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I Ceccherini

Publications and source records attributed to I Ceccherini.

At least 19 recordsLinked to original sources
PubMed2006

Clinical and genetic characterization of Italian patients affected by CINCA syndrome.

OBJECTIVE: We report the experience of the Italian Registry of patients affected by chronic infantile neurological, cutaneous, articular (CINCA) syndrome. The clinical and genetic features of 12 unrelated Italian patients with CINCA syndrome are described, focusing on the possible influence of the presence of CIAS1/cryopyrin mutations on the phenotype of the disease and on its prognosis. METHODS: The clinical features of 12 Italian CINCA patients were evaluated. Genomic DNA of the patients was sequenced using specific primers for CIAS1 and ASC genes. RESULTS: Our patients shared typical CINCA characteristics and, sometimes, remarkable perinatal events, peculiar of CIAS1-mutated patients. Seven patients carried CIAS1 missense mutation, localized within the nucleotide binding domain of cryopyrin. Four previously described mutations and three new heterozygous CIAS1 missense mutations were identified. ASC gene, encoding for a direct interactor of cryopyrin, was not mutated in Italian CINCA patients. Finally, we reported the efficacy and safety of anti-IL1 therapy (Anakinra) in seven patients with a particularly severe CINCA phenotype. CONCLUSION: Despite some common signs-used as syndrome hallmarks-we observed a high variability in symptoms, genetic results and outcomes in Italian CINCA patients. In contrast with other authors, we cannot find out any correlation between mutations in CIAS1 and CINCA severity, but we underlined the concomitance of perinatal events and mental retardation only in CIAS1 mutated subjects. Finally, we confirmed the efficacy of Anakinra treatment, both in CIAS1-mutated and non-mutated patients.

Adolescent↗
PubMed2006

Haplotypes of the human RET proto-oncogene associated with Hirschsprung disease in the Italian population derive from a single ancestral combination of alleles.

The RET proto-oncogene is the major gene involved in the complex genetics of Hirschsprung disease (HSCR), or aganglionic megacolon, showing causative loss-of-function mutations in 15-30% of the sporadic cases. Several RET polymorphisms and haplotypes have been described in association with the disease, suggesting a role for this gene in HSCR predisposition, also in the absence of mutations in the coding region. Finally, the presence of a functional variant in intron 1 has repeatedly been proposed to explain such findings. Here we report a case-control study conducted on 97 Italian HSCR sporadic patients and 85 population matched controls, using 13 RET polymorphisms distributed throughout the gene, from the basal promoter to the 3'UTR. Linkage disequilibrium and haplotype analyses have shown increased recombination between the 5' and 3' portions of the gene and an over-representation, in the cases studied, of two haplotypes sharing a common allelic combination that extends from the promoter up to intron 5. We propose that these two disease-associated haplotypes derive from a single founding locus, extending up to intron 19 and successively rearranged in correspondence with a high recombination rate region located between the proximal and distal portions of the gene. Our results suggests the possibility that a common HSCR predisposing variant, in linkage disequilibrium with such haplotypes, is located further downstream than the previously suggested interval encompassing intron 1.

Alleles↗
PubMed2004

Replication pattern of the pericentromeric region of chromosome 10q and expression of the RET protooncogene.

Regulation of the RET gene is highly specific during embryo development and is strictly tissue-specific. Control of transcription depends on mechanisms influenced by epigenetic processes, in particular, histone acetylation at regions flanking the 5' end of the gene. Since the RET gene is mapped in the pericentromeric region of the human chromosome 10, the implication of epigenetic processes is even more striking and worth to be investigated in an extended chromosomal tract. One experimental approach to study the chromatin status in relationship with gene transcription is to assess the replication timing, which we did by using fluorescent in situ hybridization in cells expressing or not expressing the RET gene. By using probes spanning a 700-kb genomic region from the RET locus toward the centromere, we found a relationship between RET expression and early replication. Different patterns were observed between cells naturally expressing RET and cells induced to expression of RET by treatment with sodium butyrate, an inhibitor of histone deacetylases. Three-dimensional analysis of the nuclear localization of fluorescent signals by confocal microscopy showed difference of localization between the RET probe and a probe for a housekeeping gene, G3PDH, located at 12p13.3, in cells that do not express RET, in accordance with previous data for other genes and chromosomal regions. However, RET-expressing cells showed a localization of signals which was not consistent with that expected for expressed genes.

