Sialyl Lewis x expression in cervical scrapes of premalignant lesions

Sialylated oligosaccharides of glycoproteins and glycolipids have been implicated in tumour progression and metastases. Altered expression of glycosidic antigens has been reported in cervical cancer. In cervix premalignant lesions, an increased expression of sialic acid has been reported. In the present study we determined the expression profiles of the glycosidic antigens Tn, sialyl Tn (sTn), Lewis a (Le^a), sialyl Lewis a (sLe^a), Lewis x (Le^x) and sialyl Lewis x (sLe^x) in cervical scrapes with cytological diagnoses of normal, low-grade squamous intraepithelial lesions (LGSIL) and high-grade squamous intraepithelial lesions (HGSIL). Cervical scrapings were collected to detect tumour antigens expressions by flow cytometry using monoclonal antibodies. Cytometry analysis of Tn, sTn, Le^a and Le^x did not reveal differences at the expression level among groups. The number of positive cells to sLe^a antigen increased in the HGSIL group with respect to the normal group (p?=?0.0495). The number of positive cells to sLe^x antigen in the samples increased with respect to the grade of squamous intraepithelial lesion (SIL) (p?<?0.001, Mann–Whitney U test). The intensity of expression of this antigen increased in the HGSIL samples with respect to normal samples (p?<?0.0068). sLe^x antigen could be a candidate to be used as biomarker for the early diagnosis of cervical cancer.     

Diet is not responsible for the presence of several oxidatively damaged DNA lesions in mouse urine

In order to eliminate the possibility that diet may influence urinary oxidative DNA lesion levels, in our experiments we used a recently developed technique involving HPLC pre-purification followed by gas chromatography with isotope dilution mass spectrometric detection. This methodology was applied for the determination of the lesions: 8-oxo-7,8-dihydroguanine (8-oxoGua), 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodGuo) and 5-(hydroxymethyl)uracil (5HMUra) in the urine of mice fed with nucleic acid free diet and normal, unrestricted diet. The mean levels of 8-oxoGua, 8-oxodGuo and 5HMUra of the animals fed the normal diet reached the mean values of [Formula: See Text], [Formula: See Text] and [Formula: See Text] After feeding the mice for 12 days with nucleic acid free diet the respective values were [Formula: See Text], [Formula: See Text] and [Formula: See Text] respectively. The results clearly demonstrate that irrespective of the diet, the excretion rates were not statistically different during the course of feeding. The respective p values for the differences between lesions in the two types of diets were: 0.13 (8-oxoGua), 0.16 (8-oxodGuo), 0.18 (5-HMUra). Our results clearly indicate that diet does not contribute to urinary excretion of the lesions in mouse model.     

A premature stop codon in the ADAMTS2 gene is likely to be responsible for dermatosparaxis in Dorper sheep

We have used polymerase chain reaction-single-strand conformational polymorphism analysis to investigate variation in exon 2 of the ADAM metalloproteinase with thrombospondin type I motif, 2 (ADAMTS2) gene in 598 sheep, including three white Dorper lambs that had a pathology consistent with dermatosparaxis. Four sequence variants (A, B, C and D) were identified at this exon, with the lambs having the dermatosparaxis phenotype being uniquely B homozygous and their mothers being B-containing heterozygous for ADAMTS2. Analysis of the amplified exon 2 sequences revealed the B variant had a nucleotide substitution that creates a premature stop codon and would notionally abbreviate the ADAMTS2 peptide. The B variant was not found in any other breed aside from the white Dorper sheep that were studied.     

