The heterozygous chromosome deletion within the band 22q11 (del22q11) is an important cause of congenital cardiovascular defects. It is the genetic basis of DiGeorge syndrome and causes the most common deletion syndrome in humans. Because the deleted region is largely conserved in the mouse, we were able to engineer a chromosome deletion (Df1) spanning a segment of the murine region homologous to the human deleted region. Here we describe heterozygously deleted (Df1/+) mice with cardiovascular abnormalities of the same type as those associated with del22q11; we have traced the embryological origin of these abnormalities to defective development of the fourth pharyngeal arch arteries. Genetic complementation of the deletion using a chromosome duplicated for the Df1 DNA segment corrects the heart defects, indicating that they are caused by reduced dosage of genes located within Df1. The Df1/+ mouse model reveals the pathogenic basis of the most clinically severe aspect of DiGeorge syndrome and uncovers a new mechanism leading to aortic arch abnormalities. These mutants represent a mouse model of a human deletion syndrome generated by chromosome engineering.

Autosomal dominant myotonia congenita or Thomsen's disease and autosomal recessive myotonia congenita or Becker's are rare nondystrophic disorders due to allelic mutations of the muscle chloride channel gene, CLCN1. We have analysed all 24 exons of the CLCN1 gene, in a panel of 20 unrelated patients (9 with dominant and 11 with recessive mytotonia congenita). We have found five novel mutations including two missense (V5631, F708L), one nonsense (C481X), one splicing (IVS19+2T->A), and one frameshift (2264delC), and also detected the recurrent R894X mutation. These account for 10 of the 22 recessive alleles examined, while no mutations were found in the dominant form. We report three novel polymorphisms (-134T/G, 898C/A and 2154T/C). Our results support high molecular heterogeneity of these myotonias in Italian population and provide new insight for the diagnosis and genetic counselling of these diseases.

Context Measurement of circulating miRNAs (c-miRNAs), as potential biomarkers of fragility fracture risk, has recently become a subject of investigation. Objective Measure, by next generation sequencing (NGS), global miRNA expression in serum samples of osteoporotic subjects vs individuals with normal bone mineral density (BMD). Design Samples were collected from patients with different bone phenotypes and/or fragility fractures who did not receive any anti-resorptive and/or bone-forming drug at the time of blood collection. Setting Samples and data were collected at 7 medical centers in Italy. Patients NGS pre-screening: 50 osteoporotic patients vs 30 individuals with normal BMD. Droplet Digital PCR (ddPCR) validation: 213 patients with different bone phenotypes, including the NGS-analyzed cohort. Results NGS identified 5 miRNAs (miR-8085, miR-320a-3p, miR-23a-3p, miR-4497, miR-145-5p) differentially expressed in osteoporosis cases without fractures vs controls. ddPCR validation confirmed a lower c-miR-23a-3p expression in osteoporotic patients, with or without fracture, than in osteopenic and normal subjects, and an increased c-miR-320a-3p expression in osteoporotic patients with fracture and a lower expression in osteoporotic patients without fracture. ddPCR analysis showed a significantly increased expression of miR-21-5p in osteoporotic patients, with or without fracture, than in osteopenic and normal subjects, not evidenced by the NGS pre-screening. Discussion Our study confirmed levels of c-miR-23a-3p and c-miR-21-5p as able to distinguish osteoporotic patients and subjects with normal BMD. Increased levels of c-miR-320a-3p resulted specifically associated with fractures, independently by BMD, suggesting c-miR-320a-3p as a prognostic indicator of fracture risk in osteoporotic patients, to be confirmed in prospective studies on incident fractures.

The vitamin D receptor (VDR) regulates bone development and calcium homeostasis, suggesting a central role in musculoskeletal diseases such as osteoporosis (OP). Several studies have examined the contribution of VDR polymorphisms and epigenetic signatures in bone metabolism and OP risk, with sometimes inconclusive results. Our study aimed to explore the association between genetic variability, expression and the methylation pattern of VDR with the risk of OP in a cohort of Caucasian patients. Genomic DNA from 139 OP, 54 osteopenic (Ope) and 73 healthy (CTR) subjects were used for genotyping the rs731236 (TaqI), rs2228570 (FokI) and rs11568820 (Cdx2) polymorphisms of the VDR gene by an allelic discrimination assay. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis of VDR expression levels and pyrosequencing analysis of a VDR promoter CpG island were carried out in a subcohort (25 OP and 25 CTR) of subjects. Data obtained showed a significantly higher OP risk for rs11568820 G/A and A/A genotypes (p = 0.05). qRT-PCR revealed lower VDR gene expression levels in the OP group compared to CTR subjects (p = 0.0009), also associated with both the rs11568820 A/A genotype (p = 0.03) and femoral fragility fractures (p = 0.05). No association was found between the methylation pattern of the region analyzed of the VDR promoter and its expression levels. Our results identify a significative association between Cdx2 rs11568820 polymorphism and OP risk. In addition, the VDR transcriptomic profile suggests a putative interconnection with OP progression, providing a useful tool to stratify OP phenotype and fragility fracture risk.

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