Occupational exposure to aflatoxin (AFB1) in poultry production.

Aflatoxin B1 (AFB1) has been recognized to produce cancer in human liver. In addition, epidemiological and laboratory studies demonstrated that the respiratory system was a target for AFB1. Exposure occurs predominantly through the food chain, but inhalation represents an additional route of exposure. The present study aimed to examine AFB1 exposure among poultry workers in Portugal. Blood samples were collected from a total of 31 poultry workers from six poultry farms. In addition, a control group (n = 30) was included comprised of workers who undertook administrative tasks. Measurement of AFB1 in serum was performed by enzyme-linked immunosorbent assay (ELISA). For examining fungi contamination, air samples were collected through an impaction method. Air sampling was obtained in pavilion interior and outside the premises, since this was the place regarded as the reference location. Using molecular methods, toxicogenic strains (aflatoxin-producing) were investigated within the group of species belonging to Aspergillus flavus complex. Eighteen poultry workers (59%) had detectable levels of AFB1 with values ranging from <1 ng/ml to 4.23 ng/ml and with a mean value of 2 ± 0.98 ng/ml. AFB1 was not detected in the serum sampled from any of the controls. Aspergillus flavus was the fungal species third most frequently found in the indoor air samples analyzed (7.2%) and was the most frequently isolated species in air samples containing only Aspergillus genus (74.5%). The presence of aflatoxigenic strains was only confirmed in outdoor air samples from one of the units, indicating the presence of a source inside the building in at least one case. Data indicate that AFB1 inhalation represents an additional risk in this occupational setting that needs to be recognized, assessed, and prevented.

Gene expression regulation and lineage evolution: the North and South tale of the hybrid polyploid Squalius alburnoides complex.

The evolution of hybrid polyploid vertebrates, their viability and their perpetuation over evolutionary time have always been questions of great interest. However, little is known about the impact of hybridization and polyploidization on the regulatory networks that guarantee the appropriate quantitative and qualitative gene expression programme. The Squalius alburnoides complex of hybrid fish is an attractive system to address these questions, as it includes a wide variety of diploid and polyploid forms, and intricate systems of genetic exchange. Through the study of genome-specific allele expression of seven housekeeping and tissue-specific genes, we found that a gene copy silencing mechanism of dosage compensation exists throughout the distribution range of the complex. Here we show that the allele-specific patterns of silencing vary within the complex, according to the geographical origin and the type of genome involved in the hybridization process. In southern populations, triploids of S. alburnoides show an overall tendency for silencing the allele from the minority genome, while northern population polyploids exhibit preferential biallelic gene expression patterns, irrespective of genomic composition. The present findings further suggest that gene copy silencing and variable expression of specific allele combinations may be important processes in vertebrate polyploid evolution.

eRF3a/GSPT1 12-GGC allele increases the susceptibility for breast cancer development.

It is now widely recognized that translation factors are involved in cancer development and that components of the translation machinery that are deregulated in cancer cells may become targets for cancer therapy. The eukaryotic Release Factor 3 (eRF3) is a GTPase that associates with eRF1 in a complex that mediates translation termination. eRF3a/GSPT1 first exon contains a (GGC)n expansion coding for proteins with different N-terminal extremities. Herein we show that the longer allele (12-GGC) is present in 5.1% (7/137) of the breast cancer patients analysed and is absent in the control population (0/135), corresponding to an increased risk for cancer development, as revealed by Odds Ratio analysis. mRNA quantification suggests that patients with the 12-GGC allele overexpress eRF3a/GSPT1 in tumor tissues relative to the normal adjacent tissues. However, using an in vivo assay for translation termination in HEK293 cells, we do not detect any difference in the activity of the eRF3a proteins encoded by the various eRF3a/GSPT1 alleles. Although the connection between the presence of eRF3a/GSPT1 12-GGC allele and tumorigenesis is still unknown, our data suggest that the presence of the 12-GGC allele provides a potential novel risk marker for various types of cancer.

DMBT1 is frequently downregulated in well-differentiated gastric carcinoma but more frequently upregulated across various gastric cancer types.

