Analysis of the p16INK4 and TP53 Tumor Suppressor Genes in Bone Sarcoma Pediatric Patients

Recent data suggest that deletion of p16INK4 and mutation of TP53 are among the most common genetic events in the development of human cancer, since the codified proteins act as brakes of the abnormal cell cycle. As the molecular events leading to the development of pediatric bone sarcomas remain unclear, we analyzed 75 osteosarcoma and Ewing sarcoma samples from 43 pediatric patients to search for alterations at the TP53 or p16INK4 tumor suppressor genes. By means of PCR-DGGE (polymerase chain reaction and denaturing gradient gel electrophoresis) we detected TP53 point mutations in 18.6% of the tumor samples, but no constitutional mutations. In the analysis of p16INK4, 7% of the samples harbored deletions of the gene but no point mutations were detected by SSCP (single strand conformation polymorphism) analysis, just the polymorphism Ala-->Thr at codon 148. These data support the hypothesis that TP53 alterations may play a role in the development of pediatric bone tumors and that the primary mechanism of inactivation of p16INK4 seems to be homozygous deletion rather than point mutation

El ensayo de micronúcleos como medida de inestabilidad genética inducida por agentes genotóxicos

Human genetic integrity is compromised by the intense industrial activity, which emphasizes the importance to determine an "acceptable" genetic damage level and to carry out routine genotoxicity assays in the populations at risk. Micronuclei are cytoplasmatic bodies of nuclear origin which correspond to genetic material that is not correctly incorporated in the daughter cells in the cellular division; they reflect the existence of chromosomal aberrations and are originated by chromosomal breaks, replication errors followed by cellular division of the DNA and/or exposure to genotoxic agents. There are several factors able to modify the number of micronuclei present in a given cell, among them are age, gender, vitamins, medical treatments, daily exposure to genotoxic agents, etc. The cytogenetic assay for the detection of micronuclei (CBMN: cytokinesis-block micronucleus) is based on the use of a chemical agent, cytochalasin-B, which is able to block cytocinesis but allowing the nuclear division, therefore yielding binucleated and monodivided cells. The micronuclei scoring is performed on 1000 binucleated cells and the starting sample may vary, although most studies are performed on peripheral blood lymphocytes. The micronuclei assay is considered a practical, universally validated and technically feasible protocol which is useful to evaluate the genetic instability induced by genotoxic agent

Dental Treatment under General Anesthesia in Healthy and Medically Compromised/Developmentally Disabled Children: A Comparative Study

Aim: To compare the type, number of procedures and working time of dental treatment provided under dental general anesthesia (DGA) in healthy and medically compromised/developmentally disabled children (MCDD children). Design: This cross-sectional prospective study involved 80 children divided into two groups of 40 children each. Group 1 consisted of healthy and Group 2 consisted of MCDD children. Results: Healthy children needed more working time than MCDD children, the means being 161±7.9 and 84±5.7 minutes, respectively (P= 0.0001). Operative dentistry and endodontic treatments showed a significant statistical difference (P= 0.0001). The means of procedures were 17±5.0 for healthy children and 11±4.8 for MCDD children (P= 0.0001). Conclusions: Healthy children needed more extensive dental treatment than MCDD children under DGA. The information from this sample of Mexican children could be used as reference for determining trends both within a facility as well as in comparing facilities in cross-population studies

Screening of the human tumor necrosis factor-alpha (TNF-α) gene promoter polymorphisms by PCR–DGGE analysis

We have designed a new PCR-DGGE technique that enables detection of base changes in the TNF-alpha gene promoter. Screening of 130 samples from Spanish children has shown that this technique accurately detects the altered band patterns induced by the presence of the polymorphisms at positions -376, -308, -238 and -163 of the promoter sequence. Although further analysis are needed to fully characterise the alterations detected, we believe that this PCR-DGGE technique is a rapid and sensitive first approach to the genetic characterisation of the TNF-alpha promote

Methotrexate in Pediatric Osteosarcoma: Response and Toxicity in Relation to Genetic Polymorphisms and Dihydrofolate Reductase and Reduced Folate Carrier 1 Expression

