Silvia Ciafrè - Academia.edu (original) (raw)
Papers by Silvia Ciafrè
Rivista europea per le scienze mediche e farmacologiche = European review for medical and pharmacological sciences = Revue européenne pour les sciences médicales et pharmacologiques
Cardiac function and morphology in chronic hemodialyzed patients are modified in consequence of b... more Cardiac function and morphology in chronic hemodialyzed patients are modified in consequence of both vascular and neurohormonal factors. In the present study we investigate on the role of prostacyclin (PGI2) vasodilator agent, during hemodialytic (HD) treatment. Twenty-four patients (13 males and 11 females; 9 hypertensive and 15 normotensive) aged 58.5 +/- 14.4 years were studied; 2.5 ml of venous blood were collected before (time 0) and 15', 120', and 240' of dialytic session. The PGI2 levels were measured in plasma, after extraction in ethyl acetate by RIA method, as levels of 6-Keto-PGF1 alpha, a stable metabolite. The results have shown as increase of PGI2 levels at 15' in hypertensive HD patients (HHD) from the begin of dialysis that increased until 240'. This phenomenon was more significant in hypertensive than in normotensive group (NHD) (p < 0.05 vs NHD). These preliminary data suggest that in HHD patients the role of PGI2 is more important than in NHD patients as regards the effects on regulation of circulatory tone. The increment of PGI2 levels could be in relation with the sympathetic activation occurred during hemodialytic treatment.
The EMBO Journal
The expression of the yeast L2 r-protein gene is controlled at the level of mRNA accumulation. Th... more The expression of the yeast L2 r-protein gene is controlled at the level of mRNA accumulation. The product of the gene appears to participate in this regulation by an autogenous feedback mechanism. This control does not operate at the level of transcription but instead affects L2 mRNA accumulation. This autogenous regulation of mRNA accumulation provides an interesting analogy to the autogenous translational regulation of r-proteins in Escherichia coli. Key words: autogenous regulation/ribosomal protein/RNA stability/yeast an autogenous feedback control in which the LI r-protein controls its synthesis by regulating the efficiency of splicing of its own pre-mRNA .
Recenti progressi in medicina
Atrial natriuretic factor (ANF) is a potent natriuretic and vasoactive (vasorelaxant) peptide loc... more Atrial natriuretic factor (ANF) is a potent natriuretic and vasoactive (vasorelaxant) peptide localized in the secretory-like atrial specific granules. The main peptide in this storage granules is the 126 amino acid proatrial natriuretic peptide, but the principal circulating form in human plasma is the 28 amino acid, alpha-human natriuretic peptide. Animal and in vitro studies have suggested that ANF modulates autonomic circulatory control, probably with a dose-dependent mechanism. Moreover, recent human studies have resulted contradictory. In particular, it is still unclear if high circulating levels of ANF, which are present in congestive heart diseases constantly, may be correlated with sympathetic nervous system activity in man. Previously we have shown that in congestive diseases there is a relation between ANF and catecholamine secretion. From these basis, the aim of this study was to investigate on the pathophysiological relations between atrial natriuretic factor (ANF) release and adrenergic activation in patients with obstructive hypertrophic cardiomyopathy (n = 6) and non obstructive hypertrophic cardiomyopathy (n = 4). Sympathetic activation in physiologic way was induced by cycloergometer sub-maximal exercise. Then specimens of venous blood were achieved for plasma determination of ANF and catecholamines pre- and post-exercise. Results have shown that in obstructive hypertrophic cardiomyopathy patients basal levels of ANF and catecholamines were higher than levels of these parameters in non obstructive hypertrophic cardiomyopathy patients.(ABSTRACT TRUNCATED AT 250 WORDS)
Recenti progressi in medicina
Endothelin-1 (ET-1) is an endothelium-derived vasoconstrictor peptide isolated from the culture s... more Endothelin-1 (ET-1) is an endothelium-derived vasoconstrictor peptide isolated from the culture supernatant of porcine aortic endothelial cells. This 21 amino-acid residue peptide has potent vasoconstrictive properties in vitro and in vivo. ET-1 action involves phosphatidylinositol turnover, calcium mobilization and protein kinase C activation. Endothelial cells have distinct receptors for different operating through hydrosoluble hormones. The aim of this study was to investigate on a possible role of angiotensin II (ANG II) to modulate the release ET-1 from human endothelial cells in vitro. These data revealed a time- and a dose-dependent increase of ET-1 production in response to ANG II. This mechanism may have important pathophysiological implications in vivo. In fact, a double-mechanism of secretion of ET-1 from endothelial cells could exist: one active in a physiological condition and an other in response to a vasoconstrictor stimuli (as well as ANG II). Furthermore, these results may suggest an additional favourable effect of ACE-inhibition in human hypertension therapy.