Butyrates↗
PubMed2003

Rescue of human RET gene expression by sodium butyrate: a novel powerful tool for molecular studies in Hirschsprung disease.

BACKGROUND: The RET gene encodes a tyrosine kinase receptor involved in different human neurocristopathies, such as specific neuroendocrine tumours and Hirschsprung disease (HSCR). Gene expression is developmentally regulated and the RET transcript is undetectable in most adult cells, including lymphocytes. The impossibility of performing functional studies on RET mRNA has to date limited the detection and characterisation of an indefinite proportion of gene anomalies that cannot be identified by conventional DNA genomic screening in HSCR cases. AIMS: Development of a protocol suitable to activate RET expression in RET negative cell lines and therefore to investigate directly RET mRNA, extending the conventional gene mutation analysis to detection of splicing anomalies and impaired expression of the RET gene. METHODS: The effect of sodium butyrate (NaB), a histone deacetylase inhibitor, on rescuing RET expression was tested by one round of reverse transcription- polymerase chain reaction from total RNA of treated lymphoblasts from both HSCR patients and control individuals. RESULTS: Analysis of RET expression was possible by NaB treatment of RET negative cells, such as lymphoblasts. This treatment allowed us to detect impaired RET expression as well as a splicing defect in two HSCR patients previously believed to be devoid of any gene abnormality. CONCLUSIONS: The full application of the proposed protocol in most of the unexplained HSCR cases will allow us to establish the precise role of RET not only in causing but also in predisposing to HSCR pathogenesis.

Alternative Splicing↗
PubMed2003

The Ret proto-oncogene in the WAG/Rij rat strain: an animal model for inherited C-cell carcinoma?

WAG/Rij rat strain has been suggested as an animal model for the study of inherited human medullary thyroid carcinoma (MTC), due to its high incidence of spontaneous C-cell thyroid tumours. Although the role of the Ret proto-oncogene mutations, as responsible for human MTC, is well established, nothing has been published concerning this putative animal model. Based upon the previously reported rat Ret sequence, exons 10, 11, 13, 14, 15, and 16, known to carry activating mutations in humans, have been analysed in the WAG/Rij rat by PCR, single strand conformational polymorphism (SSCP) and direct sequencing. Neither the germline nor MTC samples showed any Ret sequence difference in the exons when analysed in comparison to a non-MTC-susceptible rat strain. Our results indicate that Ret exons relevant in humans are not involved in WAG/Rij rat MTC, as expected, and this questions the validity of this strain as a model for the human disease, and suggests there must be additional mechanisms for the genesis and progression of rat MTC.

Animals↗
PubMed2002

Diagnostic and therapeutic approach to multiple endocrine neoplasia type 2B in pediatric patients.

Multiple endocrine neoplasia (MEN) 2B is a hereditary syndrome including medullary thyroid carcinoma (MTC), pheochromocytoma, gastrointestinal (GI) disorders, marfanoid facies, and multiple ganglioneuromas. MTC is the major cause of mortality, and often appears during the 1st decade of life. RET proto-oncogene mutations are responsible for MEN 2B. Other RET mutations cause MEN 2A syndrome, familial MTC, or Hirschsprung's disease. We studied three MEN 2B patients with the aim of delineating the best diagnostic and therapeutic protocol. The gold standards for diagnosis are histochemical study of the rectal mucosa and molecular analysis of RET, which in familial cases detects MEN 2B at a preclinical stage so that early total prophylactic thyroidectomy can be performed. In non-familial cases, the diagnosis can be suggested by the presence of GI symptoms, ganglioneuromas, and/or the typical facies. The intestinal innervation pattern, analyzed with the acetylcholinesterase technique, is pathognomonic for MEN 2B. In our protocol a rectal biopsy is, therefore, the first measure. The surgical treatment of MEN 2B is total thyroidectomy with cervical lymphadenectomy of the central compartment of the neck. When possible, this intervention should be performed prophylactically before 1 year of age.

