Integrated Approach to Highlighting the Molecular Bases of a Deep Vein Thrombosis Event in an Elite Basketball Athlete.

Acute or intense exercise can result in metabolic imbalances, muscle injuries, or reveal hidden disorders. Laboratory medicine in sports is playing an increasingly crucial role in monitoring athletes' health conditions. In this study, we designed an integrated approach to explore the causes of a deep venous thrombosis event in an elite basketball player. Since the complete blood count revealed a marked platelet count (838 × 103 µL), and thrombophilia screening tests did not reveal any significant alteration, we evaluated the thrombin generation, which highlights a state of hypercoagulability. First-level haemostasis exams showed only a slight prolongation of the activated Partial Thromboplastin Time (aPTT). Thus, screening tests for von Willebrand Disease showed a reduction in vWF parameters. Therefore, we directed our hypothesis towards a diagnosis of acquired von Willebrand disease secondary to Essential Thrombocythemia (ET). To confirm this hypothesis and highlight the molecular mechanism underlying the observed phenotype, molecular tests were performed to evaluate the presence of the most common mutations associated with ET, revealing a 52-bp deletion in the coding region of CALR exon 9. This case report highlights the importance of an integrated approach to monitoring the athletes' health status to personalise training and treatments, thus avoiding the appearance of diseases and injuries that, if underestimated, can undermine the athlete's life.

Clinical, Genetic, and Histological Characterization of Patients with Rare Neuromuscular and Mitochondrial Diseases Presenting with Different Cardiomyopathy Phenotypes.

Cardiomyopathies are mostly determined by genetic mutations affecting either cardiac muscle cell structure or function. Nevertheless, cardiomyopathies may also be part of complex clinical phenotypes in the spectrum of neuromuscular (NMD) or mitochondrial diseases (MD). The aim of this study is to describe the clinical, molecular, and histological characteristics of a consecutive cohort of patients with cardiomyopathy associated with NMDs or MDs referred to a tertiary cardiomyopathy clinic. Consecutive patients with a definitive diagnosis of NMDs and MDs presenting with a cardiomyopathy phenotype were described. Seven patients were identified: two patients with ACAD9 deficiency (Patient 1 carried the c.1240C>T (p.Arg414Cys) homozygous variant in ACAD9; Patient 2 carried the c.1240C>T (p.Arg414Cys) and the c.1646G>A (p.Ar549Gln) variants in ACAD9); two patients with MYH7-related myopathy (Patient 3 carried the c.1325G>A (p.Arg442His) variant in MYH7; Patient 4 carried the c.1357C>T (p.Arg453Cys) variant in MYH7); one patient with desminopathy (Patient 5 carried the c.46C>T (p.Arg16Cys) variant in DES); two patients with mitochondrial myopathy (Patient 6 carried the m.3243A>G variant in MT-TL1; Patient 7 carried the c.253G>A (p.Gly85Arg) and the c.1055C>T (p.Thr352Met) variants in MTO1). All patients underwent a comprehensive cardiovascular and neuromuscular evaluation, including muscle biopsy and genetic testing. This study described the clinical phenotype of rare NMDs and MDs presenting as cardiomyopathies. A multidisciplinary evaluation, combined with genetic testing, plays a main role in the diagnosis of these rare diseases, and provides information about clinical expectations, and guides management.

Protein haploinsufficiency drivers identify MYBPC3 variants that cause hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac disease. Variants in MYBPC3, the gene encoding cardiac myosin-binding protein C (cMyBP-C), are the leading cause of HCM. However, the pathogenicity status of hundreds of MYBPC3 variants found in patients remains unknown, as a consequence of our incomplete understanding of the pathomechanisms triggered by HCM-causing variants. Here, we examined 44 nontruncating MYBPC3 variants that we classified as HCM-linked or nonpathogenic according to cosegregation and population genetics criteria. We found that around half of the HCM-linked variants showed alterations in RNA splicing or protein stability, both of which can lead to cMyBP-C haploinsufficiency. These protein haploinsufficiency drivers associated with HCM pathogenicity with 100% and 94% specificity, respectively. Furthermore, we uncovered that 11% of nontruncating MYBPC3 variants currently classified as of uncertain significance in ClinVar induced one of these molecular phenotypes. Our strategy, which can be applied to other conditions induced by protein loss of function, supports the idea that cMyBP-C haploinsufficiency is a fundamental pathomechanism in HCM.