Virology, Jan 9, 2015
Adenosine deaminase acting on RNA1 (ADAR1) was previously reported to affect HIV-1 replication. W... more Adenosine deaminase acting on RNA1 (ADAR1) was previously reported to affect HIV-1 replication. We report data showing that ADAR1 interacts with the HIV-1 p55 Gag protein, the major structural protein of the immature virus capsid. Furthermore, we found that the endogenous ADAR1 is incorporated into virions purified from the supernatant of primary HIV-1-infected CD4(+) T lymphocytes. Additional experiments demonstrated that the expression of the p55 Gag protein is sufficient for ADAR1 incorporation into virus-like particles (VLPs). Overall, our data originally support the evidence that ADAR1 can be part of the cell protein array uploaded in HIV-1 particles.
International journal of molecular sciences, 2015
Spinal muscular atrophy (SMA) is an inherited neuromuscular disorder and the leading genetic caus... more Spinal muscular atrophy (SMA) is an inherited neuromuscular disorder and the leading genetic cause of death in infants. Despite the disease-causing gene, survival motor neuron (SMN1), encodes a ubiquitous protein, SMN1 deficiency preferentially affects spinal motor neurons (MNs), leaving the basis of this selective cell damage still unexplained. As neural stem cells (NSCs) are multipotent self-renewing cells that can differentiate into neurons, they represent an in vitro model for elucidating the pathogenetic mechanism of neurodegenerative diseases such as SMA. Here we characterize for the first time neural stem cells (NSCs) derived from embryonic spinal cords of a severe SMNΔ7 SMA mouse model. SMNΔ7 NSCs behave as their wild type (WT) counterparts, when we consider neurosphere formation ability and the expression levels of specific regional and self-renewal markers. However, they show a perturbed cell cycle phase distribution and an increased proliferation rate compared to wild typ...
Oncotarget, 2015
Glioblastoma multiforme (GBM) is the most common and deadliest primary brain tumor, driving patie... more Glioblastoma multiforme (GBM) is the most common and deadliest primary brain tumor, driving patients to death within 15 months after diagnosis (short term survivors, ST), with the exception of a small fraction of patients (long term survivors, LT) surviving longer than 36 months. Here we present deep sequencing data showing that peritumoral (P) areas differ from healthy white matter, but share with their respective frankly tumoral (C) samples, a number of mRNAs and microRNAs representative of extracellular matrix remodeling, TGFβ and signaling, of the involvement of cell types different from tumor cells but contributing to tumor growth, such as microglia or reactive astrocytes. Moreover, we provide evidence about RNAs differentially expressed in ST vs LT samples, suggesting the contribution of TGF-β signaling in this distinction too. We also show that the edited form of miR-376c-3p is reduced in C vs P samples and in ST tumors compared to LT ones. As a whole, our study provides new ...
Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, 2015
Background and aims: Epithelial-to-mesenchymal transition (EMT) and the reverse mesenchymal-to-ep... more Background and aims: Epithelial-to-mesenchymal transition (EMT) and the reverse mesenchymal-to-epithelial 23 transition (MET) are manifestations of cellular plasticity that imply a dynamic and profound gene expression 24 reprogramming. While a major epigenetic code controlling the coordinated regulation of a whole transcriptional 25 profile is guaranteed by DNA methylation, DNA methyltransferase (DNMT) activities in EMT/MET dynamics are 26 still largely unexplored. 27 Here, we investigated the molecular mechanisms directly linking HNF4α, the master effector of MET, to the 28 regulation of both de novo of DNMT 3A and 3B. 29 Methods: Correlation among EMT/MET markers, microRNA29 and DNMT3s expression was evaluated by 30 RT-qPCR, Western blotting and immunocytochemical analysis. Functional roles of microRNAs and DNMT3s 31 were tested by anti-miRs, microRNA precursors and chemical inhibitors. ChIP was utilized for investigating 32 HNF4α DNA binding activity. 33 Results: HNF4α silencing was sufficient to induce positive modulation of DNMT3B, in in vitro differentiated 34 hepatocytes as well as in vivo hepatocyte-specific Hnf4α knockout mice, and DNMT3A, in vitro, but not 35 DNMT1. In exploring the molecular mechanisms underlying these observations, evidence have been gathered 36 for (i) the inverse correlation between DNMT3 levels and the expression of their regulators miR-29a and miR-37 29b and (ii) the role of HNF4α as a direct regulator of miR-29a-b transcription. Notably, during TGFβ-induced 38 EMT, DNMT3s' pivotal function has been proved, thus suggesting the need for the repression of these DNMTs 39 in the maintenance of a differentiated phenotype. 40 Conclusions: HNF4α maintains hepatocyte identity by regulating miR-29a and -29b expression, which in turn 41 control epigenetic modifications by limiting DNMT3A and DNMT3B levels. 42
British journal of cancer, Jan 25, 2011
Combination of age at diagnosis, stage and MYCN amplification stratifies neuroblastoma into low-r... more Combination of age at diagnosis, stage and MYCN amplification stratifies neuroblastoma into low-risk and high-risk. We aimed to establish whether a microRNA (miRNA) signature could be associated with prognosis in both groups. Microarray expression profiling of human miRNAs and quantitative reverse-transcriptase PCR of selected miRNAs were performed on a preliminary cohort of 13 patients. Results were validated on an independent cohort of 214 patients. The relationship between miRNA expression and the overall or disease-free survival was analysed on the total cohort of 227 patients using the log-rank test and the multivariable Cox proportional hazard model. A total of 15 of 17 miRNAs that discriminated high-risk from low-risk neuroblastoma belonged to the imprinted human 14q32.31 miRNA cluster and two, miR-487b and miR-410, were significantly downregulated in the high-risk group. Multivariable analyses showed miR-487b expression as associated with overall survival and disease-free su...
International journal of immunopathology and pharmacology
Malignant gliomas, with an incidence of 5 cases per 100,000 population per year, represent the mo... more Malignant gliomas, with an incidence of 5 cases per 100,000 population per year, represent the most common primary brain tumour. They have an overall survival length of less than 2 years. Many different adjuvant therapies have been developed. Among them, Photodynamic Therapy (PDT), that is based on photochemical reactions between light and tumoral tissue selectively labelled with exogenous photosensitizing agents. Among photosensitizers, m-THPC (Temoporfin), seems to be the most promising one for the treatment of brain tumors, but, unfortunately, it causes problems of high skin photosensitivity. To by-pass this problem, we devised an intratumoral route of administration of this photosensitizer. The aim of this study is to investigate and compare the uptake of m-THPC in brain tumor and normal tissue after systemic and intratumoral administration of the drug. 30 female Wistar rats received m-THPC 12 days after C6 tumor implantation. Temoporfin was administered intratumorally in 24 rat...
Free radical research, 2007
Since it was suggested that cobalt chloride (CoCl(2)) could mimic the O(2) sensing role of mitoch... more Since it was suggested that cobalt chloride (CoCl(2)) could mimic the O(2) sensing role of mitochondria by increasing reactive oxygen species (ROS) generation during normoxia, we studied the correlation between CoCl(2)-generation of free radicals and the induction of a hypoxic cellular response in myogenic cell lines. In both L6C5 and C2C12 cell lines, exposure to CoCl(2) induced an increase of intracellular oxidants, the accumulation of HIF-1alpha protein, and the expression of vascular endothelial growth factor (VEGF) and/or iNOS genes. On the other hand, only ascorbic acid, but not trolox, was effective in lowering the CoCl(2) gene up-regulation. Neither the cytotoxicity nor the apoptosis induced by CoCl(2) in skeletal muscle cells were modified by culture supplementation with either ascorbic acid or trolox. Thus, CoCl(2) treatment of myogenic cell lines may represent a useful and convenient in vitro model to study gene modulation induced by hypoxia in skeletal muscle, although c...
Cancer research, Jan 15, 2001
Several reports have suggested that the mechanism of protection induced by antiidiotypic vaccinat... more Several reports have suggested that the mechanism of protection induced by antiidiotypic vaccination against low-grade lymphoproliferative disorders is likely to be antibody mediated. Here we test the hypothesis that DNA vaccination with the short peptide encompassing the complementary-determining region 3 hypervariable region of immunoglobulin heavy chain (VH-CDR3) may elicit a specific antibody immune response able to recognize the native antigens in the form required for therapy. As a test system, we used the VH-CDR3 sequences derived from two patients with non-Hodgkin's B lymphomas (PA, AS) and one patient with hairy cell leukemia (BA) to immunize outbred Swiss mice. This experimental model could mimic a clinical setting in which different patients present distinct HLA haplotypes. Individual tumor-specific VH-CDR3 sequences were amplified by a two-step procedure and directly cloned into multigenic plasmid vectors (pRC100 and derived) with and without mouse interleukin 2 (mIL...
