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MODY2 is the most prevalent monogenic form of diabetes in Italy with an estimated prevalence of about 0.5–1.5%. MODY2 is potentially indistinguishable from other forms of diabetes, however, its identification impacts on patients’ quality of life and healthcare resources. Unfortunately, DNA direct sequencing as diagnostic test is not readily accessible and expensive. In addition current guidelines, aiming to establish when the test should be performed, proved a poor detection rate. Aim of this study is to propose a reliable and easy-to-use tool to identify candidate patients for MODY2 genetic testing. We designed and validated a diagnostic flowchart in the attempt to improve the detection rate and to increase the number of properly requested tests. The flowchart, called 7-iF, consists of 7 binary ‘‘yes or no’’ questions and its unequivocal output is an indication for whether testing or not. We tested the 7-iF to estimate its clinical utility in comparison to the clinical suspicion alone. The 7-iF, in a prospective 2-year study (921 diabetic children) showed a precision of about the 76%. Using retrospective data, the 7-iF showed a precision in identifying MODY2 patients of about 80% compared to the 40% of the clinical suspicion. On the other hand, despite a relatively high number of missing MODY2 patients, the 7-iF would not suggest the test for 90% of the non-MODY2 patients, demonstrating that a wide application of this method might 1) help less experienced clinicians in suspecting MODY2 patients and 2) reducing the number of unnecessary tests. With the 7-iF, a clinician can feel confident of identifying a potential case of MODY2 and suggest the molecular test without fear of wasting time and money. A Qaly-type analysis estimated an increase in the patients’ quality of life and savings for the health care system of about 9 million euros per year.

Background: Several studies have shown an elevated prevalence of coeliac disease (CD) in sibs of coeliac patients (risk 8-12%). Aim and method: This study seeks to evaluate the risk that sibs of children with CD will also develop CD. This cohort of 188 Italian families was composed of probands with CD, at least one sib, and both parents. CD status was determined and HLA-DQ genotyping performed for all family members. The study also used a data set of Italian triads (127 probands and both their parents) also genotyped for HLA-DQ. Results: The overall risk that a sib of a CD patient will develop the disease is estimated at 10% in this sample. The risk estimate ranges from 0,1 to 29% when HLA-DQ information of the proband, parents and sib is considered. We found a negligible risk (lower than 1%) for 40% of the sibs of probands, a risk greater than 1% but less than 10% for 30%, and finally a high or very high risk (above 25%) in one third of families. Conclusion: These results make it possible to provide more accurate information to parents with child with CD about the real risk for another child. An antenatal estimate of the order of risk of CD is now possible. Specific follow-up can thus be offered for babies at high risk.

MODY2 is the most prevalent monogenic form of diabetes in Italy with an estimated prevalence of about 0.5-1.5%. MODY2 is potentially indistinguishable from other forms of diabetes, however, its identification impacts on patients' quality of life and healthcare resources. Unfortunately, DNA direct sequencing as diagnostic test is not readily accessible and expensive. In addition current guidelines, aiming to establish when the test should be performed, proved a poor detection rate. Aim of this study is to propose a reliable and easy-to-use tool to identify candidate patients for MODY2 genetic testing. We designed and validated a diagnostic flowchart in the attempt to improve the detection rate and to increase the number of properly requested tests. The flowchart, called 7-iF, consists of 7 binary "yes or no" questions and its unequivocal output is an indication for whether testing or not. We tested the 7-iF to estimate its clinical utility in comparison to the clinical suspicion alone. The 7-iF, in a prospective 2-year study (921 diabetic children) showed a precision of about the 76%. Using retrospective data, the 7-iF showed a precision in identifying MODY2 patients of about 80% compared to the 40% of the clinical suspicion. On the other hand, despite a relatively high number of missing MODY2 patients, the 7-iF would not suggest the test for 90% of the non-MODY2 patients, demonstrating that a wide application of this method might 1) help less experienced clinicians in suspecting MODY2 patients and 2) reducing the number of unnecessary tests. With the 7-iF, a clinician can feel confident of identifying a potential case of MODY2 and suggest the molecular test without fear of wasting time and money. A Qaly-type analysis estimated an increase in the patients' quality of life and savings for the health care system of about 9 million euros per year.

