9 results on '"Finotti, Alessia"'
Search Results
2. UPF1 silenced cellular model systems for screening of read-through agents active on β039 thalassemia point mutation
- Author
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Salvatori, Francesca, Pappadà, Mariangela, Breveglieri, Giulia, D’Aversa, Elisabetta, Finotti, Alessia, Lampronti, Ilaria, Gambari, Roberto, and Borgatti, Monica
- Published
- 2018
- Full Text
- View/download PDF
3. An Aγ-globin G->A gene polymorphism associated with β039 thalassemia globin gene and high fetal hemoglobin production.
- Author
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Breveglieri, Giulia, Bianchi, Nicoletta, Cosenza, Lucia Carmela, Gamberini, Maria Rita, Chiavilli, Francesco, Zuccato, Cristina, Montagner, Giulia, Borgatti, Monica, Lampronti, Ilaria, Finotti, Alessia, and Gambari, Roberto
- Subjects
GENETIC polymorphisms ,THALASSEMIA ,GLOBIN genes ,FETAL hemoglobin ,GENETIC mutation ,ERYTHROCYTE membranes - Abstract
Background: Increase of the expression of γ-globin gene and high production of fetal hemoglobin (HbF) in β-thalassemia patients is widely accepted as associated with a milder or even asymptomatic disease. The search for HbF-associated polymorphisms (such as the XmnI, BCL11A and MYB polymorphisms) has recently gained great attention, in order to stratify β-thalassemia patients with respect to expectancy of the first transfusion, need for annual intake of blood, response to HbF inducers (the most studied of which is hydroxyurea). Methods: Aγ-globin gene sequencing was performed on genomic DNA isolated from a total of 75 β-thalassemia patients, including 31 β
0 39/β0 39, 33 β0 39/β+ IVSI-110, 9 β+ IVSI-110/β+ IVSI-110, one β0 IVSI-1/β+ IVSI-6 and one β0 39/β+ IVSI-6. Results: The results show that the rs368698783 polymorphism is present in β-thalassemia patients in the 5'UTR sequence (+25) of the Aγ-globin gene, known to affect the LYAR (human homologue of mouse Ly-1 antibody reactive clone) binding site 5'-GGTTAT-3'. This Aγ(+25 G->A) polymorphism is associated with the Gγ-globin-XmnI polymorphism and both are linked with the β0 39-globin gene, but not with the β+ IVSI-110-globin gene. In agreement with the expectation that this mutation alters the LYAR binding activity, we found that the Aγ(+25 G->A) and Gγ-globin-XmnI polymorphisms are associated with high HbF in erythroid precursor cells isolated from β0 39/β0 39 thalassemia patients. Conclusions: As a potential explanation of our findings, we hypothesize that in β-thalassemia the Gγ-globin-XmnI/Aγ-globin-(G->A) genotype is frequently under genetic linkage with β0 -thalassemia mutations, but not with the β+ -thalassemia mutation here studied (i.e. β+ IVSI-110) and that this genetic combination has been selected within the population of β0-thalassemia patients, due to functional association with high HbF. Here we describe the characterization of the rs368698783 (+25 G->A) polymorphism of the Aγ-globin gene associated in β0 39 thalassemia patients with high HbF in erythroid precursor cells. [ABSTRACT FROM AUTHOR]- Published