Advances in experimental medicine and biology, 1998
Cell Biology International Reports, 1990
Vaccine, 2006
The high toll of death among first-week infants is due to infections occurring at the end of preg... more The high toll of death among first-week infants is due to infections occurring at the end of pregnancy, during birth or by breastfeeding. This problem significantly concerns industrialized countries also. To prevent the typical "first-week infections", a vaccine would be protective as early as at the birth. In utero DNA immunization has demonstrated the effectiveness in inducing specific immunity in newborns. We have already published results of a 2-year follow-up showing long-term safety, protective antibody titers at birth and long-term immune memory, following intramuscular in utero anti-HBV DNA immunization in 90-days pig fetuses.
Nephron, 1997
Indexes of myocardial ischemia and vasoconstrictive hormonal release were evaluated in order to i... more Indexes of myocardial ischemia and vasoconstrictive hormonal release were evaluated in order to investigate the difference between essential hypertension and hypertension during chronic renal failure. Arterial hypertension induces several cardiovascular alterations that reflect themselves either on the heart and/or on the coronary blood flow enhancing the cardiovascular risk. Since chronic renal failure can influence the neuroendocrine response, various mechanisms involved in hypertension during chronic renal failure are still unclear. High endothelin 1 (ET-1) levels have been found both in arterial hypertension and during chronic renal failure. Interestingly, either ET-1 or catecholamines seem also to be implied in the pathogenesis of myocardial ischemia. 20 hypertensive uremic and 20 essentially hypertensive patients underwent echocardiographic wall motion and wall thickening analysis performed at baseline and immediately after the end of exercise. Simultaneously, myocardial perfusion was evaluated by 99mTc-MIBI-SPECT. In addition, plasma norepinephrine and ET-1 concentrations were measured at baseline and at peak exercise. The segmental radionuclide analysis showed a greater ischemic degree in hypertensive uremic patients. Yet, we were able to identify one or more regions of the left ventricle in which both systolic thickening measurements and wall motion after exercise were impaired. After exercise, wall thickening impairment was correlated with both wall motion abnormalities (r = 0.72, p < 0.01) and MIBI ischemic grade (r = 0.82, p < 0.001). Basal and after-exercise plasmatic norepinephrine and endothelin levels were higher in hypertensive uremic than in essentially hypertensive patients. Moreover, there was a significant correlation between increments in norepinephrine concentration and MIBI perfusion defects, and between the increment in ET-1 concentration and both MIBI perfusion defects, or kinetic alterations assessed by wall motion as well as by wall thickening. This is the first cross-sectional study in which a higher degree of myocardial ischemia has been observed in hypertensive uremic patients combined with an enhanced plasma release of both norepinephrine and ET-1. This phenomenon may contribute to enhance the cardiovascular risk of these patients.
Nucleic Acids Research, 2011
MicroRNAs (miRNAs) are potent negative regulators of gene expression involved in all aspects of c... more MicroRNAs (miRNAs) are potent negative regulators of gene expression involved in all aspects of cell biology. They finely modulate virtually all physiological pathways in metazoans, and are deeply implicated in all main pathologies, among which cancer. Mir-221 and miR-222, two closely related miRNAs encoded in cluster from a genomic region on chromosome X, are strongly upregulated in several forms of human tumours. In this work, we report that the ectopic modulation of NF-kB modifies miR-221/222 expression in prostate carcinoma and glioblastoma cell lines, where we had previously shown their oncogenic activity. We identify two separate distal regions upstream of miR-221/222 promoter which are bound by the NF-kB subunit p65 and drive efficient transcription in luciferase reporter assays; consistently, the site-directed mutagenesis disrupting p65 binding sites or the ectopical inhibition of NF-kB activity significantly reduce luciferase activity. In the most distal enhancer region, we also define a binding site for c-Jun, and we show that the binding of this factor cooperates with that of p65, fully accounting for the observed upregulation of miR-221/222. Thus our work uncovers an additional mechanism through which NF-kB and c-Jun, two transcription factors deeply involved in cancer onset and progression, contribute to oncogenesis, by inducing miR-221/222 transcription.
