CEO), the active metabolite of VC, with measurement of the induced etheno-DNA adducts before and after repair. MATERIALS AND METHODS: Immortalized lymphoblast cell lines from seven VC workers (four homozygous wild-type and three homozygous variant for the 399 XRCC1 polymorphism) were exposed to CEO, and etheno-adenosine (epsilonA) adduct levels were determined by enzyme-linked immunosorbent assay (ELISA) pre-exposure and at 0, 4, 8 and 24 h following exposure. RESULTS: The average epsilonA adduct levels were statistically significantly higher in the variant cells compared to the wild-type cells at 8 and 24 h following exposure (P Conclusion: These results are consistent with the epidemiologic findings of the types of VC-induced biomarkers observed in exposed individuals and the mutational spectra found in the resultant tumors as well as the key role that BER, especially XRCC1, plays in this carcinogenic pathway.

Conformational effects of a common codon 751 polymorphism on the C-terminal domain of the xeroderma pigmentosum D protein.

AIM: The xeroderma pigmentosum D (XPD) protein is a DNA helicase involved in the repair of DNA damage, including nucleotide excision repair (NER) and transcription-coupled repair (TCR). The C-terminal domain of XPD has been implicated in interactions with other components of the TFIIH complex, and it is also the site of a common genetic polymorphism in XPD at amino acid residue 751 (Lys->Gln). Some evidence suggests that this polymorphism may alter DNA repair capacity and increase cancer risk. The aim of this study was to investigate whether these effects could be attributable to conformational changes in XPD induced by the polymorphism. MATERIALS AND METHODS: Molecular dynamics techniques were used to predict the structure of the wild-type and polymorphic forms of the C-terminal domain of XPD and differences in structure produced by the polymorphic substitution were determined. RESULTS: The results indicate that, although the general configuration of both proteins is similar, the substitution produces a significant conformational change immediately N-terminal to the site of the polymorphism. CONCLUSION: These results provide support for the hypothesis that this polymorphism in XPD could affect DNA repair capability, and hence cancer risk, by altering the structure of the C-terminal domain.

Serum growth factors in asbestosis patients.

Various growth factors, including platelet-derived growth factor (PDGF) and transforming growth factor (TGF)-beta, have been implicated in the pathogenesis of asbestos-induced disease. PDGF and TGF-beta levels were determined by enzyme-linked immunosorbent assays in the banked serum samples of a cohort of workers with asbestosis, and the relationships of the growth factor levels to the subsequent development of cancer and to the radiographic severity and progression of asbestosis in the cohort were examined. Serum levels of PDGF and TGF-beta were found to be unrelated to the development of cancer, and serum levels of PDGF were found to be unrelated to the severity and progression of asbestosis. However, serum levels of TGF-beta were found to be statistically significantly related to disease severity (p = 0.01), increasing approximately 2.4-fold from ILO radiographic category 0 to category 3, and they were marginally related to disease progression (p = 0.07), in multivariate analysis controlling for other contributory factors including cumulative asbestos exposure. This suggests that serum TGF-beta may be a useful biomarker for asbestos-induced fibrotic disease.

Gene-environment interactions between DNA repair polymorphisms and exposure to the carcinogen vinyl chloride.

We have recently suggested that polymorphisms in metabolism and repair pathways may play a role in modulating the effects of exposure to the carcinogen vinyl chloride in the production of biomarkers of its mutagenic damage. The aim of the present study was to extend these observations by examining gene-environment interactions between several common polymorphisms in the DNA repair genes XRCC1 and ERCC2/XPD and vinyl chloride exposure on the production of vinyl chloride-induced biomarkers of mutation. A cohort of 546 French vinyl chloride workers were genotyped for the XRCC1 codon 194 (Arg>Trp; rs1799782), 280 (Arg>His; rs25489) and 399 (Arg>Gln; rs25487) polymorphisms and the ERCC2/XPD codon 312 (Asp>Asn; rs1799793) and 751 (Lys>Gln; rs13181) polymorphisms. The results demonstrated a statistically significant allele dosage effect of the XRCC1 399 variant on the production of the vinyl chloride-induced mutant p53 biomarker, even after controlling for confounders including cumulative vinyl chloride exposure (p = 0.03), with a potentially supramultiplicative gene-environment interaction. In addition, the results demonstrate statistically significant allele dosage effects of the ERCC2/XPD 312 and 751 variants on the production of the vinyl chloride-induced mutant ras-p21 biomarker, even after controlling for confounders including cumulative vinyl chloride exposure (p < 0.0001 and p = 0.0006, respectively), with a potentially supramultiplicative gene-environment interaction for the codon 751 allele. Finally, the results suggest potential supramultiplicative gene-gene interactions between CYP2E1 (c2 allele; rs3813867) and ERCC2/XPD polymorphisms that are consistent with the proposed carcinogenic pathway for vinyl chloride, which requires metabolic activation by CYP2E1 to reactive intermediates that form DNA adducts that, if not removed by DNA repair mechanisms, result in oncogenic mutations.