Aims: Many authors do not recommend hypoglycemic treatment during pregnancy in women affected by monogenic diabetes due to heterozygous glucokinase (GCK) mutations (MODY 2) in case of affected fetus, because maternal hyperglycemia would be necessary to achieve a normal birthweight. We aimed to evaluate differences in birthweight between MODY 2 affected children according to the parent who carried the mutation. Methods: We retrospectively studied 48 MODY 2 affected children, whose mothers did not receive hypoglycemic treatment during pregnancy, divided into two groups according to the presence of the mutation in the mother (group A) or in the father (group B). Data were extracted from the database of the Regional Centre of Pediatric Diabetology of the University of Campania, Naples, collected from 1996 to 2016. We analyzed birthweight and centile birthweight. Results: Percentage of small for gestational age was significantly higher in group B than in group A. We found three large for gestational age in the group that inherited the deficit from the mother, all with the same novel GCK mutation (p.Lys458-Cys461del). Conclusions: We hypothesize that not all MODY 2 affected fetuses need the same levels of hyperglycemia to have an appropriate growth, maybe because different kinds of GCK mutations may result in different phenotypes. Consequently, a “tailored therapy” of maternal hyperglycemia, based on fetal growth frequently monitored through ultrasounds, is essential in MODY 2 pregnancies.

Wolfram Syndrome (WS) is a very rare genetic disorder characterized by several symptoms that occur from childhood to adulthood. Usually, the first clinical sign is non-autoimmune diabetes even if other clinical features (optic subatrophy, neurosensorial deafness, diabetes insipidus) may be present in an early state and may be diagnosed after diabetes’ onset. Prognosis is poor, and the death occurs at the median age of 39 years as a consequence of progressive respiratory impairment, secondary to brain atrophy and neurological failure. The aim of this paper is the description of the metabolic treatment of the WS. We reported the experience of long treatment in patients with this syndrome diagnosed in pediatric age and followed also in adult age. It is known that there is a correlation between metabolic control of diabetes, the onset of other associated symptoms, and the progression of the neurodegenerative alterations. Therefore, a multidisciplinary approach is necessary in order to prevent, treat and carefully monitor all the comorbidities that may occur. An extensive understanding of WS from pathophysiology to novel possible therapy is fundamental and further studies are needed to better manage this devastating disease and to guarantee to patients a better quality of life and a longer life expectancy.

Maturity onset diabetes of the young (MODY) is the most common form of monogenic diabetes in Europe, affecting between 1 and 6% of diabetic patients. It comprises a group of heterogeneous genetic disorders characterized by early onset of diabetes (commonly before age 25), absence of autoimmunity, and beta-cell dysfunction. So far, mutations in 14 different genes involved in glucose homeostasis and pancreatic development [1] have been associated with this disease. Although it is an autosomal dominant disorder, de novo mutations should be taken into consideration in patients without a family history of diabetes. Most cases of MODY are due to mutations in GCK, HNF1a, HNF4a and HNF1b, previously known as MODY2, MODY3, MODY1, and MODY5, respectively. Among these, HNF1b is an active transcription factor that forms homodimers or heterodimers with HNF1a and plays a fundamental role in kidney development, nephron differentiation, and pancreatic growth and differentiation. Mutations in this gene lead to congenital anomalies of the kidney and urinary tract, genital malformations, pancreatic atrophy with endocrine and exocrine deficiency [2]. Diabetes usually onsets in early adulthood and frequently requires insulin treatment. We present an unusual case of a prenatal diagnosis that revealed a mutation in the HNF1b gene responsible for neonatal hyperglycemia in a pregnant woman affected by X-linked ectodermal dysplasia.

Maturity Onset Diabetes of the Young (MODY), the most frequent form of monogenic diabetes, comprises a group of heterogeneous disorders, characterized by non-autoimmune diabetes due to mutations of at least 14 different genes. We report a case of a 38-years-old patient with non-autoimmune diabetes, where molecular analysis evidenced a large deletion on chromosome 17q12 including several genes, among them HNF1β associated to MODY5. The analysis allowed us to clarify the complex phenotype of the patient including, in addition to diabetes, intellectual disability, seizures, kidney cysts and facial dimorphisms. This case shows that diabetes when caused by large deletions, can be just one of the symptoms of a “clinical syndrome” that includes other features due to the deletion of neighboring genes and confirms the important role of the molecular test to obtain a correct diagnosis in a patient with a suspicion of monogenic diabetes.