To determine the influence of the genotype and the level of expression of different enzymes involved in folate metabolism on the response to and toxicity of high-dose methotrexate treatment in pediatric osteosarcomas. STUDY DESIGN: DHFR and Reduced folate carrier 1 (RFC1) semiquantitative expression was analyzed in 34 primary and metastatic osteosarcoma tissues by real-time polymerase chain reaction. The following polymorphisms were also analyzed in peripheral blood from 96 children with osteosarcoma and 110 control subjects: C677T, A1298C (MTHFR), G80A (RFC1), A2756G (MTR), C1420T (SHMT), the 28bp-repeat polymorphism, and 1494del6 of the TYMS gene. Treatment toxicity was scored after each cycle according to criteria from the World Health Organization. RESULTS: DHFR and RFC1 expression was lower in initial osteosarcoma biopsy specimens than in metastases (P = .024 and P = .041, respectively). RFC1 expression was moderately decreased in samples with poor histologic response to preoperative treatment (P = .053). Patients with osteosarcoma with G3/G4 hematologic toxicity were more frequently TT than CT/CC for C677T/MTHFR (P = .023) and GG for A2756G/MTR (P = .048 and P = .057 for gastrointestinal and hematologic toxicity, respectively). CONCLUSIONS: The role of C677T/MTHFR and A2756G/MTR on chemotherapy-induced toxicity should be further investigated in pediatric osteosarcomas receiving high-dose methotrexate. Altered expression of DHFR and RFC1 is a feasible mechanism by which osteosarcoma cells become resistant to methotrexate

Looking for blazars in a sample of unidentified high-energy emitting Fermi sources

Context. Based on their overwhelming dominance among associated Fermi γ-ray catalogue sources, it is expected that a large fraction of the unidentified Fermi objects are blazars. Through crossmatching between the positions of unidentified γ-ray sources from the First Fermi Catalog of γ-ray sources emitting above 10 GeV (1FHL) and the ROSAT and Swift/XRT catalogues of X-ray objects and between pointed XRT observations, a sample of 36 potential associations was found in previous works with less than 15 arcsec of positional offset. One-third of them have recently been classified; the remainder, though believed to belong to the blazar class, still lack spectroscopic classifications. Aims. We study the optical spectrum of the putative counterparts of these unidentified gamma-ray sources in order to find their redshifts and to determine their nature and main spectral characteristics. Methods. An observational campaign was carried out on the putative counterparts of 13 1FHL sources using medium-resolution optical spectroscopy from the Osservatorio Astronomico di Bologna in Loiano, Italy; the Telescopio Nazionale Galileo and the Nordic Optical Telescope, both in the Canary Islands, Spain; and the Observatorio Astronómico Nacional San Pedro Mártir in Baja California, Mexico. Results. We were able to classify 14 new objects based on their continuum shapes and spectral features. Conclusions. Twelve new blazars were found, along with one new quasar and one new narrow line Seyfert 1 (NLS1) to be potentially associated with the 1FHL sources of our sample. Redshifts or lower limits were obtained when possible alongside central black hole mass and luminosity estimates for the NLS1 and the quasar.Fil: Marchesini, Ezequiel Joaquín. Universidad Nacional de la Plata. Facultad de Ciencias Astronómicas y Geofísicas; ArgentinaFil: Masetti, Nicola. Istituto di Astrofisica Spaziale e Fisica Cosmica di Bologna; ItaliaFil: Chavushyan, V.. Instituto Nacional de Astrofísica, Óptica y Electrónica; MéxicoFil: Cellone, Sergio Aldo. Universidad Nacional de la Plata. Facultad de Ciencias Astronómicas y Geofísicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Astrofísica La Plata. Universidad Nacional de La Plata. Facultad de Ciencias Astronómicas y Geofísicas. Instituto de Astrofísica La Plata; ArgentinaFil: Andruchow, Ileana. Universidad Nacional de la Plata. Facultad de Ciencias Astronómicas y Geofísicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto de Astrofísica La Plata. Universidad Nacional de La Plata. Facultad de Ciencias Astronómicas y Geofísicas. Instituto de Astrofísica La Plata; ArgentinaFil: Bassani, L.. Istituto di Astrofisica Spaziale e Fisica Cosmica di Bologna; ItaliaFil: Bazzano, A.. Istituto di Astrofisica e Planetologia Spaziali; ItaliaFil: Jiménez-Bailón, E.. Universidad Nacional Autónoma de México; MéxicoFil: Landi, R.. Istituto di Astrofisica Spaziale e Fisica Cosmica di Bologna; ItaliaFil: Malizia, A.. Istituto di Astrofisica Spaziale e Fisica Cosmica di Bologna; ItaliaFil: Palazzi, E.. Istituto di Astrofisica Spaziale e Fisica Cosmica di Bologna; ItaliaFil: Patiño Álvarez, V.. Instituto Nacional de Astrofísica, Óptica y Electrónica; MéxicoFil: Rodríguez Castillo, G. A.. Osservatorio Astronomico di Roma; ItaliaFil: Stephen, J. B.. Istituto di Astrofisica Spaziale e Fisica Cosmica di Bologna; ItaliaFil: Ubertini, P.. Istituto di Astrofisica e Planetologia Spaziali; Itali