- 2017
- Full Text
- View/download PDF
4. A validated cellular biobank for β-thalassemia.
- Author
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Cosenza, Lucia Carmela, Breda, Laura, Breveglieri, Giulia, Zuccato, Cristina, Finotti, Alessia, Lampronti, Ilaria, Borgatti, Monica, Chiavilli, Francesco, Gamberini, Maria Rita, Satta, Stefania, Manunza, Laura, De Martis, Franca Rosa, Moi, Paolo, Rivella, Stefano, Gambari, Roberto, and Bianchi, Nicoletta
- Subjects
THALASSEMIA ,GENE therapy ,PHLEBOTOMISTS ,REPRODUCIBLE research ,ERYTHROCYTES ,MEDICAL research - Abstract
Background: Cellular biobanking is a key resource for collaborative networks planning to use same cells in studies aimed at solving a variety of biological and biomedical issues. This approach is of great importance in studies on β-thalassemia, since the recruitment of patients and collection of specimens can represent a crucial and often limiting factor in the experimental planning.Methods: Erythroid precursor cells were obtained from 72 patients, mostly β-thalassemic, expanded and cryopreserved. Expression of globin genes was analyzed by real time RT-qPCR. Hemoglobin production was studied by HPLC.Results: In this paper we describe the production and validation of a Thal-Biobank constituted by expanded erythroid precursor cells from β-thalassemia patients. The biobanked samples were validated for maintenance of their phenotype after (a) cell isolation from same patients during independent phlebotomies, (b) freezing step in different biobanked cryovials, (c) thawing step and analysis at different time points. Reproducibility was confirmed by shipping the frozen biobanked cells to different laboratories, where the cells were thawed, cultured and analyzed using the same standardized procedures. The biobanked cells were stratified on the basis of their baseline level of fetal hemoglobin production and exposed to fetal hemoglobin inducers.Conclusion: The use of biobanked cells allows stratification of the patients with respect to fetal hemoglobin production and can be used for determining the response to the fetal hemoglobin inducer hydroxyurea and to gene therapy protocols with reproducible results. [ABSTRACT FROM AUTHOR]- Published
- 2016
- Full Text
- View/download PDF
5. Regulation of IL-8 gene expression in gliomas by microRNA miR-93.
- Author
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Fabbri, Enrica, Brognara, Eleonora, Montagner, Giulia, Ghimenton, Claudio, Eccher, Albino, Cantù, Cinzia, Khalil, Susanna, Bezzerri, Valentino, Provezza, Lisa, Bianchi, Nicoletta, Finotti, Alessia, Borgatti, Monica, Moretto, Giuseppe, Chilosi, Marco, Cabrini, Giulio, and Gambari, Roberto
- Subjects
INTERLEUKIN-8 ,GENE expression ,GLIOMAS ,MICRORNA ,VASCULAR endothelial growth factors ,NEOVASCULARIZATION - Abstract
Background: Different strategies have been proposed to target neoangiogenesis in gliomas, besides those targeting Vascular Endothelial Growth Factor (VEGF). The chemokine Interleukin-8 (IL-8) has been shown to possess both tumorigenic and proangiogenic properties. Although different pathways of induction of IL-8 gene expression have been already elucidated, few data are available on its post-transcriptional regulation in gliomas.Methods: Here we investigated the role of the microRNA miR-93 on the expression levels of IL-8 and other pro-inflammatory genes by RT-qPCR and Bio-Plex analysis. We used different disease model systems, including clinical samples from glioma patients and two glioma cell lines, U251 and T98G.Results: IL-8 and VEGF transcripts are highly expressed in low and high grade gliomas in respect to reference healthy brain; miR-93 expression is also increased and inversely correlated with transcription of IL-8 and VEGF genes. Computational analysis showed the presence of miR-93 consensus sequences in the 3'UTR region of both VEGF and IL-8 mRNAs, predicting possible interaction with miR-93 and suggesting a potential regulatory role of this microRNA. In vitro transfection with pre-miR-93 and antagomiR-93 inversely modulated VEGF and IL-8 gene expression and protein release when the glioma cell line U251 was considered. Similar data were obtained on IL-8 gene regulation in the other glioma cell line analyzed, T98G. The effect of pre-miR-93 and antagomiR-93 in U251 cells has been extended to the secretion of a panel of cytokines, chemokines and growth factors, which consolidated the concept of a role of miR-93 in IL-8 and VEGF gene expression and evidenced a potential regulatory role also for MCP-1 and PDGF (also involved in angiogenesis).Conclusion: In conclusion, our results suggest an increasing role of miR-93 in regulating the level of expression of several genes involved in the angiogenesis of gliomas. [ABSTRACT FROM AUTHOR]- Published