The penetratin sequence in the anticancer PNC-28 peptide causes tumor cell necrosis rather than apoptosis of human pancreatic cancer cells.

BACKGROUND: PNC-27 and PNC-28 are p53-derived peptides from the human double minute (hdm-2) binding domain attached to penetratin. These peptides induce tumor cell necrosis of cancer cells, but not normal cells. The anticancer activity and mechanism of PNC-28 (p53 aa17-26-penetratin) was specifically studied against human pancreatic cancer. METHODS: MiaPaCa-2 cells were treated with PNC-28. Necrosis was determined by measuring lactate dehydrogenase (LDH) and apoptosis as assayed for measuring elevation of proapoptotic proteins. PNC-29, an unrelated peptide, and hdm-2-binding domain p53 aa12-26 without penetratin (PNC-26) were used as controls. Since there is evidence that penetratin is required for tumor cell necrosis, we tested "naked" p53 peptide without penetratin by transfecting a plasmid that encodes p53 aa17-26 segment of PNC-28 into MiaPaCa-2 and an untransformed rat pancreatic acinar cell line, BMRPA1. Time-lapse electron microscopy was employed to further elucidate anticancer mechanism. RESULTS: Treatment with PNC-28 does not result in the elevation of proapoptotic proteins found in p53-induced apoptosis, but elicits rapid release of LDH, indicative of tumor cell necrosis. Accordingly, we observed membrane pore formation and dose-dependent killing. In direct contrast, transfected MiaPaCa-2 cells underwent apoptosis, and not necrosis, as evidenced by expression of high levels of caspases-3 and 7 and annexin V with background levels of LDH. CONCLUSION: These results suggest that PNC-28 may be effective in treating human pancreatic cancer. The penetratin sequence appears to be responsible for the fundamental change in the mechanism of action, inducing rapid necrosis initiated by membrane pore formation. Cancer cell death by apoptosis was observed in the absence of penetratin.

Nonmalignant respiratory effects of chronic arsenic exposure from drinking water among never-smokers in Bangladesh.

BACKGROUND: Arsenic from drinking water has been associated with malignant and nonmalignant respiratory illnesses. The association with nonmalignant respiratory illnesses has not been well established because the assessments of respiratory symptoms may be influenced by recall bias or interviewer bias because participants had visible skin lesions. OBJECTIVES: We examined the relationship of the serum level of Clara cell protein CC16--a novel biomarker for respiratory illnesses--with well As, total urinary As, and urinary As methylation indices. METHODS: We conducted a cross-sectional study in nonsmoking individuals (n = 241) selected from a large cohort with a wide range of As exposure (0.1-761 microg/L) from drinking water in Bangladesh. Total urinary As, urinary As metabolites, and serum CC16 were measured in urine and serum samples collected at baseline of the parent cohort study. RESULTS: We observed an inverse association between urinary As and serum CC16 among persons with skin lesions (beta = -0.13, p = 0.01). We also observed a positive association between secondary methylation index in urinary As and CC16 levels (beta = 0.12, p = 0.05) in the overall study population; the association was stronger among people without skin lesions (beta = 0.18, p = 0.04), indicating that increased methylation capability may be protective against As-induced respiratory damage. In a subsample of study participants undergoing spirometric measures (n = 31), we observed inverse associations between urinary As and predictive FEV(1) (forced expiratory volume measured in 1 sec) (r = -0.37; FEV(1)/forced vital capacity ratio and primary methylation index (r = -0.42, p = 0.01). CONCLUSIONS: The findings suggest that serum CC16 may be a useful biomarker of epithelial lung damage in individuals with arsenical skin lesions. Also, we observed the deleterious respiratory effects of As exposure at concentrations lower than reported in earlier studies.

CYP2E1 mRNA expression, genetic polymorphisms in peripheral blood lymphocytes and liver abnormalities in Chinese VCM-exposed workers.

OBJECTIVE: To study the relationship between expression of cytochrome P4502E1 (CYP2E1) in human lymphocytes, variant CYP2E1 genotype, exposure to vinyl chloride monomer (VCM), and liver abnormalities in VCM-exposed workers. METHODS: A case-control study was performed on 90 male occupationally exposed workers and 42 matched male nonexposed controls. Data were collected based on health surveillance, workplace investigation and questionnaire Survey. Total RNA and DNA were isolated from peripheral blood lymphocytes, and CYP2E1 mRNA expression was determined using RT-PCR, and the presence of CYP2E1 polymorphisms was identified based on PCR-RFLP. RESULTS: The mRNA expression of CYP2E1 in exposed workers (0.89+/-0.46) was significantly higher than in nonexposed controls (0.61+/-0.35) (P < 0.01). Logistic regression analysis demonstrated a statistically significant association between CYP2E1 mRNA expression levels and liver abnormalities in the VCM-exposed workers (OR = 3.66, P < 0.05). The genotype frequency for CYP2E1 variants among VCM-exposed workers was not significantly different between workers with liver abnormalities and those without. CONCLUSIONS: Liver abnormalities in subjects exposed to VCM are positively associated with expression of peripheral blood lymphocyte mRNA, which is significantly increased in exposed workers compared to nonexposed controls. Therefore, CYP2E1 mRNA levels may be useful for health surveillance and protection of VCM-exposed workers.

