Definition of a region of loss of heterozygosity at chromosome 11q in cervical carcinoma.

Chromosomal deletions at segment 11q23-q24 have been identified in a variety of human epithelial tumors, including cervical carcinoma (CC), indicating the presence in this region of at least a tumor suppressor gene (TSG) involved in the development of these neoplasms. To localize the 11q deletion target more precisely, 54 primary cervical carcinomas were examined for loss of heterozygosity (LOH) using a panel of microsatellite DNA markers mapping to 11p.15 and spanning region 11q23-qter. Nineteen tumors were found to have LOH at chromosome 11q. The highest frequency of LOH was observed at locus APOC-3, located in 11q23.1-q23.2, which was deleted in 42% of the informative cases. In contrast, LOH was infrequent at distal 11q in current series of CC. The smallest common region of loss included APOC-3 and was defined distally by marker D11S925 in region 11q23. The present data strongly suggest that the 11q suppressor gene(s) involved in cervical tumorigenesis is likely to be located at chromosome region 11q22-q23.     

New method to measure minisatellite variant repeat variation in population genetic studies

The classical analysis of minisatellite variant repeat (MVR) variation using modular structures is limited by the lack of knowledge of the mutational process involved in the evolution of most of the minisatellites. In this study a new method to measure MVR variation and to calculate genetic distances using MVR codes is proposed. The method is based on the statistical similarity of MVR patterns and considers the complete variability of the minisatellite, enabling meaningful comparisons of closely related populations. As an example, the method has been applied to analyze variation in MSY1 (DYF155S1) in five sets of data from European and North African populations. Am. J. Hum. Biol. 14:421–428, 2002.     

Replicative senescence in patients with chronic kidney failure

BACKGROUND: Chronic activation of immunocompetent cells may lead to stress-induced premature senescence (SIPS); these senescent cells are characterized by a decrease in telomere length. The present study evaluates SIPS in circulating immunocompetent cells from predialysis patients, patients on hemodialysis, and in renal transplant patients with normal renal function. METHODS: Determination of telomere length by flow-fluorescence in situ hybridization (FISH), expression of surface molecules, and evaluation of apoptosis was performed by flow cytometry. RESULTS: In uremic predialysis patients, we observed a subpopulation of lymphocytes with short telomeres. However, in this population of patients we did not observe SIPS mononuclear cells. In hemodialysis patients, we found a subpopulation of SIPS mononuclear cells that also showed phenotypic changes of proinflammatory activity. Finally, transplant patients with normal renal function also exhibited a subpopulation of SIPS lymphocytes, which can be attributed to chronic lymphocyte activation induced by the major histocompatibility complex. CONCLUSION: In chronic kidney disease patients, immunocompetent cells undergo SIPS, a process associated with chronic cell activation and induced by numerous stimuli including uremia, hemodialysis membranes, and bacterial products. Because SIPS immunocompetent cells are activated cells with proinflammatory features and live longer in peripheral blood, it is likely that SIPS cells contribute significantly to the chronic inflammatory state of patients with advanced renal failure.     

Extending STR markers in Y chromosome haplotypes

Two multiplex reactions were developed to amplify 16 Y-STRs (DYS19, DYS385, DYS389 I and II, DYS390, DYS391, DYS392, DYS393, DYS437, DYS438, DYS439, GATA A7.1, GATA A7.2, GATA A10, GATA C4, GATA H4). Here we extend previous population studies done in a sample from northern Portugal for the GATA A7.1, GATA A7.2, GATA C4 and GATA H4 loci. A total of 199 different haplotypes identified by the 16 Y-STR markers were observed in a sample of 208 male individuals, of which 190 were unique and 9 were found twice. The overall haplotype diversity was 0.9996. The haplotype diversity of the Y-STR set composed of the 8 new markers is higher than the Y-STR core set included in the Y-STR haplotype reference database. Sequence structure of new alleles for GATA C4 and GATA H4 is reported. The usefulness of the inclusion of this new set of Y-STRs in forensic casework was also assessed. The increase in haplotype diversity with the addition of any new Y-STR marker to the 8 Y-STR core set is dependent not only on the gene diversity (positively) but also (negatively) on the degree of gametic association between the markers and the haplotypes previously defined. For instance, in our sample the addition of the DYS437, DYS438 and GATA A7.2 to a 13-locus set increased haplotype diversity only by 0.0001.     