Rivista europea per le scienze mediche e farmacologiche = European review for medical and pharmacological sciences = Revue européenne pour les sciences médicales et pharmacologiques
Cardiac function and morphology in chronic hemodialyzed patients are modified in consequence of b... more Cardiac function and morphology in chronic hemodialyzed patients are modified in consequence of both vascular and neurohormonal factors. In the present study we investigate on the role of prostacyclin (PGI2) vasodilator agent, during hemodialytic (HD) treatment. Twenty-four patients (13 males and 11 females; 9 hypertensive and 15 normotensive) aged 58.5 +/- 14.4 years were studied; 2.5 ml of venous blood were collected before (time 0) and 15&amp;#39;, 120&amp;#39;, and 240&amp;#39; of dialytic session. The PGI2 levels were measured in plasma, after extraction in ethyl acetate by RIA method, as levels of 6-Keto-PGF1 alpha, a stable metabolite. The results have shown as increase of PGI2 levels at 15&amp;#39; in hypertensive HD patients (HHD) from the begin of dialysis that increased until 240&amp;#39;. This phenomenon was more significant in hypertensive than in normotensive group (NHD) (p &amp;lt; 0.05 vs NHD). These preliminary data suggest that in HHD patients the role of PGI2 is more important than in NHD patients as regards the effects on regulation of circulatory tone. The increment of PGI2 levels could be in relation with the sympathetic activation occurred during hemodialytic treatment.
The EMBO Journal
The expression of the yeast L2 r-protein gene is controlled at the level of mRNA accumulation. Th... more The expression of the yeast L2 r-protein gene is controlled at the level of mRNA accumulation. The product of the gene appears to participate in this regulation by an autogenous feedback mechanism. This control does not operate at the level of transcription but instead affects L2 mRNA accumulation. This autogenous regulation of mRNA accumulation provides an interesting analogy to the autogenous translational regulation of r-proteins in Escherichia coli. Key words: autogenous regulation/ribosomal protein/RNA stability/yeast an autogenous feedback control in which the LI r-protein controls its synthesis by regulating the efficiency of splicing of its own pre-mRNA .
Recenti progressi in medicina
Atrial natriuretic factor (ANF) is a potent natriuretic and vasoactive (vasorelaxant) peptide loc... more Atrial natriuretic factor (ANF) is a potent natriuretic and vasoactive (vasorelaxant) peptide localized in the secretory-like atrial specific granules. The main peptide in this storage granules is the 126 amino acid proatrial natriuretic peptide, but the principal circulating form in human plasma is the 28 amino acid, alpha-human natriuretic peptide. Animal and in vitro studies have suggested that ANF modulates autonomic circulatory control, probably with a dose-dependent mechanism. Moreover, recent human studies have resulted contradictory. In particular, it is still unclear if high circulating levels of ANF, which are present in congestive heart diseases constantly, may be correlated with sympathetic nervous system activity in man. Previously we have shown that in congestive diseases there is a relation between ANF and catecholamine secretion. From these basis, the aim of this study was to investigate on the pathophysiological relations between atrial natriuretic factor (ANF) release and adrenergic activation in patients with obstructive hypertrophic cardiomyopathy (n = 6) and non obstructive hypertrophic cardiomyopathy (n = 4). Sympathetic activation in physiologic way was induced by cycloergometer sub-maximal exercise. Then specimens of venous blood were achieved for plasma determination of ANF and catecholamines pre- and post-exercise. Results have shown that in obstructive hypertrophic cardiomyopathy patients basal levels of ANF and catecholamines were higher than levels of these parameters in non obstructive hypertrophic cardiomyopathy patients.(ABSTRACT TRUNCATED AT 250 WORDS)
Recenti progressi in medicina
Endothelin-1 (ET-1) is an endothelium-derived vasoconstrictor peptide isolated from the culture s... more Endothelin-1 (ET-1) is an endothelium-derived vasoconstrictor peptide isolated from the culture supernatant of porcine aortic endothelial cells. This 21 amino-acid residue peptide has potent vasoconstrictive properties in vitro and in vivo. ET-1 action involves phosphatidylinositol turnover, calcium mobilization and protein kinase C activation. Endothelial cells have distinct receptors for different operating through hydrosoluble hormones. The aim of this study was to investigate on a possible role of angiotensin II (ANG II) to modulate the release ET-1 from human endothelial cells in vitro. These data revealed a time- and a dose-dependent increase of ET-1 production in response to ANG II. This mechanism may have important pathophysiological implications in vivo. In fact, a double-mechanism of secretion of ET-1 from endothelial cells could exist: one active in a physiological condition and an other in response to a vasoconstrictor stimuli (as well as ANG II). Furthermore, these results may suggest an additional favourable effect of ACE-inhibition in human hypertension therapy.
