Valutazione del background genetico di una coorte di individui dislessici mediante l'utilizzo di tecnologie ad alta processivit\ue0

Il progresso tecnologico e la riduzione dei costi ha significativamente contribuito all'identificazioni delle cause genetiche di diverse malattie genetiche multifattoriali. In questo lavoro di dottorato riporto i risultati di un analisi genetica di una coorte di ragazzi dislessici utilizzando delle tecnologie ad alta processivit\ue0 come Next Generation Sequencing e SNP-array ad alta densit\ue0. La coorte in studio consiste di 49 soggetti con dislessia e 52 soggetti con dislessia e altre Disabilit\ue0 Specifiche di Apprendimento (disortografia, disgrafia, discalculia). Tutti i campioni sono stati sequenziati utilizzando la piattaforma Ion Torrent, focalizzandosi sulle regioni codificanti e sulle giunzioni esone-introne, in 12 geni candidati (CMIP, CNTNAP2, CYP19A1, DCDC2, DIP2A, DYX1C1, GCFC2, KIAA0319, KIAA0319L, MRPL19, ROBO1, S100B), focalizzandosi sulle varianti non descritte in letteratura e in quelle rare (MAF G). Inoltre, diverse CNV sono state identificate che sovrappongono dei geni associati a problemi con il linguaggio, ma nessuna delle CNV con i geni riportati sopra. Infine, una copia di fratelli sono portatori di diverse duplicazioni localizzate nella regione 16p13.11, una regione di suscettibilit\ue0 ai disordini di neurosviluppo. Il presente lavoro arrichisse la nostra conoscenza sul background genetico della coorte in studio. Nello stesso momento i risultati ottenuti devono essere ulteriormente analizzati per poter attribuire un ruolo a quanto possibile certo sul loro contributo all'insorgenza della dislessia.Technological improvements and continued cost reduction have significantly contributed to the progress of identifying the genetic causes of complex traits. Here we report the results of a genetic screening on a dyslexia cohort combining targeted next generation sequencing and high density SNP array. The study cohort consists of 49 subjects with dyslexia and 52 subjects with dyslexia and other specific learning disabilities (dysorthographia, dysgraphia, dyscalculia). All samples were sequenced on Ion Torrent platform, targeting the coding regions and their exone-intron boundaries of 12 candidate dyslexia genes (CMIP, CNTNAP2, CYP19A1, DCDC2, DIP2A, DYX1C1, GCFC2, KIAA0319, KIAA0319L, MRPL19, ROBO1, S100B), with focus on novel and rare variants. A subset of 54 samples was further analyzed, genotyping over 1.7 M markers (Multi Ethnic Global Array design, Ilumina), for copy number variation (CNV) discovery and characterization according to the literature. For this purpose, high confidence CNVs were obtained using the cnvPartition and the PennCNV calling algorithms. We report a total of 12 pathogenic predicted variants, among which two novel deleterious events (DIP2A:p.G1387* and KIAA0319:p.V774Afs*37) and a known rare splicing variant (GCFC2:c.266-2A>G). Moreover, several copy number variants were identified, overlapping some language related genes, but not any of the above sequenced genes. Finally, a sibling pair was found to harbor duplications in the chromosome band 16p13.11, a susceptibility region for several neurodevelopmental disorders. The present study enriches our knowledge about the genetic background in a dyslexia cohort. At the same time our findings emphasize the need for further research to attribute causative roles of these events for cohort phenotypes

Developmental dyslexia and its complex genetic puzzle

Genetics suggest that DD is an additive or interactive effect of multiple genetic and environmental risk factors. In 2014 an interdisciplinary collaboration between the University of Trieste and the Institute for Maternal and Child Health IRCCS \u201cBurlo Garofolo\u201d of Trieste started, with the aim of performing a genetic study in Italian families with dyslexia

Copy number variation, gene expression and histological localization of human beta-defensin 2 in patients with adeno-tonsillar hypertrophy

Both bacterial infections and innate oral immunity response participate in development of adeno-tonsillar hypertrophy (ATH). ATH can lead to obstructive sleep apnea. We investigated the beta-defensin 2 (hBD-2) encoding gene, DEFB4, by analyzing the copy number variations (CNVs) of the defensin gene cluster in patients with ATH and by correlating CNV with DEFB4 gene expression. We enrolled 79 patients with ATH, 21 of whom presented with only adenoid hypertrophy, while 58 exhibited hypertrophy of both adenoid and tonsil. CNVs of the defensin gene cluster, DEFB4 mRNA, and hBD-2 protein expression were assessed. Also, beta-defensin 2 was localized histologically using immunohistochemistry. The distribution of defensin gene cluster CNV was similar among the 79 subjects. DEFB4 expression analysis exhibited considerable inter-individual variability, but with neither specific differences among subjects nor correlation with the CNV number. Immunohistochemistry enabled localization of hBD-2 in the tonsil and adenoid epithelium. No differences in localization between the two ATH presentations were found. Inducible antimicrobial defensin peptides exhibited great inter-individual variability in terms of both CNV and gene expression, but no correlation with presentation of ATH was found