Population study of eight novel Y-chromosome STRs (DYS460, DYS461, GATA-A10, GATA-C4, GATA-H4, DYS434, DYS437, DYS439) in a southeast Iberian population: looking for highly informative Y-chromosome haplotypes

The present study analyses 8 recently described tetranucleotide microsatellites (DYS460, DYS461, GATA-A10, GATA-C4, GATA-H4, DYS434, DYS437, DYS439) in southeast Spain and out of a total of 76 individuals 67 showed different haplotypes. Out of the 67 different haplotypes, 63 were present once, 3 were found 2 times, and 1 was found 7 times (9.21%). By combining the allelic states of the present eight Y-chromosome STRs with those previously carried out on the same individuals, highly informative haplotypes could be obtained. The haplotype diversity using the basic set of Y-STRs (DYS19, DYS389-I, DYS389-II, DYS390, DYS391, DYS392, DYS393, DYS385 and DYX156Y) previously analyzed is 0.9844. For the same individuals, this value using the new set of Y-STRs is slightly lower (0.8949), while the haplotype diversity combining the two sets of primers significantly increase to 0.9868. The results obtained in the present work show the usefulness of these microsatellites for individual identification and paternity testing in forensic genetics.     

On-line hemodiafiltration reduces the proinflammatory CD14+CD16+ monocyte-derived dendritic cells: A prospective, crossover study

It is not known whether high convective transport may have a role in modulating the chronic inflammation of hemodialysis (HD) patients. The aim of this study was to evaluate the effect of on-line hemodiafiltration (OL-HDF) on proinflammatory peripheral monocytes: Percentage of CD14+CD16+ cells and their telomere length and spontaneous or bacterial DNA-induced production of cytokines (TNF-alpha and IL-6). In a prospective, crossover study, 31 patients who were on high-flux HD (HF-HD) were evaluated. Patients underwent the following sequence of treatments (4 mo each): HF-HD (basal), OL-HDF (period 1), HF-HD (period 2), OL-HDF (period 3), and HF-HD (period 4). The dialysis characteristics were similar in the two modalities; the only difference was a higher convective transport in the OL-HDF than in the HF-HD. All patients who were on OL-HDF periods showed a significantly lower number of CD14+CD16+ cells than on HF-HD (18.5 +/- 2.3 basal versus 13.6 +/- 2.9 period 1 and 13.9 +/- 2.3 period 3; P = 0.001). By contrast, HF-HD restored the number of CD14+CD16+ cells to the basal values (19.2 +/- 2.8 and 18.6 +/- 1.4, periods 2 and 4, respectively; NS). During OL-HDF periods, the reduction of CD14+CD16+ was paralleled by a decreased number of short telomere cells. Spontaneous or bacterial DNA-induced production of cytokines (TNF-alpha and IL-6) was increased in HF-HD as compared with OL-HDF. In conclusion, these results demonstrate that as compared with HF-HD, OL-HDF markedly reduces the number of proinflammatory CD14+CD16+ cells and the production of TNF-alpha and IL-6. Future studies are needed to assess the possible therapeutic effect of convective transport on chronic inflammation that is associated with HD.     

Heteroplasmy in mtDNA and the weight of evidence in forensic mtDNA analysis: a case report

Mitochondrial DNA (mtDNA) sequencing has been validated as a useful tool for forensic analysis. However, there are several aspects of the analysis which need to be considered in order to evaluate the value of the evidence. One of these aspects is related to heteroplasmy which is the state when two or more mtDNA populations occur in a single individual, cell or mitochondrion. In this report a case is described where the mtDNA profile of the blood sample of a raped woman was compared with the mtDNA profile of a single hair found in the suspect’s car. The results obtained show differences in sequence between different portions of the hair and the victim’s sequence. These differences are related to various heteroplasmy events. The concordance between the hair sample and the potential source (victim) of this sample is questionable and the strength of the evidence depends on how the sequence information is interpreted by the expert. The discussion of the results emphasises the necessity to evaluate heteroplasmic events in routine forensic work. Received: 26 January 2000 / Accepted: 19 April 2000     

Role of adhesion molecules in mononuclear cell apoptosis induced by cuprophan hemodialysis membranes