Virology, Jan 9, 2015
Adenosine deaminase acting on RNA1 (ADAR1) was previously reported to affect HIV-1 replication. W... more Adenosine deaminase acting on RNA1 (ADAR1) was previously reported to affect HIV-1 replication. We report data showing that ADAR1 interacts with the HIV-1 p55 Gag protein, the major structural protein of the immature virus capsid. Furthermore, we found that the endogenous ADAR1 is incorporated into virions purified from the supernatant of primary HIV-1-infected CD4(+) T lymphocytes. Additional experiments demonstrated that the expression of the p55 Gag protein is sufficient for ADAR1 incorporation into virus-like particles (VLPs). Overall, our data originally support the evidence that ADAR1 can be part of the cell protein array uploaded in HIV-1 particles.
International journal of molecular sciences, 2015
Spinal muscular atrophy (SMA) is an inherited neuromuscular disorder and the leading genetic caus... more Spinal muscular atrophy (SMA) is an inherited neuromuscular disorder and the leading genetic cause of death in infants. Despite the disease-causing gene, survival motor neuron (SMN1), encodes a ubiquitous protein, SMN1 deficiency preferentially affects spinal motor neurons (MNs), leaving the basis of this selective cell damage still unexplained. As neural stem cells (NSCs) are multipotent self-renewing cells that can differentiate into neurons, they represent an in vitro model for elucidating the pathogenetic mechanism of neurodegenerative diseases such as SMA. Here we characterize for the first time neural stem cells (NSCs) derived from embryonic spinal cords of a severe SMNΔ7 SMA mouse model. SMNΔ7 NSCs behave as their wild type (WT) counterparts, when we consider neurosphere formation ability and the expression levels of specific regional and self-renewal markers. However, they show a perturbed cell cycle phase distribution and an increased proliferation rate compared to wild typ...
Oncotarget, 2015
Glioblastoma multiforme (GBM) is the most common and deadliest primary brain tumor, driving patie... more Glioblastoma multiforme (GBM) is the most common and deadliest primary brain tumor, driving patients to death within 15 months after diagnosis (short term survivors, ST), with the exception of a small fraction of patients (long term survivors, LT) surviving longer than 36 months. Here we present deep sequencing data showing that peritumoral (P) areas differ from healthy white matter, but share with their respective frankly tumoral (C) samples, a number of mRNAs and microRNAs representative of extracellular matrix remodeling, TGFβ and signaling, of the involvement of cell types different from tumor cells but contributing to tumor growth, such as microglia or reactive astrocytes. Moreover, we provide evidence about RNAs differentially expressed in ST vs LT samples, suggesting the contribution of TGF-β signaling in this distinction too. We also show that the edited form of miR-376c-3p is reduced in C vs P samples and in ST tumors compared to LT ones. As a whole, our study provides new ...
Biochimica et Biophysica Acta (BBA) - Gene Regulatory Mechanisms, 2015
Background and aims: Epithelial-to-mesenchymal transition (EMT) and the reverse mesenchymal-to-ep... more Background and aims: Epithelial-to-mesenchymal transition (EMT) and the reverse mesenchymal-to-epithelial 23 transition (MET) are manifestations of cellular plasticity that imply a dynamic and profound gene expression 24 reprogramming. While a major epigenetic code controlling the coordinated regulation of a whole transcriptional 25 profile is guaranteed by DNA methylation, DNA methyltransferase (DNMT) activities in EMT/MET dynamics are 26 still largely unexplored. 27 Here, we investigated the molecular mechanisms directly linking HNF4α, the master effector of MET, to the 28 regulation of both de novo of DNMT 3A and 3B. 29 Methods: Correlation among EMT/MET markers, microRNA29 and DNMT3s expression was evaluated by 30 RT-qPCR, Western blotting and immunocytochemical analysis. Functional roles of microRNAs and DNMT3s 31 were tested by anti-miRs, microRNA precursors and chemical inhibitors. ChIP was utilized for investigating 32 HNF4α DNA binding activity. 33 Results: HNF4α silencing was sufficient to induce positive modulation of DNMT3B, in in vitro differentiated 34 hepatocytes as well as in vivo hepatocyte-specific Hnf4α knockout mice, and DNMT3A, in vitro, but not 35 DNMT1. In exploring the molecular mechanisms underlying these observations, evidence have been gathered 36 for (i) the inverse correlation between DNMT3 levels and the expression of their regulators miR-29a and miR-37 29b and (ii) the role of HNF4α as a direct regulator of miR-29a-b transcription. Notably, during TGFβ-induced 38 EMT, DNMT3s' pivotal function has been proved, thus suggesting the need for the repression of these DNMTs 39 in the maintenance of a differentiated phenotype. 40 Conclusions: HNF4α maintains hepatocyte identity by regulating miR-29a and -29b expression, which in turn 41 control epigenetic modifications by limiting DNMT3A and DNMT3B levels. 42