Germline rare variants of lectin pathway genes predispose to asymptomatic SARS-CoV-2 infection in elderly individuals.

PURPOSE: Emerging evidence suggest that infection-dependent hyperactivation of complement system (CS) may worsen COVID-19 outcome. We investigated the role of predicted high impact rare variants - referred as qualifying variants (QVs) - of CS genes in predisposing asymptomatic COVID-19 in elderly individuals, known to be more susceptible to severe disease. METHODS: Exploiting exome sequencing data and 56 CS genes, we performed a gene-based collapsing test between 164 asymptomatic subjects (aged ≥60 years) and 56,885 European individuals from the Genome Aggregation Database. We replicated this test comparing the same asymptomatic individuals with 147 hospitalized patients with COVID-19. RESULTS: We found an enrichment of QVs in 3 genes (MASP1, COLEC11, and COLEC10), which belong to the lectin pathway, in the asymptomatic cohort. Analyses of complement activity in serum showed decreased activity of lectin pathway in asymptomatic individuals with QVs. Finally, we found allelic variants associated with asymptomatic COVID-19 phenotype and with a decreased expression of MASP1, COLEC11, and COLEC10 in lung tissue. CONCLUSION: This study suggests that genetic rare variants can protect from severe COVID-19 by mitigating the activity of lectin pathway and prothrombin. The genetic data obtained through ES of 786 asymptomatic and 147 hospitalized individuals are publicly available at http://espocovid.ceinge.unina.it/.

Diagnostic and Therapeutic Potential for HNP-1, HBD-1 and HBD-4 in Pregnant Women with COVID-19.

Pregnancy is characterized by significant immunological changes and a cytokine profile, as well as vitamin deficiencies that can cause problems for the correct development of a fetus. Defensins are small antimicrobial peptides that are part of the innate immune system and are involved in several biological activities. Following that, this study aims to compare the levels of various cytokines and to investigate the role of defensins between pregnant women with confirmed COVID-19 infection and pregnant women without any defined risk factor. TNF-α, TGF-ß, IL-2 and IL-10, ß-defensins, have been evaluated by gene expression in our population. At the same time, by ELISA assay IL-6, IL-8, defensin alpha 1, defensin beta 1 and defensin beta 4 have been measured. The data obtained show that mothers affected by COVID-19 have an increase in pro-inflammatory factors (TNF-α, TGF-ß, IL-2, IL-6, IL-8) compared to controls; this increase could generate a sort of "protection of the fetus" from virus attacks. Contemporarily, we have an increase in the anti-inflammatory cytokine IL-10 and an increase in AMPs, which highlights how the mother's body is responding to the viral attack. These results allow us to hypothesize a mechanism of "trafficking" of antimicrobial peptides from the mother to the fetus that would help the fetus to protect itself from the infection in progress.

The Risk of Sudden Unexpected Cardiac Death in Children: Epidemiology, Clinical Causes, and Prevention.

"Sudden unexplained death (SUD) is a tragic event for both the family and community, particularly when it occurs in young individuals. Sudden cardiac death (SCD) represents the leading form of SUD and is defined as an unexpected event without an obvious extracardiac cause, occurring within 1 hour after the onset of symptoms. In children, the main causes of SCD are inherited cardiac disorders, whereas coronary artery diseases (congenital or acquired), congenital heart diseases, and myocarditis are rare. The present review examines the current state of knowledge regarding SCD in children, discussing the epidemiology, clinical causes, and prevention strategies."

Cardiovascular Involvement in mtDNA Disease: Diagnosis, Management, and Therapeutic Options.