Adolescent↗
PubMed2002

Specific haplotypes of the RET proto-oncogene are over-represented in patients with sporadic papillary thyroid carcinoma.

BACKGROUND: Papillary thyroid carcinoma (PTC), which may be sporadic (95%) or familial (5%), has a prevalence adjusted for age in the general population of 1:100 000. Somatic rearrangements of the RET proto-oncogene are present in up to 66% of sporadic tumours, while they are rarely found in familial cases. PURPOSE: In order to determine if some variants of this gene, or a combination of them, might predispose to PTC, we looked for an association of RET haplotype(s) in PTC cases and in controls from four countries matched for sex, age, and population. METHODS: Four single nucleotide polymorphisms (SNPs) across the RET coding sequence were typed and haplotype frequencies were estimated. Genotype and haplotype distributions were compared among these cases and controls. RESULTS: Ten haplotypes were observed, the seven most frequent of which have been previously described in sporadic Hirschsprung patients and controls. The single locus analyses suggested association of exon 2 and exon 13 SNPs with sporadic PTC. The haplotype analysis showed over-representation of one haplotype in French and Italian sporadic PTC, whereas a different haplotype was significantly under-represented in French familial PTC. CONCLUSIONS: Our data suggest that some variants of RET and some specific haplotypes may act as low penetrance alleles in the predisposition to PTC.

Age Factors↗
PubMed2001

The sensitivity of activated Cys Ret mutants to glial cell line-derived neurotrophic factor is mandatory to rescue neuroectodermic cells from apoptosis.

Hirschsprung's disease (HSCR), a frequent developmental defect of the enteric nervous system is due to loss-of-function mutations of RET, a receptor tyrosine kinase essential for the mediation of glial cell-derived neurotrophic factor (GDNF)-induced cell survival. Instead, gain-of-function Cys mutations (e.g., Cys(609), Cys(620), and Cys(634)) of the same gene are responsible for thyroid carcinoma (MEN2A/familial medullary thyroid carcinoma) by causing a covalent Ret dimerization, leading to ligand-independent activation of its tyrosine kinase. In this context, the association of Cys(609)- or Cys(620)-activating mutations with HSCR is still an unresolved paradox. To address this issue, we have compared these two mutants with the Cys(634) Ret variant, which has never been associated with HSCR, for their ability to rescue neuroectodermic cells (SK-N-MC cells) from apoptosis. We show here that despite their constitutively activated kinase, the mere expression of these three mutants does not allow cell rescue. Instead, we demonstrate that like the wild-type Ret, the Cys(634) Ret variant can trigger antiapoptotic pathways only in response to GDNF. In contrast, Cys(609) or Cys(620) mutations, which impair the terminal Ret glycosylation required for its insertion at the plasma membrane, abrogate GDNF-induced cell rescue. Taken together, these data support the idea that sensitivity to GDNF is the mandatory condition, even for constitutively activated Ret mutants, to rescue neuroectodermic cells from apoptosis. These findings may help clarify how a gain-of-function mutation can be associated with a developmental defect.

Animals↗
PubMed2001

Genomic organisation of the mouse Ret proto-oncogene.

The RET proto-oncogene is involved in the development of both kidney and neural crests derived tissues. RET deleterious mutations cause hereditary neuroendocrine tumours and congenital intestinal aganglionosis. Ongoing efforts aimed at elucidating the function of this gene include expression studies in different species and in transgenic mice. As first step in the study of Ret expression in mouse, we obtained the mouse Ret genomic structure. Intron-exon boundaries were determined and sequenced, all introns but the first one were amplified and cloned, and exons positioned in a restriction map. Mouse and human genes comparison indicates a highly conserved genomic organisation, except for exon 21 which is not conserved in mouse. A region extending 386 bp 5' to the first exon was sequenced and compared with its human counterpart. Some features, reported for the human promoter, like the absence of TATA or CAAT boxes and a high GC content, are conserved.

Animals↗
PubMed2000

Incidence of RET mutations in patients with Hirschsprung's disease.