British journal of cancer, Jan 25, 2011
Combination of age at diagnosis, stage and MYCN amplification stratifies neuroblastoma into low-r... more Combination of age at diagnosis, stage and MYCN amplification stratifies neuroblastoma into low-risk and high-risk. We aimed to establish whether a microRNA (miRNA) signature could be associated with prognosis in both groups. Microarray expression profiling of human miRNAs and quantitative reverse-transcriptase PCR of selected miRNAs were performed on a preliminary cohort of 13 patients. Results were validated on an independent cohort of 214 patients. The relationship between miRNA expression and the overall or disease-free survival was analysed on the total cohort of 227 patients using the log-rank test and the multivariable Cox proportional hazard model. A total of 15 of 17 miRNAs that discriminated high-risk from low-risk neuroblastoma belonged to the imprinted human 14q32.31 miRNA cluster and two, miR-487b and miR-410, were significantly downregulated in the high-risk group. Multivariable analyses showed miR-487b expression as associated with overall survival and disease-free su...
International journal of immunopathology and pharmacology
Malignant gliomas, with an incidence of 5 cases per 100,000 population per year, represent the mo... more Malignant gliomas, with an incidence of 5 cases per 100,000 population per year, represent the most common primary brain tumour. They have an overall survival length of less than 2 years. Many different adjuvant therapies have been developed. Among them, Photodynamic Therapy (PDT), that is based on photochemical reactions between light and tumoral tissue selectively labelled with exogenous photosensitizing agents. Among photosensitizers, m-THPC (Temoporfin), seems to be the most promising one for the treatment of brain tumors, but, unfortunately, it causes problems of high skin photosensitivity. To by-pass this problem, we devised an intratumoral route of administration of this photosensitizer. The aim of this study is to investigate and compare the uptake of m-THPC in brain tumor and normal tissue after systemic and intratumoral administration of the drug. 30 female Wistar rats received m-THPC 12 days after C6 tumor implantation. Temoporfin was administered intratumorally in 24 rat...
Free radical research, 2007
Since it was suggested that cobalt chloride (CoCl(2)) could mimic the O(2) sensing role of mitoch... more Since it was suggested that cobalt chloride (CoCl(2)) could mimic the O(2) sensing role of mitochondria by increasing reactive oxygen species (ROS) generation during normoxia, we studied the correlation between CoCl(2)-generation of free radicals and the induction of a hypoxic cellular response in myogenic cell lines. In both L6C5 and C2C12 cell lines, exposure to CoCl(2) induced an increase of intracellular oxidants, the accumulation of HIF-1alpha protein, and the expression of vascular endothelial growth factor (VEGF) and/or iNOS genes. On the other hand, only ascorbic acid, but not trolox, was effective in lowering the CoCl(2) gene up-regulation. Neither the cytotoxicity nor the apoptosis induced by CoCl(2) in skeletal muscle cells were modified by culture supplementation with either ascorbic acid or trolox. Thus, CoCl(2) treatment of myogenic cell lines may represent a useful and convenient in vitro model to study gene modulation induced by hypoxia in skeletal muscle, although c...
Cancer research, Jan 15, 2001
Several reports have suggested that the mechanism of protection induced by antiidiotypic vaccinat... more Several reports have suggested that the mechanism of protection induced by antiidiotypic vaccination against low-grade lymphoproliferative disorders is likely to be antibody mediated. Here we test the hypothesis that DNA vaccination with the short peptide encompassing the complementary-determining region 3 hypervariable region of immunoglobulin heavy chain (VH-CDR3) may elicit a specific antibody immune response able to recognize the native antigens in the form required for therapy. As a test system, we used the VH-CDR3 sequences derived from two patients with non-Hodgkin's B lymphomas (PA, AS) and one patient with hairy cell leukemia (BA) to immunize outbred Swiss mice. This experimental model could mimic a clinical setting in which different patients present distinct HLA haplotypes. Individual tumor-specific VH-CDR3 sequences were amplified by a two-step procedure and directly cloned into multigenic plasmid vectors (pRC100 and derived) with and without mouse interleukin 2 (mIL...