Mitochondrial diseases (MD) include an heterogenous group of systemic disorders caused by sporadic or inherited mutations in nuclear or mitochondrial DNA (mtDNA), causing impairment of oxidative phosphorylation system. Hypertrophic cardiomyopathy is the dominant pattern of cardiomyopathy in all forms of mtDNA disease, being observed in almost 40% of the patients. Dilated cardiomyopathy, left ventricular noncompaction, and conduction system disturbances have been also reported. In this article, the authors discuss the current clinical knowledge on MD, focusing on diagnosis and management of mitochondrial diseases caused by mtDNA mutations.

Impact of Regular Physical Activity on Aortic Diameter Progression in Paediatric Patients with Bicuspid Aortic Valve.

Patients with bicuspid aortic valve (BAV) have an increased risk of aortic dilation and aortic dissection or rupture. The impact of physical training on the natural course of aortopathy in BAV patients remains unclear. The aim of this study was to evaluate the impact of regular physical activity on aortic diameters in a consecutive cohort of paediatric patients with BAV. Consecutive paediatric BAV patients were evaluated and categorized into two groups: physically active and sedentary subjects. Only the subjects with a complete 2-year follow-up were included in the study. To evaluate the potential impact of physical activity on aortic size, aortic diameters were measured at the sinus of Valsalva and mid-ascending aorta using echocardiography. We defined aortic diameter progression the increase of aortic diameter ≥ 10% from baseline. Among 90 BAV patients (11.5 ± 3.4 years of age, 77% males), 53 (59%) were physically active subjects. Compared to sedentary, physically active subjects were not significantly more likely to have > 10% increase in sinus of Valsalva (13% vs. 8%, p-value = 0.45) or mid-ascending aorta diameter (9% vs. 13%, p-value = 0.55) at 2 years follow-up, both in subjects with sinus of Valsalva diameter progression (3.7 ± 1.0 mm vs. 3.5 ± 0.8 mm, p-value = 0.67) and in those with ascending aorta diameter progression (3.0 ± 0.8 mm vs. 3.2 ± 1.3 mm, p-value = 0.83). In our paediatric cohort of BAV patients, the prevalence and the degree of aortic diameter progression was not significantly different between physically active and sedentary subjects, suggesting that aortic dilation is unrelated to regular physical activity over a 2-year period.

Molecular Epidemiology of Mitochondrial Cardiomyopathy: A Search Among Mitochondrial and Nuclear Genes.

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.

Regulatory Noncoding and Predicted Pathogenic Coding Variants of CCR5 Predispose to Severe COVID-19.

Genome-wide association studies (GWAS) found locus 3p21.31 associated with severe COVID-19. CCR5 resides at the same locus and, given its known biological role in other infection diseases, we investigated if common noncoding and rare coding variants, affecting CCR5, can predispose to severe COVID-19. We combined single nucleotide polymorphisms (SNPs) that met the suggestive significance level (P ≤ 1 × 10-5) at the 3p21.31 locus in public GWAS datasets (6406 COVID-19 hospitalized patients and 902,088 controls) with gene expression data from 208 lung tissues, Hi-C, and Chip-seq data. Through whole exome sequencing (WES), we explored rare coding variants in 147 severe COVID-19 patients. We identified three SNPs (rs9845542, rs12639314, and rs35951367) associated with severe COVID-19 whose risk alleles correlated with low CCR5 expression in lung tissues. The rs35951367 resided in a CTFC binding site that interacts with CCR5 gene in lung tissues and was confirmed to be associated with severe COVID-19 in two independent datasets. We also identified a rare coding variant (rs34418657) associated with the risk of developing severe COVID-19. Our results suggest a biological role of CCR5 in the progression of COVID-19 as common and rare genetic variants can increase the risk of developing severe COVID-19 by affecting the functions of CCR5.

Childhood obesity: an overview of laboratory medicine, exercise and microbiome.

In the last few years, a significant increase of childhood obesity incidence unequally distributed within countries and population groups has been observed, thus representing an important public health problem associated with several health and social consequences. Obese children have more than a 50% probability of becoming obese adults, and to develop pathologies typical of obese adults, that include type 2-diabetes, dyslipidemia and hypertension. Also environmental factors, such as reduced physical activity and increased sedentary activities, may also result in increased caloric intake and/or decreased caloric expenditure. In the present review, we aimed to identify and describe a specific panel of parameters in order to evaluate and characterize the childhood obesity status useful in setting up a preventive diagnostic approach directed at improving health-related behaviors and identifying predisposing risk factors. An early identification of risk factors for childhood obesity could definitely help in setting up adequate and specific clinical treatments.

