Search

ObjectivesThis study aims to evaluate in a real-life Italian multicenter cohort of axial spondyloarthritis (axSpA) (1) the 4-year effectiveness and safety of secukinumab, (2) the drug retention rate (DRR), and (3) the impact of the line of bDMARDs treatment, subtype of axSpA, and sex on achieving low disease activity (LDA) and very low disease activity (VLDA).MethodsConsecutive axSpA patients receiving secukinumab between 2016 and 2023 were prospectively evaluated. Data on disease characteristics, previous/ongoing treatments, comorbidities, and follow-up duration were collected. Treatment response was evaluated at 6 and 12 months after initiation and yearly up to 48 months (T48). DRR and effectiveness outcomes were evaluated according to bDMARDs treatment, axSpA subtype, and sex. Infections and adverse events (AEs) were recorded.ResultsWe enrolled 272 patients (48.2% male; median age, 51; 39.7% HLA-B27+; 40.4% nr-axSpA), of whom 30.9% were naïve to secukinumab. Overall, secukinumab yielded improvement in effectiveness outcomes; the naïve patients maintained lower disease activity vs. the non-naïve ones. At T48, the LDA and VLDA rates were higher in naïve patients and in male individuals. Treatment was discontinued in 104 patients due to primary/secondary loss of effectiveness and in 34 patients due to AEs. The DRR at T48 was 67.4% in the whole population, regardless of treatment line, axSpA subtype, and sex.ConclusionsSecukinumab was safe and effective in all axSpA patients irrespective of treatment line, disease subtype, and sex. The patients achieved sustained 4-year remission and DRR.

Adult T-cell leukemia/lymphoma (ATL) is a T-cell lymphoproliferative neoplasm caused by the human T-cell leukemia virus type 1 (HTLV-1). Two viral proteins, Tax-1 and HBZ play important roles in HTLV-1 infectivity and in HTLV-1-associated pathologies by altering key pathways of cell homeostasis. However, the molecular mechanisms through which the two viral proteins, particularly HBZ, induce and/or sustain the oncogenic process are still largely elusive. Previous results suggested that HBZ interaction with nuclear factors may alter cell cycle and cell proliferation. To have a more complete picture of the HBZ interactions, we investigated in detail the endogenous HBZ interactome in leukemic cells by immunoprecipitating the HBZ-interacting complexes of ATL-2 leukemic cells, followed by tandem mass spectrometry analyses. RNA seq analysis was performed to decipher the differential gene expression and splicing modifications related to HTLV-1. Here we compared ATL-2 with MOLT-4, a non HTLV-1 derived leukemic T cell line and further compared with HBZ-induced modifications in an isogenic system composed by Jurkat T cells and stably HBZ transfected Jurkat derivatives. The endogenous HBZ interactome of ATL-2 cells identified 249 interactors covering three main clusters corresponding to protein families mainly involved in mRNA splicing, nonsense-mediated RNA decay (NMD) and JAK-STAT signaling pathway. Here we analyzed in detail the cluster involved in RNA splicing. RNAseq analysis showed that HBZ specifically altered the transcription of many genes, including crucial oncogenes, by affecting different splicing events. Consistently, the two RNA helicases, members of the RNA splicing family, DDX5 and its paralog DDX17, recently shown to be involved in alternative splicing of cellular genes after NF-κB activation by HTLV-1 Tax-1, interacted and partially co-localized with HBZ. For the first time, a complete picture of the endogenous HBZ interactome was elucidated. The wide interaction of HBZ with molecules involved in RNA splicing and the subsequent transcriptome alteration strongly suggests an unprecedented complex role of the viral oncogene in the establishment of the leukemic state.