Well-differentiated gastric carcinomas are considered to represent a distinct entity emerging via specific molecular changes different from those found in other gastric carcinoma types. The gene deleted in malignant brain tumours 1 (DMBT1) at 10q25.3-q26.1 codes for a protein presumably involved in cell differentiation and protection and has been proposed as a candidate tumour suppressor for brain and epithelial cancer. One study reported a loss of DMBT1 expression in 12.5% (5/40) of gastric cancer samples. Here, we examined in more detail DMBT1 protein and mRNA expression in 78 primary gastric tumour samples and corresponding normal gastric mucosa. DMBT1 was expressed in all non-tumour gastric mucosa tissues. Eleven out of 71 (15%) gastric tumours were negative for the DMBT1 protein in immunohistochemical analyses. Lack of DMBT1 expression was significantly more frequently found in well-differentiated gastric tumours (6/18 well-differentiated tumours vs. 5/53 other subtypes; P=0.025). Quantitative RT-PCR revealed a downregulation of the DMBT1 mRNA for 8/21 (38%) cases, while the remaining 13 cases (62%) displayed a substantial upregulation. Our data suggest that a loss of DMBT1 expression may preferentially take place in well-differentiated gastric carcinoma. However, an upregulation of DMBT1 expression is more frequently found across all gastric cancer types.

Genotoxic effects in occupational exposure to formaldehyde: A study in anatomy and pathology laboratories and formaldehyde-resins production.

BACKGROUND: According to the Report on Carcinogens, formaldehyde ranks 25th in the overall U.S. chemical production, with more than 5 million tons produced each year. Given its economic importance and widespread use, many people are exposed to formaldehyde environmentally and/or occupationally. Presently, the International Agency for Research on Cancer classifies formaldehyde as carcinogenic to humans (Group 1), based on sufficient evidence in humans and in experimental animals. Manyfold in vitro studies clearly indicated that formaldehyde can induce genotoxic effects in proliferating cultured mammalian cells. Furthermore, some in vivo studies have found changes in epithelial cells and in peripheral blood lymphocytes related to formaldehyde exposure. METHODS: A study was carried out in Portugal, using 80 workers occupationally exposed to formaldehyde vapours: 30 workers from formaldehyde and formaldehyde-based resins production factory and 50 from 10 pathology and anatomy laboratories. A control group of 85 non-exposed subjects was considered. Exposure assessment was performed by applying simultaneously two techniques of air monitoring: NIOSH Method 2541 and Photo Ionization Detection equipment with simultaneously video recording. Evaluation of genotoxic effects was performed by application of micronucleus test in exfoliated epithelial cells from buccal mucosa and peripheral blood lymphocytes. RESULTS: Time-weighted average concentrations not exceeded the reference value (0.75 ppm) in the two occupational settings studied. Ceiling concentrations, on the other hand, were higher than reference value (0.3 ppm) in both. The frequency of micronucleus in peripheral blood lymphocytes and in epithelial cells was significantly higher in both exposed groups than in the control group (p<0.001). Moreover, the frequency of micronucleus in peripheral blood lymphocytes was significantly higher in the laboratories group than in the factory workers (p < 0.05). A moderate positive correlation was found between duration of occupational exposure to formaldehyde (years of exposure) and micronucleus frequency in peripheral blood lymphocytes (r=0.401; p<0.001) and in epithelial cells (r=0.209; p < 0.01). CONCLUSIONS: The population studied is exposed to high peak concentrations of formaldehyde with a long-term exposure. These two aspects, cumulatively, can be the cause of the observed genotoxic endpoint effects. The association of these cytogenetic effects with formaldehyde exposure gives important information to risk assessment process and may also be used to assess health risks for exposed workers.

Differential expression of GSPT1 GGCn alleles in cancer.

The human eukaryotic release factor 3a (eRF3a), encoded by the G1 to S phase transition 1 gene (GSPT1; alias eRF3a), is upregulated in various human cancers. GSPT1 contains a GGC(n) polymorphism in exon 1, encoding a polyglycine expansion in the N-terminal of the protein. The longer allele, GGC(12), was previously shown to be associated to cancer. The GGC(12) allele was present in 2.2% of colorectal cancer patients but was absent in Crohn disease patients and in the control group. Real-time quantitative RT-PCR analysis showed that the GGC(12) allele was present at up to 10-fold higher transcription levels than the GGC(10) allele (P < 0.001). No GSPT1 amplifications were detected, and there was no correlation between the length of the alleles and methylation levels of the CpG sites inside the GGC expansion. Using flow cytometry, we compared the levels of apoptosis and proliferation rates between cell lines with different genotypes, but detected no significant differences. Finally, we used a cytokinesis-block micronucleus assay to evaluate the frequency of micronuclei in the same cell lines. Cell lines with the longer alleles had higher frequencies of micronuclei in binucleated cells, which is probably a result of defects in mitotic spindle formation. Altogether, these findings indicate that GSPT1 should be considered a potential proto-oncogene.