The Hidden Fragility in the Heart of the Athletes: A Review of Genetic Biomarkers.

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.

Molecular Basis of Inflammation in the Pathogenesis of Cardiomyopathies.

Cardiomyopathies (CMPs) represent a diverse group of heart muscle diseases, grouped into specific morphological and functional phenotypes. CMPs are associated with mutations in sarcomeric and non-sarcomeric genes, with several suspected epigenetic and environmental mechanisms involved in determining penetrance and expressivity. The understanding of the underlying molecular mechanisms of myocardial diseases is fundamental to achieving a proper management and treatment of these disorders. Among these, inflammation seems to play an important role in the pathogenesis of CMPs. The aim of the present study is to review the current knowledge on the role of inflammation and the immune system activation in the pathogenesis of CMPs and to identify potential molecular targets for a tailored anti-inflammatory treatment.

Laboratory medicine: health evaluation in elite athletes.

The need to evaluate the health status of an athlete represents a crucial aim in preventive and protective sports science in order to identify the best diagnostic strategy to improve performance and reduce risks related to physical exercise. In the present review we aim to define the main biochemical and haematological markers that vary significantly during and after sports training to identify risk factors, at competitive and professional levels and to highlight the set up of a specific parameter's panel for elite athletes. Moreover, we also intend to consider additional biomarkers, still under investigation, which could further contribute to laboratory sports medicine and provide reliable data that can be used by athlete's competent staff in order to establish personal attitudes and prevent sports injuries.

Biochemical and molecular characterization of 3-Methylcrotonylglycinuria in an Italian asymptomatic girl.

3-Methylcrotonylglycinuria is an organic aciduria resulting from deficiency of 3-methylcrotonyl-CoA carboxylase (3-MCC), a biotin-dependent mitochondrial enzym carboxylating 3-methylcrotonyl-CoA to 3-methylglutaconyl-CoA during leucine catabolism. Its deficiency, due to mutations on MCCC1 and MCCC2 genes, leads to accumulation of 3-methylcrotonyl-CoA metabolites in blood and/or urine, primarily 3-hydroxyisovaleryl-carnitine (C5-OH) in plasma and 3-methylcrotonyl-glycine (3-MCG) and 3-hydroxyisovaleric acid (3-HIVA) in the urine. The phenotype of 3-MCC deficiency is highly variable, ranging from severe neurological abnormalities and death in infancy to asymptomatic adults. Here we report the biochemical and molecular characterization of an Italian asymptomatic girl, positive for the newborn screening test. Molecular analysis showed two mutations in the MCCC2 gene, an already described missense mutation, c.691A > T (p.I231F), and a novel splicing mutation, c.1150-1G > A. We characterized the expression profile of the splice mutation by functional studies.

A rare case of sterol-C4-methyl oxidase deficiency in a young Italian male: Biochemical and molecular characterization.

Inborn defects of cholesterol biosynthesis are metabolic disorders presenting with multi-organ and tissue anomalies. An autosomal recessive defect involving the demethylating enzyme C4-methyl sterol (SC4MOL) has been reported in only 4 patients so far. In infancy, all patients were affected by microcephaly, bilateral congenital cataracts, growth delay, psoriasiform dermatitis, immune dysfunction, and intellectual disability. Herein, we describe a new case of SC4MOL deficiency in which a 19-year-old Italian male was affected by bilateral congenital cataracts, growth delay and learning disabilities, behavioral disorders and small stature, but not microcephaly. Our patient had abundant scalp dandruff, without other skin manifestations. Analysis of the blood sterol profile showed accumulation of C4-monomethyl and C4-dimethyl sterols suggesting a deficiency of the SC4MOL enzyme. Sequencing of the MSMO1 gene (also known as the "SC4MOL" gene) confirmed mutations in each allele (c.731A>G, p.Y244C, which is already known, and c.605G>A, p.G202E, which is a novel variant). His father carried c.731A>G mutation, whereas his mother carried c.605G>A. Thus, the combination of multiple skills and methodologies, in particular, blood sterol profiling and genetic analysis, led to the diagnosis of a new case of a very rare defect of cholesterol biosynthesis. Consequently, we suggest that these two analyses should be performed as soon as possible in all undiagnosed patients affected by bilateral cataracts and developmental delay.