- 2015
- Full Text
- View/download PDF
6. Complexation to Cationic Microspheres of Double-Stranded Peptide Nucleic Acid-DNA Chimeras Exhibiting Decoy Activity.
- Author
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Mischiati, Carlo, Sereni, Alessia, Finotti, Alessia, Breda, Laura, Cortesi, Rita, Nastruzzi, Claudio, Romanelli, Alessandra, Saviano, Michele, Bianchi, Nicoletta, Pedone, Carlo, Borgatti, Monica, and Gambari, Roberto
- Subjects
NUCLEIC acids ,DNA ,CHIMERAS (Botany) ,MICROSPHERES ,GENE therapy - Abstract
The major aim of this paper was to determine whether cationic microspheres (CM), consisting of the permeable polymer Eudragit
® RS 100 plus the cationic surfactant dioctadecyl-dimethyl-ammonium bromide (DDAB18 ), could bind to double-stranded peptide nucleic acid PNA-DNA-PNA (PDP) chimeras exhibiting decoy activity against NF-κB transcription factors. Microspheres were produced by the ‘solvent evaporation method’ and centrifugation at 500, 1,000 and 3,000 rpm to obtain different-sized microparticles. Microsphere morphology, size and size distribution were determined by optical and electron microscopy observations. In order to determine their binding activity, double-stranded DNA-based and PDP-based decoy molecules were incubated with different amounts of microparticles in the presence of 100 ng of either32 P-labeled DNA-DNA or DNA-PDP hybrid molecules or cold PDP-PDP hybrids. The complexes were analyzed by agarose gel electrophoresis. The resistance of32 P-labeled DNA-DNA and DNA-PDP molecules in the presence of serum or cellular extracts was evaluated after binding to CM by gel electrophoresis analysis. DDAB18 Eudragit RS 100 microspheres are able to bind to DNA-PDP and PDP-PDP hybrids, to deliver these molecules to target cells and to protect DNA-PDP molecules from enzymatic degradation in simulated biological fluids. In addition, when assayed in ex vivo conditions, DDAB18 Eudragit RS 100 microspheres exhibited low toxicity. The results presented in this paper demonstrate that CM can be considered suitable formulations for pharmacogenomic therapy employing double-stranded PDP chimeras. Copyright © 2004 National Science Council, ROC and S. Karger AG, Basel [ABSTRACT FROM AUTHOR]- Published
- 2004
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- View/download PDF
7. UPF1 silenced cellular model systems for screening of read-through agents active on β039 thalassemia point mutation.
- Author
-
Salvatori, Francesca, Pappadà, Mariangela, Breveglieri, Giulia, D'Aversa, Elisabetta, Finotti, Alessia, Lampronti, Ilaria, Gambari, Roberto, and Borgatti, Monica
- Subjects
THALASSEMIA treatment ,NONSENSE mutation ,MESSENGER RNA ,STOP codons ,PHARMACOLOGY - Abstract
Background: Nonsense mutations promote premature translational termination, introducing stop codons within the coding region of mRNAs and causing inherited diseases, including thalassemia. For instance, in β
0 39 thalassemia the CAG (glutamine) codon is mutated to the UAG stop codon, leading to premature translation termination and to mRNA destabilization through the well described NMD (nonsense-mediated mRNA decay). In order to develop an approach facilitating translation and, therefore, protection from NMD, ribosomal read-through molecules, such as aminoglycoside antibiotics, have been tested on mRNAs carrying premature stop codons. These findings have introduced new hopes for the development of a pharmacological approach to the β0 39 thalassemia therapy. While several strategies, designed to enhance translational read-through, have been reported to inhibit NMD efficiency concomitantly, experimental tools for systematic analysis of mammalian NMD inhibition by translational read-through are lacking. Results: We developed a human cellular model of the β0 39 thalassemia mutation with UPF-1 suppressed and showing a partial NMD suppression. Conclusions: This novel cellular model could be used for the screening of molecules exhibiting preferential read-through activity allowing a great rescue of the mutated transcripts. [ABSTRACT FROM AUTHOR]- Published
- 2018
- Full Text
- View/download PDF
8. An Aγ-globin G->A gene polymorphism associated with β 0 39 thalassemia globin gene and high fetal hemoglobin production.