hTERT methylation and expression in gastric cancer

Gastric cancer is the second most prevalent cause of cancer death worldwide. DNA methylation is a common event in gastric carcinogenesis. hTERT seems to be the rate-limiting determinant of telomerase activation, which is responsible for stability and life span. hTERT hypermethylation has been associated with telomerase expression. In the present study, we investigated the promoter methylation status and hTERT protein expression in gastric cancer and normal mucosa samples. One hundred and nine gastric cancer and 53 normal mucosa samples were investigated through methylation-specific PCR. Immunohistochemistry was analysed using peroxidase in 55 gastric cancer and 18 normal gastric mucosa samples. This is the first study evaluating hTERT methylation status in gastric carcinogenesis. We did not observe hTERT protein expression in normal gastric mucosa. Moreover, hTERT expression was observed in 80% of tumours and was associated with gastric cancer (p?<?0.0001). Partial methylation was the most frequent pattern in gastric samples, even in normal mucosa. The frequency of specimens presenting hypermethylation was significantly higher in tumours than in normal mucosa samples (p?=?0.0002), although the presence of hypermethylated promoter was not associated with a higher frequency of hTERT expression. A low correlation between hTERT protein expression and methylation was verified in gastric cancer samples. There was a clear difference in the frequency of hTERT expression and methylation within tumoral and non-tumoral tissues. Methylation status and telomerase expression may be useful for the diagnosis of gastric cancer and may have an impact on the anti-telomerase strategy for cancer therapy.     

DNA repair is responsible for the presence of oxidatively damaged DNA lesions in urine

The repair of oxidatively damaged DNA is integral to the maintenance of genomic stability, and hence prevention of a wide variety of pathological conditions, such as aging, cancer and cardiovascular disease. The ability to non-invasively assess DNA repair may provide information regarding repair pathways, variability in repair capacity, and susceptibility to disease. The development of assays to measure urinary DNA lesions offered this potential, although it rapidly became clear that possible contribution from diet and cell turnover may influence urinary lesion levels. Whilst early studies attempted to address these issues, up until now, much of the data appears conflicting. However, recent work from our laboratories, in which human volunteers were fed highly oxidatively modified 15N-labelled DNA demonstrates that diet does not appear to contribute to urinary levels of 8-hydroxyguanine and 7,8-dihydro-8-oxo-2'-deoxyguanosine. Furthermore, we propose that a number of literature reports form an argument against a contribution from cell death. Indeed we, and others, have presented evidence, which strongly suggests the involvement of cell death to be minimal. Taken together, these data would appear to rule out various confounding factors, leaving DNA repair pathways as the principal source of urinary purine, if not DNA, lesions enabling such measurements to be used as indicators of repair.     

DNA damage from micronutrient deficiencies is likely to be a major cause of cancer

A deficiency of any of the micronutrients: folic acid, Vitamin B12, Vitamin B6, niacin, Vitamin C, Vitamin E, iron, or zinc, mimics radiation in damaging DNA by causing single- and double-strand breaks, oxidative lesions, or both. For example, the percentage of the US population that has a low intake (<50% of the RDA) for each of these eight micronutrients ranges from 2 to >20%. A level of folate deficiency causing chromosome breaks was present in approximately 10% of the US population, and in a much higher percentage of the poor. Folate deficiency causes extensive incorporation of uracil into human DNA (4 million/cell), leading to chromosomal breaks. This mechanism is the likely cause of the increased colon cancer risk associated with low folate intake. Some evidence, and mechanistic considerations, suggest that Vitamin B12 (14% US elderly) and B6 (10% of US) deficiencies also cause high uracil and chromosome breaks. Micronutrient deficiency may explain, in good part, why the quarter of the population that eats the fewest fruits and vegetables (five portions a day is advised) has about double the cancer rate for most types of cancer when compared to the quarter with the highest intake. For example, 80% of American children and adolescents and 68% of adults do not eat five portions a day. Common micronutrient deficiencies are likely to damage DNA by the same mechanism as radiation and many chemicals, appear to be orders of magnitude more important, and should be compared for perspective. Remedying micronutrient deficiencies should lead to a major improvement in health and an increase in longevity at low cost.     