Advances in experimental medicine and biology, 1998
Cell Biology International Reports, 1990
Vaccine, 2006
The high toll of death among first-week infants is due to infections occurring at the end of preg... more The high toll of death among first-week infants is due to infections occurring at the end of pregnancy, during birth or by breastfeeding. This problem significantly concerns industrialized countries also. To prevent the typical "first-week infections", a vaccine would be protective as early as at the birth. In utero DNA immunization has demonstrated the effectiveness in inducing specific immunity in newborns. We have already published results of a 2-year follow-up showing long-term safety, protective antibody titers at birth and long-term immune memory, following intramuscular in utero anti-HBV DNA immunization in 90-days pig fetuses.
Nephron, 1997
Indexes of myocardial ischemia and vasoconstrictive hormonal release were evaluated in order to i... more Indexes of myocardial ischemia and vasoconstrictive hormonal release were evaluated in order to investigate the difference between essential hypertension and hypertension during chronic renal failure. Arterial hypertension induces several cardiovascular alterations that reflect themselves either on the heart and/or on the coronary blood flow enhancing the cardiovascular risk. Since chronic renal failure can influence the neuroendocrine response, various mechanisms involved in hypertension during chronic renal failure are still unclear. High endothelin 1 (ET-1) levels have been found both in arterial hypertension and during chronic renal failure. Interestingly, either ET-1 or catecholamines seem also to be implied in the pathogenesis of myocardial ischemia. 20 hypertensive uremic and 20 essentially hypertensive patients underwent echocardiographic wall motion and wall thickening analysis performed at baseline and immediately after the end of exercise. Simultaneously, myocardial perfusion was evaluated by 99mTc-MIBI-SPECT. In addition, plasma norepinephrine and ET-1 concentrations were measured at baseline and at peak exercise. The segmental radionuclide analysis showed a greater ischemic degree in hypertensive uremic patients. Yet, we were able to identify one or more regions of the left ventricle in which both systolic thickening measurements and wall motion after exercise were impaired. After exercise, wall thickening impairment was correlated with both wall motion abnormalities (r = 0.72, p < 0.01) and MIBI ischemic grade (r = 0.82, p < 0.001). Basal and after-exercise plasmatic norepinephrine and endothelin levels were higher in hypertensive uremic than in essentially hypertensive patients. Moreover, there was a significant correlation between increments in norepinephrine concentration and MIBI perfusion defects, and between the increment in ET-1 concentration and both MIBI perfusion defects, or kinetic alterations assessed by wall motion as well as by wall thickening. This is the first cross-sectional study in which a higher degree of myocardial ischemia has been observed in hypertensive uremic patients combined with an enhanced plasma release of both norepinephrine and ET-1. This phenomenon may contribute to enhance the cardiovascular risk of these patients.
Nucleic Acids Research, 2011
MicroRNAs (miRNAs) are potent negative regulators of gene expression involved in all aspects of c... more MicroRNAs (miRNAs) are potent negative regulators of gene expression involved in all aspects of cell biology. They finely modulate virtually all physiological pathways in metazoans, and are deeply implicated in all main pathologies, among which cancer. Mir-221 and miR-222, two closely related miRNAs encoded in cluster from a genomic region on chromosome X, are strongly upregulated in several forms of human tumours. In this work, we report that the ectopic modulation of NF-kB modifies miR-221/222 expression in prostate carcinoma and glioblastoma cell lines, where we had previously shown their oncogenic activity. We identify two separate distal regions upstream of miR-221/222 promoter which are bound by the NF-kB subunit p65 and drive efficient transcription in luciferase reporter assays; consistently, the site-directed mutagenesis disrupting p65 binding sites or the ectopical inhibition of NF-kB activity significantly reduce luciferase activity. In the most distal enhancer region, we also define a binding site for c-Jun, and we show that the binding of this factor cooperates with that of p65, fully accounting for the observed upregulation of miR-221/222. Thus our work uncovers an additional mechanism through which NF-kB and c-Jun, two transcription factors deeply involved in cancer onset and progression, contribute to oncogenesis, by inducing miR-221/222 transcription.