Sudden cardiac death (SCD) is a devastating event which can also affect people in apparent good health, such as young athletes. It is known that intense and continuous exercise along with a genetic background that predisposes a person to the risk of fatal arrhythmias is a trigger for SCD. Therefore, knowledge of the athlete’s genetic conditions underlying the onset of SCD must be extended, in order to develop new effective prevention and/or therapeutic strategies. Arrhythmic features occur across a broad spectrum of cardiac diseases, sometimes presenting with overlapping phenotypes. The genetic basis of arrhythmogenic disorders has been greatly highlighted in the last 30 years, and has shown marked heterogeneity. The advent of next-generation sequencing has constantly updated our understanding of the genetic basis of arrhythmogenic diseases and is laying the foundation for precision medicine. With the exception of a few clinical cases involving a single athlete showing a highly suspected phenotype for the presence of a heart disease, there are few studies to date that analysed the applicability of genetic testing on cohorts of athletes. This evidence shows that genetic testing can contribute to the diagnosis of up to 13% of athletes; however, the presence of clinical markers is essential. This review aims to provide a reference collection on current knowledge of the genetic basis of sudden cardiac death in athletes and to review updated evidence on the effectiveness of genetic testing in early identification of athletes at risk for SCD.

Congenital diabetes mellitus is a rare disorder characterized by hyperglycemia that occurs shortly after birth. We define "Diabetes of Infancy" if hyperglycemia onset before 6 months of life. From the clinical point of view, we distinguish two main types of diabetes of infancy: transient (TNDM), which remits spontaneously, and permanent (PNDM), which requires lifelong treatment. TNDM may relapse later in life. About 50% of cases are transient (TNDM) and 50% permanent. Clinical manifestations include severe intrauterine growth retardation, hyperglycemia and dehydration. A wide range of different associated clinical signs including facial dysmorphism, deafness and neurological, cardiac, kidney or urinary tract anomalies are reported. Developmental delay and learning difficulties may also be observed. In this paper we review all the causes of congenital diabetes and all genes and syndromes involved in this pathology. The discovery of the pathogenesis of most forms of congenital diabetes has made it possible to adapt the therapy to the diagnosis and in the forms of alteration of the potassium channels of the pancreatic Beta cells the switch from insulin to glibenclamide per os has greatly improved the quality of life. Congenital diabetes, although it is a very rare form, has been at the must of research in recent years especially for pathogenesis and pharmacogenetics. The most striking difference compared to the more frequent autoimmune diabetes in children (type 1 diabetes) is the possibility of treatment with hypoglycemic agents and the apparent lower frequency of chronic complications.

Type 1 diabetes mellitus (DM) is characterized by irreversible, autoimmune, pancreatic β-cell destruction. During the disease, some patients experience a phase of Partial Clinical Remission (PCR) known as “honeymoon.” This is a transitory period that is characterized by insulin production by residual β cells following DM diagnosis and initiating the insulin therapy. In this study, we aimed to evaluate the influence of insulin production on immune system after the onset of diabetes, and we showed that the duration of honeymoon period could be related to the onset of other autoimmune conditions. For this retrospective study, 159 children aged between 11 and 18 years with type 1 DM were eligible. They have been diagnosed diabetes at least 10 years ago and use exogenous insulin. Our results showed that younger age at the onset of Type 1 DM in children, predicts Celiac Disease. Female sex and low HCO3 levels at the onset of DM had a high predictive value on patients who did not experience longer Partial Clinical Remission phase. Patients with higher BMI at the diagnosis of DM experienced shorter honeymoon period than the average. Smaller of our patients who diagnosed just DM have more than 297 days honeymoon period with respect to patients with one associated autoimmune disease. This may be due to a continuous and prolonged stimulation of immune system during the period of honeymoon that predispose the patient to develop other TH1 diseases. The patients who experienced more than 297 days Partial Clinical Remission seem under risk of developing one other autoimmune disease more than the patients who experienced less than 297 days Partial Clinical Remission. We have to consider that this observation is very intriguing because many protocols spring-up to try prolonging the honeymoon period in patients with autoimmune DM. If this aim is important from a metabolic point of view, long follow-ups are needed to be sure that the risk of other autoimmune diseases does not increase.

