A Combined Targeted and Whole Exome Sequencing Approach Identified Novel Candidate Genes Involved in Heritable Pulmonary Arterial Hypertension

The pathogenesis of idiopathic and heritable forms of pulmonary arterial hypertension is still not completely understood, even though several causative genes have been proposed, so that a third of patients remains genetically unresolved. Here we applied a multistep approach to extend identification of the genetic bases of such a disease by searching for novel candidate genes/pathways. Twenty-eight patients belonging to 18 families were screened for BMPR2 mutations and BMPR2-negative samples were tested for 12 additional candidate genes by means of a specific massive parallel sequencing-based assay. Finally, whole exome sequencing was performed on four patients showing no mutations at known disease genes, as well as on their unaffected parents. In addition to EIF2AK4, which has been already suggested to be associated with pulmonary veno-occlusive disease, we identified the novel candidate genes ATP13A3, CD248, EFCAB4B, involved in lung vascular remodeling that represent reliable drivers contributing to the disease according to their biological functions/inheritance patterns. Therefore, our results suggest that combining gene panel and whole exome sequencing provides new insights useful for the genetic diagnosis of familial and idiopathic pulmonary arterial hypertension, as well as for the identification of biological pathways that will be potentially targeted by new therapeutic strategies

Disease status, reasons for discontinuation and adverse events in 1038 Italian children with juvenile idiopathic arthritis treated with etanercept

Background: Data from routine clinical practice are needed to further define the efficacy and safety of biologic medications in children with juvenile idiopathic arthritis (JIA). The aim of this analysis was to investigate the disease status, reasons for discontinuation and adverse events in Italian JIA patients treated with etanercept (ETN). Methods: In 2013, all centers of the Italian Pediatric Rheumatology Study Group were asked to make a census of patients given ETN after January 2000. Patients were classified in three groups: group 1 = patients still taking ETN; group 2 = patients discontinued from ETN for any reasons; group 3 = patients lost to follow-up while receiving ETN. All three groups received a retrospective assessment; patients in group 1 also underwent a cross-sectional assessment. Results: 1038 patients were enrolled by 23 centers: 422 (40.7%) were in group 1, 462 (44.5%) in group 2, and 154 (14.8%) in group 3. Median duration of ETN therapy was 2.5 years. At cross-sectional assessment, 41.8% to 48.6% of patients in group 1 met formal criteria for inactive disease, whereas 52.4% of patients in group 2 and 55.8% of patients in group 3 were judged in clinical remission by their caring physician at last visit. A relatively greater proportion of patients with systemic arthritis were discontinued or lost to follow-up. Parent evaluations at cross-sectional visit in group 1 showed that 52.4% of patients had normal physical function, very few had impairment in quality of life, 51.2% had no pain, 76% had no morning stiffness, and 82.7% of parents were satisfied with their child's illness outcome. Clinically significant adverse events were reported for 27.8% of patients and ETN was discontinued for side effects in 9.5%. The most common adverse events were new onset or recurrent uveitis (10.2%), infections (6.6%), injection site reactions (4.4%), and neuropsychiatric (3.1%), gastrointestinal (2.4%), and hematological disorders (2.1%). Ten patients developed an inflammatory bowel disease and 2 had a malignancy. One patient died of a fulminant streptococcal sepsis. Conclusions: Around half of the patients achieved complete disease quiescence under treatment with ETN. The medication was overall well tolerated, as only one quarter of patients experienced clinically significant adverse events and less than 10% had treatment discontinued for toxicity

Validation of reference genes for qRT-PCR analysis in Megoura viciae (Hemiptera Aphididae)

Quantitative reverse transcription polymerase chain reaction (qRT-PCR) has become one of the most sensitive methods to monitor gene expression. An important and often neglected requirement of this kind of study is the validation of appropriate reference genes with the most possible stable expression levels across samples groups. In this paper, several candidates were tested in all the four nymphal instars and the adult morphs (winged and apterous) of aphid Megoura viciae Buckton (Hemiptera Aphididae), an important pest of broad bean, in order to obtain reference genes for future works on Megoura viciae gene expression. Since the use of multiple reference genes is recommended for an accurate normalization, eight candidate genes were tested, encoding respectively for: ribosomal protein L32 (RPL32), NADH dehydrogenase (ubiquinone) flavoprotein 1 (NADH), succinate dehydrogenase complex subunit A (SUCC), ribosomal protein S9 (RPS9), TATA-box binding protein (TATA), actin (ACT), β-tubulin (TBU) and ubiquitin-conjugating protein (UBIQ). Three software programs and the comparative ΔCT method were used to compare and rank the candidate genes and RefFinder - a web-based comprehensive tool that integrates all the four methods in a single index - indicated RPS9-RPL32 as the best couple. In addition, our study showed that a common-used reference gene, β-actin, achieved the worst score among our candidates. Finally, differences between the “classic” normalization with β-actin as a reference gene and the normalization using the best reference genes according to our work were highlighted using for the first time a Megoura viciae odorant binding protein, OBP4, as the target gene