Allelic Complexity in Long QT Syndrome: A Family-Case Study.

Congenital long QT syndrome (LQTS) is associated with high genetic and allelic heterogeneity. In some cases, more than one genetic variant is identified in the same (compound heterozygosity) or different (digenic heterozygosity) genes, and subjects with multiple pathogenic mutations may have a more severe disease. Standard-of-care clinical genetic testing for this and other arrhythmia susceptibility syndromes improves the identification of complex genotypes. Therefore, it is important to distinguish between pathogenic mutations and benign rare variants. We identified four genetic variants (KCNQ1-p.R583H, KCNH2-p.C108Y, KCNH2-p.K897T, and KCNE1-p.G38S) in an LQTS family. On the basis of in silico analysis, clinical data from our family, and the evidence from previous studies, we analyzed two mutated channels, KCNQ1-p.R583H and KCNH2-p.C108Y, using the whole-cell patch clamp technique. We found that KCNQ1-p.R583H was not associated with a severe functional impairment, whereas KCNH2-p.C108Y, a novel variant, encoded a non-functional channel that exerts dominant-negative effects on the wild-type. Notably, the common variants KCNH2-p.K897T and KCNE1-p.G38S were previously reported to produce more severe phenotypes when combined with disease-causing alleles. Our results indicate that the novel KCNH2-C108Y variant can be a pathogenic LQTS mutation, whereas KCNQ1-p.R583H, KCNH2-p.K897T, and KCNE1-p.G38S could be LQTS modifiers.

The multi-faceted aspects of the complex cardiac Nav1.5 protein in membrane function and pathophysiology.

The aim of this mini-review is to draw together the main concepts and findings that have emerged from recent studies of the cardiac channel protein Nav1.5. This complex protein is encoded by the SCN5A gene that, in its mutated form, is implicated in various diseases, particularly channelopathies, specifically at cardiac tissue level. Here we describe the structural, and functional aspects of Nav1.5 including post-translational modifications in normal conditions, and the main human channelopathies in which this protein may be the cause or trigger. Lastly, we also briefly discuss interacting proteins that are relevant for these channel functions in normal and disease conditions.

Mulibrey nanism: Two novel mutations in a child identified by Array CGH and DNA sequencing.

In childhood, several rare genetic diseases have overlapping symptoms and signs, including those regarding growth alterations, thus the differential diagnosis is sometimes difficult. The proband, aged 3 years, was suspected to have Silver-Russel syndrome because of intrauterine growth retardation, postnatal growth retardation, typical facial dysmorphic features, macrocephaly, body asymmetry, and bilateral fifth finger clinodactyly. Other features were left atrial and ventricular enlargement and patent foramen ovale. Total X-ray skeleton showed hypoplasia of the twelfth rib bilaterally and of the coccyx, slender long bones with thick cortex, and narrow medullary channels. The genetic investigation did not confirm Silver-Russel syndrome. At the age of 5 the patient developed an additional sign: hepatomegaly. Array CGH revealed a 147 kb deletion (involving TRIM 37 and SKA2 genes) on one allele of chromosome 17, inherited from his mother. These results suggested Mulibrey nanism. The clinical features were found to fit this hypothesis. Sequencing of the TRIM 37 gene showed a single base change at a splicing locus, inherited from his father that provoked a truncated protein. The combined use of Array CGH and DNA sequencing confirmed diagnosis of Mulibrey nanism. The large deletion involving the SKA2 gene, along with the increased frequency of malignant tumours in mulibrey patients, suggests closed monitoring for cancer of our patient and his mother. Array CGH should be performed as first tier test in all infants with multiple anomalies. The clinician should reconsider the clinical features when the genetics suggests this. © 2016 Wiley Periodicals, Inc.