- Author
-
Breveglieri G, Bianchi N, Cosenza LC, Gamberini MR, Chiavilli F, Zuccato C, Montagner G, Borgatti M, Lampronti I, Finotti A, and Gambari R
- Subjects
- Binding Sites genetics, DNA-Binding Proteins metabolism, Female, Humans, Linkage Disequilibrium, Male, Nuclear Proteins metabolism, Point Mutation, Sequence Analysis, DNA, gamma-Globins metabolism, Fetal Hemoglobin biosynthesis, Polymorphism, Genetic, beta-Thalassemia genetics, gamma-Globins genetics
- Abstract
Background: Increase of the expression of γ-globin gene and high production of fetal hemoglobin (HbF) in β-thalassemia patients is widely accepted as associated with a milder or even asymptomatic disease. The search for HbF-associated polymorphisms (such as the XmnI, BCL11A and MYB polymorphisms) has recently gained great attention, in order to stratify β-thalassemia patients with respect to expectancy of the first transfusion, need for annual intake of blood, response to HbF inducers (the most studied of which is hydroxyurea)., Methods: Aγ-globin gene sequencing was performed on genomic DNA isolated from a total of 75 β-thalassemia patients, including 31 β
0 39/β0 39, 33 β0 39/β+ IVSI-110, 9 β+ IVSI-110/β+ IVSI-110, one β0 IVSI-1/β+ IVSI-6 and one β0 39/β+ IVSI-6., Results: The results show that the rs368698783 polymorphism is present in β-thalassemia patients in the 5'UTR sequence (+25) of the Aγ-globin gene, known to affect the LYAR (human homologue of mouse Ly-1 antibody reactive clone) binding site 5'-GGTTAT-3'. This Aγ(+25 G->A) polymorphism is associated with the Gγ-globin-XmnI polymorphism and both are linked with the β0 39-globin gene, but not with the β+ IVSI-110-globin gene. In agreement with the expectation that this mutation alters the LYAR binding activity, we found that the Aγ(+25 G->A) and Gγ-globin-XmnI polymorphisms are associated with high HbF in erythroid precursor cells isolated from β0 39/β0 39 thalassemia patients., Conclusions: As a potential explanation of our findings, we hypothesize that in β-thalassemia the Gγ-globin-XmnI/Aγ-globin-(G->A) genotype is frequently under genetic linkage with β0 -thalassemia mutations, but not with the β+ -thalassemia mutation here studied (i.e. β+ IVSI-110) and that this genetic combination has been selected within the population of β0 -thalassemia patients, due to functional association with high HbF. Here we describe the characterization of the rs368698783 (+25 G->A) polymorphism of the Aγ-globin gene associated in β0 39 thalassemia patients with high HbF in erythroid precursor cells.- Published
- 2017
- Full Text
- View/download PDF
9. Upstream stimulatory factors are involved in the P1 promoter directed transcription of the A beta H-J-J locus.
- Author
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Finotti A, Treves S, Zorzato F, Gambari R, and Feriotto G
- Subjects
- Base Sequence, Binding Sites, Cell Line, Tumor, Genes, Reporter, HeLa Cells, Humans, Mixed Function Oxygenases metabolism, Molecular Sequence Data, Mutation, Upstream Stimulatory Factors genetics, Mixed Function Oxygenases genetics, Promoter Regions, Genetic, Transcription, Genetic, Upstream Stimulatory Factors metabolism
- Abstract
Background: Alternative splicing of the locus A beta H-J-J generates functionally distinct proteins: the enzyme aspartyl (asparaginyl) beta-hydroxylase (AAH), truncated homologs of AAH with a role in calcium homeostasis humbug and junctate and a structural protein of the sarcoplasmic reticulum membranes junctin. AAH and humbug are over expressed in a broad range of malignant neoplasms. We have previously reported that this locus contains two promoters, P1 and P2. While AAH and humbug are expressed in most tissues under the regulation of the P1 promoter, AAH, junctin and junctate are predominantly expressed in excitable tissues under the control of the P2 promoter. We previously demonstrated that Sp transcription factors positively regulate the P1 promoter., Results: In the present study, we extended the functional characterization of the P1 promoter of the A beta H-J-J locus. We demonstrated by quantitative Real-time RT-PCR that mRNAs from the P1 promoter are actively transcribed in all the human cell lines analysed. To investigate the transcription mechanism we transiently transfected HeLa cells with sequentially deleted reporter constructs containing different regions of the -661/+81 P1 nucleotide sequence. Our results showed that (i) this promoter fragment is a powerful activator of the reporter gene in HeLa cell line, (ii) the region spanning 512 bp upstream of the transcription start site exhibits maximal level of transcriptional activity, (iii) progressive deletions from -512 gradually reduce reporter expression. The region responsible for maximal transcription contains an E-box site; we characterized the molecular interactions between USF1/2 with this E-box element by electrophoretic mobility shift assay and supershift analysis. In addition, our USF1 and USF2 chromatin immunoprecipitation results demonstrate that these transcription factors bind the P1 promoter in vivo. A functional role of USF1/USF2 in upregulating P1-directed transcription was demonstrated by analysis of the effects of (i) in vitro mutagenesis of the P1/E-box binding site, (ii) RNA interference targeting USF1 transcripts., Conclusion: Our results suggest that USF factors positively regulate the core of P1 promoter, and, together with our previously data, we can conclude that both Sp and USF DNA interaction and transcription activity are involved in the P1 promoter dependent expression of AAH and humbug.
- Published
- 2008
- Full Text
- View/download PDF
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