Effects of cardiac resynchronization therapy on right ventricular function during rest and exercise, as assessed by radionuclide angiography, and on NT-proBNP levels

Aim: We carried out this study to investigate mid-term effects of cardiac resynchronization therapy (CRT) on right ventricular (RV) function and neurohormonal response, expressed by N-terminal pro-brain natriuretic peptide (NT-proBNP), in heart failure patients stratified by baseline RV ejection fraction (RVEF). Methods and Results: Thirty-six patients with nonischemic dilated cardiomyopathy underwent technetium-99m radionuclide angiography with bicycle exercise immediately after CRT implantation (during spontaneous rhythm and after CRT activation) and 3 months later. Plasma NT proBNP was assessed before implantation and after 3 months. At baseline, RVEF was impaired ( 6435%) in 14 patients, preserved (>35%) in 22. At 3 months, RVEF improved during rest and exercise (P = .02) in patients with impaired RV function, while remaining unchanged in patients with preserved RV function. Rest and exercise RV dyssynchrony decreased in both groups at follow-up (P < .05). A similar mid-term improvement in left ventricular (LV) function and NT-proBNP was observed in patients with impaired and preserved RVEF. In the former, the decrease in NT-proBNP correlated with the improvements both in LV and RV dyssynchrony and functions. Conclusion: CRT may improve RV performance, during rest and exercise, and neurohormonal response in heart failure patients with nonischemic dilated cardiomyopathy and baseline RV dysfunction. RV dysfunction should not be considered per se a primary criterion for excluding candidacy to CRT

In vitro thrombogenicity of drug-eluting and bare metal stents

Aims: We sought to investigate the thrombogenicity of different DES and BMS in an in vitro system of stent perfusion. Material and methods: The experimental model consisted of a peristaltic pump connected to 4 parallel silicone tubes in which different stents were deployed. Blood was drawn from healthy volunteers and the amount of stent surfaced-induced thrombus deposition was determined using 125I-fibrinogen. Results: Compared to Resolute, Biomatrix and Vision, Xience was associated with the lowest amount of stent surface-induced thrombus formation, with a significant difference compared to Vision (125I-fibrinogen median value deposition [IQ range]: 50 ng [25–98] versus 560 ng [320–1520], respectively, p < 0.05), but not to other DES. In the second set of experiments Fluoropolymer-coated BMS not eluting drug was associated with a significant 3-fold reduction in 125I-fibrinogen deposition (245 ng [80–300]) compared to Vision (625 ng [320–760], p < 0.05), but a 7-fold increase compared to Xience (35 ng [20–60], p < 0.05). Finally Xience was associated with a significantly greater absorption of albumin compared to BMS. Conclusions: In an in vitro system of stent perfusion, Xience was associated with the lowest amount of stent surface-induced thrombus formation compared with Resolute, Biomatrix and Vision, with a noted synergistic effect between the fluoropolymer and the drug

Gestione del Covid-19 in età pediatrica: documento di consenso

Coronavirus disease 2019 (Covid-19) caused by SARS-CoV-2 has rapidly spread, becoming the first pandemic of the 21st century by death toll. Children appear to be less affected than adults, with a milder clinical presentation and a significantly lower mortality rate. However, serious complica-tions can occur in childhood, such as Covid-19 temporally related multisystem inflammatory syn-drome (MIS-C). Some aspects of SARS-CoV-2 infection in children and adolescents remain un-clear and the optimal treatment has not been defined. The Working Group on Covid-19 in Paediatrics of the Emilia-Romagna Region (RE-CO-PED) has produced a consensus document with practical recommendations based on a systematic review of the literature and on the clinical experi-ence of the expert group. Evidence is reported regarding prevention measures, diagnostic tools as well as home and hospital therapeutic management of complicated cases (MIS-C). The educational and psychological effects of the pandemic in the paediatric and adolescent age are reported, with the need to define coordinated interventions (between paediatricians, neurospychiatrists, psycholo-gists and educational services) for the prevention and treatment of documented emotional, relational and educational consequences caused by the lockdown, school closures and social distances