BACKGROUND: RET mutations have been reported variously to affect 7% to 41% of Hirschsprung's disease (HSCR) patients depending on familial or sporadic occurrence of the disease, length of aganglionosis and possible association with other disease phenotypes. The authors report a study of the incidence of RET mutations in unselected HSCR patients from two regional centers and correlate their genotypes and phenotypes. METHODS: The records of HSCR patients treated in 2 regional centers with a combined population of 5 million were reviewed, and blood samples were obtained from 57 patients. During the same period, 39 patients with similar demographic data refused or provided insufficient blood for study. DNA was extracted, and the 21 exons of the RET protooncogene were screened for mutations using denaturing gradient gel electrophoresis (DGGE). RESULTS: Of 57 patients, 48 were sporadic, and 9 were familial. Lengths of aganglionosis were total colonic, 4; long, 11; short, 39; ultrashort, 1; unclassified, 2. Associated anomalies were present in 20. Causative mutations were identified in 4 (7%): missense or "silent" in 3 (exons 5, 11, 13) and deletion in 1. The silent mutation of exon 11 recently has been shown to have effects on correct RET mRNA splicing. One mutation occurred in total colonic aganglionosis (25%), 1 in long segment dysganglionosis (9%), and 2 in short segment aganglionosis (5%). Surprisingly, all these mutations occurred in sporadic cases (10%). Five patients (9%) had rare polymorphic alleles at exon 14 (n = 1) and exon 18 (n = 4). Fifty patients (88%) showed common polymorphic alleles (sequence variants) in 1 or more exons (> 4, n = 5). CONCLUSIONS: RET mutation as a primary cause for Hirschsprung's disease in the general surgical population is less frequent than previously thought. This observation is compatible with the hypothesis that HSCR could be a polygenic disease caused by additive subclinical effects of more than one gene, including RET.

Drosophila Proteins↗
PubMed2000

A single-nucleotide polymorphic variant of the RET proto-oncogene is underrepresented in sporadic Hirschsprung disease.

Hirschsprung disease (HSCR) is an inherited disorder characterised by absence of intrinsic ganglion cells in the distal gastrointestinal tract. Different susceptibility genes, involved in either the Ret-tyrosine kinase or the endothelin signalling pathways, contribute to HSCR phenotype. Interestingly, alterations of these genes are detected in only 30-50% of all HSCR patients, suggesting the involvement of modifier genes and/or additional genetic or environmental risk factors. In complex disorders common polymorphic variants can be associated with the disease phenotype, thus modifying the risk of recurrence. To investigate whether sequence variants of the RET proto-oncogene may be associated with the development of the HSCR phenotype, we analysed 92 Italian patients for the 2508C > T synonymous substitution in exon 14 (S836S) finding that the T allele is clearly less frequent than in control individuals (Fisher exact test P = 0.0002). On the other hand, this RET variant allele is overrepresented in patients affected with medullary thyroid carcinoma. Assuming a direct effect of this single-nucleotide polymorphism in predisposing to RET associated pathologies, we have performed functional tests which excluded any possible involvement of the C and T alleles in DNA-protein binding, transcript stability and RNA splicing and editing.

Alleles↗
PubMed2000

Pathogenesis of Hirschsprung's disease.

Hirschsprung's disease is an inherited disorder showing incomplete penetrance and variable expressivity. Genetic mapping and mutation screening of candidate genes, together with the study of several natural and knockout animal models, clearly have shown the involvement of several different genes in the pathogenesis of Hirschsprung's disease. Among these genes, the RET proto-oncogene accounts for the highest proportion of both familial and sporadic cases, with a wide range of mutations scattered along its entire coding region. The low detection rate of RET mutations in Hirschsprung patients also led to different hypotheses, such as the existence of additional Hirschsprung genes. Different animal and human genetic studies have identified 6 Hirschsprung genes: RET proto-oncogene (RET), endothelin 3 (EDN3), endothelin B receptor gene (EDNRB), glial-cell-line-derived neurotrophic factor (GDNF), endothelin converting enzyme (ECE1), gene encoding the Sry-related transcription factor SOX10 (SOX10). Microenvironmental factors also can play a role in the pathogenesis of aganglionosis. The developmental process of the crest-derived progenitor cells is sensitive to the level of different molecules. The expression deficit of different factors (GDNF, NTN) in the hindgut, in the absence of genetic mutations, could determine a missed activation of the receptor system, causing enteric neuroblast migration arrest.

Child↗