Mitochondrial Cardiomyopathy (MCM) is a common manifestation of multi-organ Mitochondrial Diseases (MDs), occasionally present in non-syndromic cases. Diagnosis of MCM is complex because of wide clinical and genetic heterogeneity and requires medical, laboratory, and neuroimaging investigations. Currently, the molecular screening for MCM is fundamental part of MDs management and allows achieving the definitive diagnosis. In this article, we review the current genetic knowledge associated with MDs, focusing on diagnosis of MCM and MDs showing cardiac involvement. We searched for publications on mitochondrial and nuclear genes involved in MCM, mainly focusing on genetic screening based on targeted gene panels for the molecular diagnosis of the MCM, by using Next Generation Sequencing. Here we report twelve case reports, four case-control studies, eleven retrospective studies, and two prospective studies, for a total of twenty-nine papers concerning the evaluation of cardiac manifestations in mitochondrial diseases. From the analysis of published causal mutations, we identified 130 genes to be associated with mitochondrial heart diseases. A large proportion of these genes (34.3%) encode for key proteins involved in the oxidative phosphorylation system (OXPHOS), either as directly OXPHOS subunits (22.8%), and as OXPHOS assembly factors (11.5%). Mutations in several mitochondrial tRNA genes have been also reported in multi-organ or isolated MCM (15.3%). This review highlights the main disease-genes, identified by extensive genetic analysis, which could be included as target genes in next generation panels for the molecular diagnosis of patients with clinical suspect of mitochondrial cardiomyopathies.

Choroideremia (CHM), an X-linked degen- eration of the retinal pigmented epithelium (RPE), pho- toreceptors, and choroid, ultimately leads to blindness. It is caused by loss-of-function of the CHM gene prod- uct, the Rab escort protein 1 (REP1) that is involved, together with its homologue REP2, in prenylation of Rab GTPases, key regulators of intracellular vesicular traf- fic. Here, we report the molecular characterization of 20 unrelated Italian families affected by CHM. We identi- fied 19 different mutations, nine of which are new. In most cases, we analyzed the effect of the mutations at the mRNA level. Furthermore, we demonstrated, by in vitro trancription/translation assays, that the mutated mRNAs produced truncated proteins in all cases but one. In fact, we also identified a novel REP1 missense variant (c.1520A>G; p.H507R) associated to CHM. Thus far, only two other CHM-associated missense mutations have been identified, one of which was a splicing alteration. We investigated the impact of the p.H507R amino acid change on REP1 structure and function, thus providing the first experimental demonstration that correlates a mis- sense mutation in CHM with a functional impairment of REP1. Overall, our results indicate that the REP1-Rab geranyl-geranyl transferase interaction and consequently REP1-mediated Rab prenylation is essential for RPE and photoreceptor function. Hum Mutat 32:1460-1469, 2011. C 2011 Wiley Periodicals, Inc.

BACKGROUND: MODY diabetes includes rare familiar forms due to genetic mutations resulting in β-cell dysfunction. MODY 3 is due to mutations in the gene transcription factor HNF-1α, with diabetes diagnosis in adolescence or early adult life. Few data are available about MODY 3 in pregnancy. CASE REPORT: A 36-year-old Italian woman came to our unit at the 5th week of pregnancy. She was diagnosed with diabetes at 18 years, with negative autoimmunity and a strong familiarity for diabetes. She was treated with gliclazide and metformin. She had a previous pregnancy in which she was treated with insulin, giving birth at 38 weeks to a 3.210 kg baby girl, who showed neonatal hypoglycemia. We switched her to insulin treatment according to guidelines. We asked for genetic molecular testing, resulting in a HNF-1α gene mutation. A US examination at 7 weeks revealed a twin, bicorial, biamniotic pregnancy. At 37 weeks of gestation, she gave birth to two normal-weight baby girls; only one showed neonatal hypoglycemia and a genetic test revealed that she was affected by HNF-1α gene mutation. Subsequently, entire family of the woman was tested, showing that the father, the sister and the first daughter had the same HNF-1α mutation. DISCUSSION: A MODY 3 foetus needs a near-normal maternal glycemic control, because the exposure to intrauterine hyperglycemia can lead to an earlier age of diabetes onset. Neonatal hypoglycemia is generally observed in MODY 1 infants, but it is possible to hypothesize that some HNF-1α mutations could lead to a functionally impaired protein that might dysregulate HNF-4α expression determining hypoglycemia.