Genetic profiling of autoinflammatory disorders in patients with periodic fever: a prospective study

Periodic fever syndromes (PFS) are an emerging group of autoinflammatory disorders. Clinical overlap exists and multiple genetic analyses may be needed to assist diagnosis. We evaluated the diagnostic value of a 5-gene sequencing panel (5GP) in patients with undiagnosed PFS

Impact of methylmercury and other heavy metals exposure on neurocognitive function in children of 7 years old: study protocol of the follow-up

BACKGROUND: The extent to which prenatal low-level mercury (Hg) exposure through maternal fish intake and heavy metals exposure affect children neurodevelopment is controversial and may appear in long term. In 2007 a prospective cohort, the Northern Adriatic Cohort II (NAC-II), was established to investigate the association between prenatal Hg exposure from maternal fish consumption and child neurodevelopment. 900 pregnant women were enrolled. 632 and 470 children underwent neurodevelopmental evaluation, respectively, at 18 and 40 months of age. The NAC-II cohort is a part of the Mediterranean cohort in "Public health impact of long-term, low-level, mixed element exposure in susceptible population strata" project.METHODS: This protocol describes the follow-up assessment of the effects of prenatal low level Hg and other heavy metals exposure on the developing nervous system of the children born within the NAC-II and reached the age of 7 years. Child diet components are estimated through a Diet Diary. Child hair and urine are collected for determination of Hg level. In addition, levels of other potentially neurotoxic metals, namely Manganese, Cadmium, Lead, Arsenic and Selenium are also measured in the same matrices.DiscussionThis protocol extends to the first years of schooling age the evaluation of the neurotoxicant effect of Mercury and of the other heavy metals on children's neurodevelopment, adjusting for the potential confounders such as the lifestyles and the social economic status of children's families. Longitudinal analysis of neurodevelopment, assessed in different ages (18, 40 months and 7 years), are performed

Molecular epidemiology of Usher syndrome in Italy

Purpose: Usher syndrome is an autosomal recessive disorder characterized by hearing and vision loss. Usher syndrome is divided into three clinical subclasses (type 1, type 2, and type 3), which differ in terms of the severity and progression of hearing loss and the presence or absence of vestibular symptoms. Usher syndrome is defined by significant genetic heterogeneity, with at least 12 distinct loci described and 9 genes identified. This study aims to provide a molecular epidemiology report of Usher syndrome in Italy. Methods: Molecular data have been obtained on 75 unrelated Italian patients using the most up-to date technology available for the screening of Usher syndrome gene mutations, i.e., the genotyping microarray developed by Asper Biotech (Tartu, Estonia), which simultaneously investigates 612 different marker positions using the well established arrayed primer extension methodology (APEX). Results: Using this method, we found that 12% of cases (9 out of 75) harbored homozygous or compound heterozygous mutations in the gene positions analyzed, whereas 20% (15 out of 75) of the patients were characterized by the presence of only one mutated allele based on the positions analyzed. One patient was found to be compound heterozygous for mutations in two different genes and this represents an example of possible digenic inheritance in Usher syndrome. A total of 66.6% of cases (50 out of 75) were found to be completely negative for the presence of Usher syndrome gene mutations in the detected positions. Mutations detected by the array were confirmed by direct sequencing. Conclusions: These findings highlight the efficacy of the APEX-based genotyping approach in the molecular assessment of Usher patients, suggesting the presence of alleles not yet identified and/or the involvement of additional putative genes that may account for the pathogenesis of Usher syndrome

Vertebral defects in patients with Peters plus syndrome and mutations in B3GALTL

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Abnormal expression of leiomyoma cytoskeletal proteins involved in cell migration

Uterine leiomyomas are monoclonal tumors. Several factors are involved in the neoplastic transformation of the myometrium. In our study we focused on dysregulated cytoskeletal proteins in the leiomyoma as compared to the myometrium. Paired tissue samples of ten leiomyomas and adjacent myometria were obtained and analyzed by two\u2011dimensional gel electrophoresis (2-DE). Mass spectrometry was used for protein identification, and western blotting for 2-DE data validation. The values of ten cytoskeletal proteins were found to be significantly different: eight proteins were upregulated in the leiomyoma and two proteins were downregulated. Three of the upregulated proteins (myosin regulatory light polypeptide 9, four and a half LIM domains protein 1 and LIM and SH3 domain protein 1) are involved in cell migration, while downregulated protein transgelin is involved in replicative senescence. Myosin regulatory light polypeptide 9 (MYL9) was further validated by western blotting because it is considered to be a cell migration marker in several cancers and could play a key role in leiomyoma development. Our data demonstrate significant alterations in the expression of cytoskeletal proteins involved in leiomyoma growth. A better understanding of the involvement of cytoskeletal proteins in leiomyoma pathogenesis may contribute to the identification of new therapeutic targets and the development of new pharmacological approaches