A novel mechanism for p53 to regulate its target gene ECK in signaling apoptosis.

Transcription factor p53 regulates its target genes through binding to DNA consensus sequence and activating the promoters of its downstream genes. The conventional p53 consensus binding sequence was defined as two copies of the 10-bp motif 5'-PuPuPuC(A/T)(T/A)GPyPyPy-3' with a spacer of 0 to 13 bp, which exists in the regulatory regions of some p53 target genes. However, there is no such p53 consensus sequence in the promoters of a number of p53-responsive genes, suggesting that there might be other mechanisms whereby p53 transactivates the promoters of its target genes. We report here that p53 uses a novel binding mechanism to regulate the transcription of epithelial cell kinase (ECK), a receptor protein-tyrosine kinase implicated in signal transduction. We show that p53 binds to a 10-bp perfect palindromic decanucleotide (GTGACGTCAC) in the ECK promoter, activates the ECK promoter, and increases the transcription of ECK. This palindrome is required for p53-mediated transactivation of the ECK promoter. ECK is highly responsive to oxidative damage that leads to cell death. Ectopic expression of ECK causes spontaneous apoptosis in breast cancer cells. We found that ectopic expression of a mutant ECK fails to induce apoptosis in cancer cells. Our findings show that p53 is a transcriptional regulator of ECK in mediating apoptosis. The discovery of the novel p53-binding motif in the promoter may lead to the identification of a new class of p53 target genes.

Conformational effects of a common codon 399 polymorphism on the BRCT1 domain of the XRCC1 protein.

The X-ray cross-complementing-1 (XRCC1) protein functions as a scaffold that coordinates the activity of the cellular machinery involved in base excision repair (BER) of DNA damage. The BRCT1 domain of XRCC1 is responsible for interacting with several of the key components of the BER machinery, and it is also the site of a common genetic polymorphism in XRCC1 at amino acid residue 399 (Arg --> Gln). Experimental and epidemiologic evidence suggest that this polymorphism may alter BER capacity and increase cancer risk. The aim of this study was to investigate whether these effects could be attributable to conformational changes in XRCC1 induced by the polymorphism. Molecular dynamics techniques were used to predict the structure of the wild-type and polymorphic forms of the BRCT1 domain of XRCC1, and differences in structure produced by the polymorphic substitution were determined. The results indicate that, although the general configuration of both proteins is similar and there is little actual deviation at the site of the polymorphism itself, the substitution produces significant conformational changes at several other sites in the BRCT1 domain, including the loss of secondary structural features such as alpha helices that may be critical for protein-protein interactions. These results provide support for the hypothesis that this polymorphism in XRCC1 could affect DNA repair capability by altering the structure of the BRCT1 domain and thus the ability of XRCC1 to coordinate BER.

Non-hospital based registered nurses and the risk of bloodborne pathogen exposure.

The aim of this study was to assess the risk of blood and body fluid exposure among non-hospital based registered nurses (RNs) employed in New York State. The study population was mainly unionized public sector workers, employed in state institutions. A self-administered questionnaire was completed by a random stratified sample of members of the New York State Nurses Association and registered nurse members of the New York State Public Employees Federation. Results were reviewed by participatory action research (PAR) teams to identify opportunities for improvement. Nine percent of respondents reported at least one needlestick injury in the 12-month period prior to the study. The percutaneous injury (PI) rate was 13.8 per 100 person years. Under-reporting was common; 49% of all PIs were never formally reported and 70% never received any post-exposure care. Primary reasons for not reporting included: time constraints, fear, and lack of information on reporting. Significant correlates of needlestick injuries included tenure, patient load, hours worked, lack of compliance with standard precautions, handling needles and other sharps, poor safety climate, and inadequate training and availability of safety devices (p<0.05). PAR teams identified several risk reduction strategies, with an emphasis on safety devices. Non-hospital based RNs are at risk for bloodborne exposure at rates comparable to hospital based RNs; underreporting is an important obstacle to infection prevention, and primary and secondary risk management strategies appeared to be poorly implemented. Intervention research is warranted to evaluate improved risk reduction practices tailored to this population of RNs.