BACKGROUND/AIM: Hemodialysis with Cuprophan (CU) membranes induces mononuclear cell activation, leading to increased expression of adhesion molecules, formation of cell aggregates, and apoptosis. It is likely that structure(s) of the CU membrane interact with mononuclear cell surface molecules which transduce biochemical signals to the cell. Interactions between adhesion molecules and extracellular matrix have been implicated in cell activation, proliferation, and/or apoptosis. In the present work, we study whether adhesion molecules may be involved in CU-induced mononuclear cell aggregation and/or apoptosis. METHODS: The present study was performed using THP-1 cells, a human monocytic cell line, cultured in the presence of the CU membrane. CD11b and CD54 expression was studied with fluorescent monoclonal antibodies. Cell aggregation was quantified using a phase-contrast microscope. Apoptosis was evaluated by either light microscopy or annexin V labeling. RESULTS: The results show that incubation of CU membranes with the proteins CD11b, CD18, and CD54 or the blockade of these cell surface molecules with specific monoclonal antibodies inhibited the CU-induced aggregation and apoptosis in a dose-dependent manner. CONCLUSION: These results suggest that CU membranes interact selectively with these specific proteins to induce cell activation which ultimately results in apoptosis.     

Rare HRAS1 alleles are a risk factor for the development of brain tumors.

BACKGROUND: The highly polymorphic HRAS1 minisatellite locus, located 1 kilobase downstream from the H-ras1 gene, has been associated with increased susceptibility to a variety of cancers. Microsatellite instability (MI), another molecular abnormality observed in human neoplasms, most likely reflects an increased mutation rate and also is thought to underlie cancer predisposition. The purpose of this study was to investigate the association between rare HRAS1 alleles and brain tumors and to correlate the HRAS1 allelotype with MI and clinicopathologic features. METHODS: Ninety-four patients with primary brain tumors (52 gliomas, 32 meningiomas, and 10 schwannomas) and 109 healthy control individuals were studied. The size of HRAS1 alleles was determined by fluorescent detection in an automated DNA sequencer. The interspersion pattern was assessed by the minisatellite variant repeat-polymerase chain reaction technique. RESULTS: Twenty of 94 (21.28%) patients with brain tumors had at least one rare allele, compared with 13 of 109 (11.92%) in the control population (Fisher exact test; P = 0.0329). The presence of rare alleles was associated with an increased risk of brain tumors (odds ratio, 1.99; 95% confidence interval, 0.93-4.27). The overrepresentation of rare alleles in tumor patients mainly reflects the higher frequency observed in the glioma group (P = 0.0086). The authors did not detect association between the presence of rare HRAS1 alleles and MI in their series. No significant difference in the distribution of these alleles was found when tumors were compared according to other clinicopathologic variables. CONCLUSIONS: The presence of rare HRAS1 alleles is associated with an increased risk for the development of glial neoplasms (OR = 2.72; 95% CI, 1.17-6.32). The lack of association between rare HRAS1 polymorphisms and MI suggests that these two genetic factors are not likely to be expression of the same underlying defect.     

Skp2, p27kip1 and EGFR assessment in head and neck squamous cell carcinoma: prognostic implications

EGFR, p27kip1 and Skp2, have been implicated in human cancer development. We have studied these molecules in a search for molecular markers that may have prognostic value in head and neck squamous cell carcinomas (HNSCC). Tissue samples of 62 patients were collected and immunohistochemical analysis was carried out for p27kip1, Skp2 and EGFR protein evaluation. Western blot analysis of p27kip1 was performed. The p27kip1 expression is frequently down-regulated in HNSCC (44.4%, 20/45 cases). The immunohistochemical analysis showed p27kip1 cytoplasmic retention in 7/38 tumors. Strong Skp2 signals were detected at the invasive edge of the tumor in cells lacking p27kip1 staining. We found a high EGFR staining in 49% (23/47) of the cases. The staining tended to be more frequent in lymph node-positive cases. The dysplastic tissue exhibited no Skp2 immunoreactivity, whereas 51.06% (24/47) of invasive tumors expressed high levels. Of note is that Skp2 overexpression was the only factor that significantly correlated with a shorter overall survival in multivariate analysis (p=0.048). Our results suggest that Skp2 is a useful prognostic marker for HNSCC management.