Background: Maturity onset diabetes of the young type 2 (or GCK MODY) is a genetic form of diabetes mellitus provided by mutations in the glucokinase gene (GCK). Methodology/Principal Findings: We screened the GCK gene by direct sequencing in 30 patients from South Italy with suspected MODY. The mutation-induced structural alterations in the protein were analyzed by molecular modelling. The patients' biochemical, clinical and anamnestic data were obtained. Mutations were detected in 16/30 patients (53%); 9 of the 12 mutations identified were novel (p.Glu70Asp, p.Phe123Leu, p.Asp132Asn, p.His137Asp, p.Gly162Asp, p.Thr168Ala, p.Arg392Ser, p.Glu290X, p.Gln106_Met107delinsLeu) and are in regions involved in structural rearrangements required for catalysis. The prevalence of mutation sites was higher in the small domain (7/12: -59%) than in the large (4/12: 33%) domain or in the connection (1/12:8%) region of the protein. Mild diabetic phenotypes were detected in almost all patients [mean (SD) OGTT=7.8 mMol/L (1.8)] and mean triglyceride levels were lower in mutated than in unmutated GCK patients [p=0.04]. Conclusions: The prevalence of GCK MODY is high in southern Italy, and the GCK small domain is a hot spot for MODY mutations. Both the severity of the GCK mutation and the genetic background seem to play a relevant role in the GCK MODY phenotyope. Indeed, a partial genotype-phenotype correlation was identified in related patients (3 pairs of siblings) but nut in two unrelated children bearing the same mutations. Thus, the molecular approach allows the physician to confirm the diagnosis and to predict severity of the mutation. © 2008 Tinto et al.

Background: Genotyping based on short tandem repeat (STR) regions is widely used in human identification and parentage testing, in gene mapping studies, and as an approach to studies on the etiopathogenesis and diagnosis of hereditary diseases. We wished to study a new analytical approach that uses capillary electrophoresis and multicolor fluorescence in place of slab gel electrophoresis. Methods: We evaluated the efficiency for parentage and forensic purposes of the AmpFLSTR Profiler PlusTM typing kit that is used with the ABI Prism 310 Genetic Analyzer (System-2 STR), and that of a widely used panel of nine STRs analyzed with conventional slab-gel electrophoresis followed by radioactive detection (System-1 STR). System-2 STR, based on automated capillary electrophoresis and automated sizing of the alleles by Genotyper 2.0 software, was used to determine the allele frequency of the nine loci in 157 Caucasian subjects from southern Italy. On the basis of the data obtained, we submitted 40 trios to parentage testing. Results: A higher median probability of paternity attribution and power of exclusion were obtained with System-2 STR vs System-1 STR: respectively, 99.99% and 99.95% (P

Malignant hyperthermia (MH) is an inherited autosomal dominant pharmacogenetic disorder and is the major cause of anaesthesia-induced death. Malignant hyperthermia susceptibility is usually diagnosed by the in vitro contracture test (IVCT) performed on fresh muscle biopsies exposed to caffeine and halothane, respectively. Around 50% of affected families are linked to the ryanodine receptor (RYR1) gene. The human RYR1 gene maps to chromosome 19q13.1 and encodes a protein that associates as a homotetramer and acts as a calcium-release channel from the sarcoplasmic reticulum. To date, 17 mutations have been identified in the coding region of the RYR1 gene and appear to be associated to the MH-susceptible phenotype. Here we describe a rare case of discordance between genotype (characterised by the presence of the Arg614Cys mutation in the RYR1 gene) and MH-normal typed phenotype. Although the IVCT remains a very reliable procedure for the assessment of MH status, genetic data can provide in some cases an additional aid to clinical diagnosis.

Type 2 Maturity Onset Diabetes of the Young (MODY2) is a monogenic autosomal disease characterized by a primary defect in insulin secretion and hyperglycemia. It results from GCK gene mutations that impair enzyme activity. Between 2006 and 2010, we investigated GCK mutations in 66 diabetic children from southern Italy with suspected MODY2. Denaturing High Performance Liquid Chromatography (DHPLC) and sequence analysis revealed 19 GCK mutations in 28 children, six of which were novel: p.Glu40Asp, p.Val154Leu, p.Arg447Glyfs, p.Lys458_Cys461del, p.Glu395_Arg397del and c.580-2A>T. We evaluated the effect of these 19 mutations using bioinformatic tools such as Polymorphism Phenotyping (Polyphen), Sorting Intolerant From Tolerant (SIFT) and in silico modelling. We also conducted a functional study to evaluate the pathogenic significance of seven mutations that are among the most severe mutations found in our population, and have never been characterized: p.Glu70Asp, p.His137Asp, p.Phe150Tyr, p.Val154Leu, p.Gly162Asp, p.Arg303Trp and p.Arg392Ser. These seven mutations, by altering one or more kinetic parameters, reduced enzyme catalytic activity by >40%. All mutations except p.Glu70Asp displayed thermal-instability, indeed >50% of enzyme activity was lost at 50°C/30 min. Thus, these seven mutations play a pathogenic role in MODY2 insurgence. In conclusion, this report revealed six novel GCK mutations and sheds some light on the structure-function relationship of human GCK mutations and MODY2.