DNA methylation is not increased in mouse-human somatic cell hybrids

The level of DNA methylation in three mouse-human cell lines that retained different human chromosomes and in the parental mouse and human lines has been determined by high-pressure liquid chromatography (HPLC). The level of methylation is similar in the hybrid and parental cells, indicating that interspecific somatic cell hybridization followed by preferential chromosome segregation can occur without an increase in overall DNA methylation.     

Identification of boron transporter genes likely to be responsible for tolerance to boron toxicity in wheat and barley

Tolerance to boron (B) toxicity in cereals is known to be associated with reduced tissue accumulation of B. Genes from roots of B-tolerant cultivars of wheat and barley with high similarities to previously reported B efflux transporters from Arabidopsis and rice were cloned. Expression of these genes was strongly correlated with the ability of tolerant genotypes to lower the concentration of B in roots. The gene from barley located to chromosome 4. Backcross lines containing a B tolerance locus on chromosome 4 showed tolerance in proportion to the level of expression of the transporter gene, whereas those lacking the locus were sensitive to B and had very low levels of gene expression. The results are consistent with a widespread mechanism of tolerance to high B based on efflux of B from root cells.     

Developmental patterns of chromatin structure and DNA methylation responsible for epigenetic expression of a maize regulatory gene

Epigenetic regulatory mechanisms heritably alter patterns of gene expression without changes in DNA sequence. Epigenetic states are often correlated with developmentally imposed alterations in genomic DNA methylation and local chromatin structure. Pl-Blotched is a stable epigenetic allele of the maize anthocyanin regulatory gene, purple plant1(pl). Pl-Blotched plants display a variegated pattern of pigmentation that contrasts sharply with the uniformly dark purple pigmentation of plants carrying the dominant Pl-Rhoades allele. Previously, we showed that the lower level of pigmentation in Pl-Blotched is correlated with lower pl mRNA levels and increased DNA methylation at some sites. To explore how DNA methylation, chromatin structure, and developmental stage might contribute to the expression of Pl-Blotched, we used methylation-sensitive restriction enzymes and DNaseI sensitivity assays to compare the methylation status and chromatin structure of Pl-Blotched and Pl-Rhoades at different stages in development. Both alleles exhibit developmentally sensitive changes in methylation. In Pl-Blotched, methylation of two diagnostic HpaII/MspI sites increases progressively, coincident with the juvenile-to-adult transition in growth. In seedlings, the chromatin encompassing the coding region of the gene is less sensitive to DNaseI digestion in Pl-Blotched than in Pl-Rhoades. Developmental maturation from seedling to adult is accompanied by expansion of this closed chromatin domain to include the promoter and downstream flanking sequences. We provide evidence to show that chromatin structure, rather than DNA methylation, is the primary epigenetic determinant for the phenotypic differences between Pl-Blotched and Pl-Rhoades.     

Alternative splicing is a developmental switch for hTERT expression

1. Download : Download high-res image (116KB)
2. Download : Download full-size image

     

Characteristics of high grade dyskaryotic cervical smears likely to be missed on rapid rescreening

OBJECTIVE: To determine if there is a type of high grade dyskaryotic cervical smear that is likely to be missed on rapid rescreening. STUDY DESIGN: Fifty high grade dyskaryotic smears that had originally been incorrectly reported as negative (FN) were admixed with 100 true negative smears. Each smear in the set was rapidly reviewed at least 40 times. The FN smears that were picked out on > 50% of screenings were compared with those that were passed as unremarkable on > 50% of screenings for features of the dyskaryotic cell population. RESULTS: Significant differences between the two types of FN smear were present in five aspects of the dyskaryotic cell population. A FN smear is more likely to be missed on rapid rescreening than to be selected for review if it has few dyskaryotic cells; if the dyskaryotic cells are small, with pale nuclei; and if they are scattered singly rather than in groups or syncytia. CONCLUSION: A type of severely dyskaryotic smear is likely to evade rapid rescreening as well as routine screening. This suggests that even when rapid rescreening is used as a quality assurance measure, the "zero-error standard" is unlikely to be attained.