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Biomedical subjects

Daniela Maurici

Publications and source records attributed to Daniela Maurici.

8 recordsLinked to original sources
PubMed2006

Meeting report: Validation of toxicogenomics-based test systems: ECVAM-ICCVAM/NICEATM considerations for regulatory use.

This is the report of the first workshop "Validation of Toxicogenomics-Based Test Systems" held 11-12 December 2003 in Ispra, Italy. The workshop was hosted by the European Centre for the Validation of Alternative Methods (ECVAM) and organized jointly by ECVAM, the U.S. Interagency Coordinating Committee on the Validation of Alternative Methods (ICCVAM), and the National Toxicology Program (NTP) Interagency Center for the Evaluation of Alternative Toxicological Methods (NICEATM). The primary aim of the workshop was for participants to discuss and define principles applicable to the validation of toxicogenomics platforms as well as validation of specific toxicologic test methods that incorporate toxicogenomics technologies. The workshop was viewed as an opportunity for initiating a dialogue between technologic experts, regulators, and the principal validation bodies and for identifying those factors to which the validation process would be applicable. It was felt that to do so now, as the technology is evolving and associated challenges are identified, would be a basis for the future validation of the technology when it reaches the appropriate stage. Because of the complexity of the issue, different aspects of the validation of toxicogenomics-based test methods were covered. The three focus areas include a) biologic validation of toxicogenomics-based test methods for regulatory decision making, b) technical and bioinformatics aspects related to validation, and c) validation issues as they relate to regulatory acceptance and use of toxicogenomics-based test methods. In this report we summarize the discussions and describe in detail the recommendations for future direction and priorities.

Animal Testing Alternatives↗
PubMed2005

Carcinogenicity.

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Animal Testing Alternatives↗
PubMed2004

New insights into the mechanisms involved in renal proximal tubular damage induced in vitro by ochratoxin A.

The mycotoxin ochratoxin A is a contaminant of human and animal food products. It is a potent nephrotoxin known to damage the proximal tubule. The aim of this work was to investigate the effects of ochratoxin A on a porcine renal proximal tubular epithelial cell line (LLC-PK1), and to identify sensitive endpoints revealing damage at the epithelial barrier level and at the molecular level. Cells exposed for 24 h to 5-10 microM ochratoxin indicated a clear damage to the intactness of the epithelial barrier, as shown by measurements of trans-epithelial resistance and zonula occludens-1 protein expression. At the mitochondrial level we observed alterations of the normal functions, such as an increase of the membrane potential, the formation of straight extensions, and the formation of giant mitochondria. At higher ochratoxin concentrations (50 microM), at which cytotoxicity assays revealed a significant toxicity, alterations of the cytoskeleton organization and induction of apoptosis were evident. In addition, we analyzed the expression of genes by using a cDNA macroarray. Our data indicate that ochratoxin-induced nephrotoxicity can be detected at the barrier and at the mitochondrial level at rather low concentrations, at which conventional cytotoxicity assays are unable to reveal toxic effects.

Animals↗
PubMed2004

The tumor suppressor gene TP53: implications for cancer management and therapy.

The p53 protein is an inducible transcription factor with multiple anti-proliferative roles in response to genotoxic damage; unprogrammed proliferative stimuli; and deprivation of oxygen, nutrients, or ribonucleotides. Inactivation of the TP53 gene by mutation or deletion is the most common event in human cancer. Loss of p53 function compromises genetic homeostasis in cells exposed to mutagens and prevents normal cytotoxic responses to cancer therapies. Genetic and pharmacological approaches are being developed with the ultimate goal of restoring or controlling p53 functions in cancer patients. Genetic interventions aiming at expressing wild-type TP53 in cancer cells, either by retroviral or adenoviral transfer, have met limited clinical success. However, recently, the use of a defective adenovirus (ONYX-015) that selectively kills p53-incompetent cells has shown promising effects in pre-clinical and clinical studies. Pharmacological methods are under development to either stimulate wild-type p53 protein function or induce p53 mutant proteins to resume wild-type functions. These methods are based on small chemicals (CP-31388, PRIMA-1), peptides (CDB3), or single-chain Fv antibody fragments corresponding to defined p53 domains. In addition, detection of mutant TP53 may also serve as a marker for early cancer detection, prediction, and prognosis. In this review, we discuss the mechanisms underlying these approaches and their perspectives for cancer therapy.

Female↗
PubMed2002

Initiation of human astrocytoma by clonal evolution of cells with progressive loss of p53 functions in a patient with a 283H TP53 germ-line mutation: evidence for a precursor lesion.