Translation termination and protein folding pathway genes are not correlated in gastric cancer.

BACKGROUND: The eukaryotic release factor 3 (eRF3) has been shown to affect both tubulin and actin cytoskeleton, suggesting a role in cytoskeleton assembly, mitotic spindle formation and chromosome segregation. Also, direct interactions between eRF3 and subunits of the cytosolic chaperonin CCT have been described. Moreover, both eRF3a and CCT subunits have been described to be up-regulated in cancer tissues. Our aim was to evaluate the hypothesis that eRF3 expression levels are correlated with the expression of genes encoding proteins involved in the tubulin folding pathways. METHODS: Relative expression levels of eRF1, eRF3a/GSPT1, PFDN4, CCT2, CCT4, and TBCA genes in tumour samples relative to their adjacent normal tissues were investigated using real time-polymerase chain reaction in 20 gastric cancer patients. RESULTS: The expression levels of eRF3a/GSPT1 were not correlated with the expression levels of the other genes studied. However, significant correlations were detected between the other genes, both within intestinal and diffuse type tumours. CONCLUSIONS: eRF3a/GSPT1 expression at the mRNA level is independent from both cell translation rates and from the expression of the genes involved in tubulin-folding pathways. The differences in the patterns of expression of the genes studied support the hypothesis of genetically independent pathways in the origin of intestinal and diffuse type gastric tumours.

Modulation of translation factor's gene expression by histone deacetylase inhibitors in breast cancer cells.

The histone deacetylase inhibitors sodium butyrate (NaBu) and trichostatin A (TSA) exhibit anti-proliferative activity by causing cell cycle arrest and apoptosis. The mechanisms by which NaBu and TSA cause apoptosis and cell cycle arrest are not yet completely clarified, although these agents are known to modulate the expression of several genes including cell-cycle- and apoptosis-related genes. The enzymes involved in the process of translation have important roles in controlling cell growth and apoptosis, and several of these translation factors have been described as having a causal role in the development of cancer. The expression patterns of the translation mechanism, namely of the elongation factors eEF1A1 and eEF1A2, and of the termination factors eRF1 and eRF3, were studied in the breast cancer cell line MCF-7 by real-time quantitative reverse transcription-polymerase chain reaction after a 24-h treatment with NaBu and TSA. NaBu induced inhibition of translation factors' transcription, whereas TSA caused an increase in mRNA levels. Thus, these two agents may modulate the expression of translation factors through different pathways. We propose that the inhibition caused by NaBu may, in part, be responsible for the cell cycle arrest and apoptosis induced by this agent in MCF-7 cells.

Genetic structure of an endangered Portuguese semiferal pony breed, the Garrano.

The present study intends to survey the genetic variability of an endangered semiferal Portuguese native pony breed, the Garrano. Thirteen microsatellite markers were examined in 277 animals born in 1998, belonging to eight subpopulations corresponding to eight northern Portuguese geographic regions. Mean heterozygosity (H(o)) in the Garrano breed was 0.732, ranging from 0.531 to 0.857 across subpopulations. Allelic frequencies and diversity differed significantly between regions, suggesting the existence of genetic differentiation within the breed confirmed by the population differentiation estimator F(ST). Allele sharing genetic distance (Dps) was used to determine the relationship between the analyzed subpopulations, some of which are diverging significantly from the others. Relationships among six Iberian horse breeds (including the Garrano) were assessed through the restricted maximum likelihood method, which clusters the Garrano with another Celtic pony, the Spanish Asturcon, traditionally bred in geographical continuity with the Garrano. Results reveal a statistically significant deficit of heterozygotes within the Garrano breed (F(IT) = 0.031, p < 0.05). Regular monitoring should therefore be implemented so that the effect of genetic drift within subpopulations, enhanced by inbreeding, may be successfully minimized.