Little is known about the genetic and molecular events leading to the early stages of human astrocytoma formation. To examine this issue, we analyzed the significance of sequential accumulation of two somatic point mutations (R267W and E258D) in the TP53 gene during the initiation of astrocytoma in a patient born with a single germ-line p53 point mutation (R283H). We adapted a p53 transcriptional assay in yeast to establish the temporal occurrence and allelic distribution of the p53 mutations present in the patient and characterized these mutations through functional assays and structural modeling. Our results show that the first somatic mutation occurred at codon 267 on the p53 allele harboring the germ-line mutation R283H, whereas the second somatic mutation occurred in the remaining wild-type (wt) allele at codon 258. These two mutations induced the formation of tumor cells with the genotype p53(267W+283H/258D), which comprised 70% of the cells in the primary WHO grade II astrocytoma. Another 8% of cells within the tumor had the partially mutated genotype p53(267W+283H/WT) and represented the remnants of a clinically undetectable intermediate stage of astrocytic neoplastic transformation. The remaining 22% of cells had the constitutive p53(283H/WT) genotype and likely consisted of nontumor cells. Functional analysis of the p53 alleles present in the patient's tumor indicated that the germ-line p53(R283H) could transactivate the CDKN1A((p21, WAF1, cip1, SDI1)) but not the BAX gene and retained the ability to induce growth arrest of human glioblastoma cells. The p53(R267W+R283H) and p53(E258D) were incapable of transactivating either promoter or inducing growth arrest. Modeling of p53 interaction with DNA suggests that R283H mutation may weaken the sequence-specific interaction of p53 lysine 120 with the BAX gene but not the CDKN1A p53-responsive elements. Taken together, these results have characterized, for the first time, the genetic events defining a clinically undetectable precursor lesion leading to a grade II astrocytoma. They also suggest that astrocytoma initiation in this patient resulted from monoclonal evolution driven by a sequential loss of proapoptotic and growth arrest functions of p53.

Adult↗
PubMed2002

Restoration of wild-type conformation and activity of a temperature-sensitive mutant of p53 (p53(V272M)) by the cytoprotective aminothiol WR1065 in the esophageal cancer cell line TE-1.

The aminothiol WR1065, the active metabolite of the cytoprotector amifostine, exerts its antimutagenic effects through free-radical scavenging and other unknown mechanisms. In an earlier report, we showed that WR1065 activates wild-type p53 in MCF-7 cells, leading to p53-dependent arrest in the G(1) phase of the cell cycle. To determine whether WR1065 activates p53 by modulating protein conformation, we analyzed its effects on p53 conformation and activity in the esophageal cancer cell line TE-1. This cell line contains a mutation in codon 272 of p53 (p53(V272M), with methionine instead of a valine), conferring temperature-sensitive properties to the p53 protein. At the nonpermissive temperature (37 degrees C), p53(V272M) adopts the mutant p53 conformation (nonreactive with the antibody PAb1620), does not bind specifically to DNA, and is not activated in response to DNA-damaging treatment. However, treatment with 0.5-4 mM WR1065 partially restored wild-type conformation at 37 degrees C, stimulated DNA binding activity, and increased the expression of p53 target genes WAF-1, GADD45, and MDM2, leading to cell-cycle arrest in G(1). These results suggest that WR1065 activates p53 through a mechanism distinct from DNA-damage signaling, which involves modulation of p53 protein conformation.

Cell Cycle↗
PubMedDate not supplied

Modulation of different stress pathways after styrene and styrene-7,8-oxide exposure in HepG2 cell line and normal human hepatocytes.

Styrene is one of the most important monomers produced worldwide. IARC classified styrene as a possible carcinogen to humans (group 2B). Styrene-7,8-oxide (SO) is the main reactive metabolite of styrene, and it is found to be genotoxic in several in vitro test systems. Styrene and styrene-7,8-oxide (SO) toxicity to HepG2 cells was investigated by evaluating end-points such as heat shock proteins (Hsps), metallothioneins (MT), apoptosis-related proteins, accumulation of styrene within the cells and expression of two isoforms of cytochrome P450. The potential activity of styrene and styrene-7,8-oxide in modulating gene expression was also investigated. The results showed induction of Hsp70, metallothioneins, BclX(S/L) and c-myc expression and a decrease in Bax expression in HepG2 after treatments, confirming that these compounds activated protective mechanisms. Moreover, up-regulation of TGFbeta2 and TGFbetaRIII in HepG2 cells was found after exposure to styrene, while in human primary hepatocytes these genes were down-regulated after both treatments. Finally, it was found that styrene and SO treatments did not induce CYP1A2 and CYP2E1 protein expression. In conclusion, both compounds caused toxic stress in HepG2 cells, with SO being more toxic; in the meantime, a different effect of the two compounds in HepG2 cells and primary human hepatocytes was observed regarding their activity in gene modulation.

Carcinogens↗