The 16p13.11 microdeletion, whose prevalence in the general population is about 0.04%, is known in literature as a predisposition factor to neurodevelopmental disorders, being found in about 0.13% of patients with schizophrenia, in 0.5–0.6% of patient with epilepsy, cognitive impairment, autism spectrum disorder (ASD) and aggressiveness. The goal of this study was to identify a specific gene set pattern unique for the affected patients in comparison with other familial components. Due to the incomplete penetrance of this copy number variant (CNV), we studied by whole exome sequencing (WES), with particular regard of 850 SFARI genes, three families with an affected member carrier of inherited 16p13.11 and 16p13.11p12.3 microdeletion and one family with an affected member with a de novo 16p13.11 microdeletion. By combining a deductive approach together with personalized network models, we identified gene signatures potentially capable of explaining the clinical phenotype. Candidate variants in genes of interest were identified as possibly involved in determining the neurological phenotype of the four patients, such as compound heterozygosity in CECR2, variants in MTOR and RICTOR genes, compound heterozygous single nucleotide variants in the LRRK2 gene. Moreover, genes present in the microdeletion region were partially present as central nodes, with a focus on NDE1. No additional pathogenetic or uncertain CNVs were found in all four patients. No significant variants were detected in genes included in the microdeletion in patients 1, 2 and 3, excluding the finding of unmasked recessive variants. In conclusion, WES is a fundamental tool in the genetic investigation of patients having a predisposing variant, which is not sufficient to define the clinical phenotype. Moreover, the analysis of WES data using Systems medicine tools, such as personalized network models, led to the prioritization of genes on a high throughput scale and to discover variants in genes that were not prioritized at first.

Different strategies have been developed to help parents with introducing new or disliked vegetables. Nonetheless, many parents of preschoolers struggle against children's refusal to eat vegetables. In this study, we aimed to evaluate the effectiveness of e-books in promoting positive attitudes toward vegetables through repeated visual exposures. A total of 61 families with preschoolers joined the See & Eat study and received an e-book about one of two vegetables chosen from a list of 24. Parents provided ratings of children's willingness to taste, intake, and liking of the chosen vegetables before and after reading the e-book; parents also evaluated their children's food fussiness and their agreement with respect to three mealtime goals of the family. Using a 2 (vegetable: target or non-target) × 2 (time: pre-test or post-test) within-subjects analysis, results from 53 families revealed a significant increase in children's willingness to taste, intake, and liking at post-test of both target and non-target vegetables. Following a two-week parent-child e-book reading intervention, children's food fussiness and parents' endorsement of positive mealtime goals slightly but significantly increased. Results suggest that e-books are effective in encouraging healthy eating among preschoolers and that the positive effect of e-book reading can generalize to other vegetables.

Gene set enrichment analysis (GSEA) is a powerful tool to associate a disease phenotype to a group of genes/proteins. GSEA attributes a specific weight to each gene/protein in the input list that depends on a metric of choice, which is usually represented by quantitative expression data. However, expression data are not always available. Here, GSEA based on betweenness centrality of a protein–protein interaction (PPI) network is described and applied to two cases, where an expression metric is missing. First, personalized PPI networks were generated from genes displaying alterations (assessed by array comparative genomic hybridization and whole exome sequencing) in four probands bearing a 16p13.11 microdeletion in common and several other point variants. Patients showed disease phenotypes linked to neurodevelopment. All networks were assembled around a cluster of first interactors of altered genes with high betweenness centrality. All four clusters included genes known to be involved in neurodevelopmental disorders with different centrality. Moreover, the GSEA results pointed out to the evidence of “cell cycle” among enriched pathways. Second, a large interaction network obtained by merging proteomics studies on three neurodegenerative disorders was analyzed from the topological point of view. We observed that most central proteins are often linked to Parkinson’s disease. The selection of these proteins improved the specificity of GSEA, with “Metabolism of amino acids and derivatives” and “Cellular response to stress or external stimuli” as top-ranked enriched pathways. In conclusion, betweenness centrality revealed to be a suitable metric for GSEA. Thus, centrality-based GSEA represents an opportunity for precision medicine and network medicine.

Mutations in PARK2 gene are the most frequent cause of familial forms of Parkinson’s disease (PD). This gene encodes Parkin, an E3 ubiquitin ligase involved in several cellular mechanisms, including mitophagy. Parkin loss-of-function is responsible for the cellular accumulation of damaged mitochondria, which in turn determines an increment of reactive oxygen species (ROS) levels, lower ATP production, and apoptosis activation. Given the importance of mitochondrial dysfunction and mitophagy impairment in PD pathogenesis, the aim of the present study was to investigate both total and mitochondrial proteome alterations in human skin fibroblasts of PARK2-mutated patients. To this end, both total and mitochondria-enriched protein fractions from fibroblasts of five PARK2-mutated patients and five control subjects were analyzed by quantitative shotgun proteomics to identify proteins specifically altered by Parkin mutations (mass spectrometry proteomics data have been submitted to ProteomeXchange with the identifier PXD015880). Both the network-based and gene set enrichment analyses pointed out pathways in which Rab GTPase proteins are involved. To have a more comprehensive view of the mitochondrial alterations due to PARK2 mutations, we investigated the impact of Parkin loss on mitochondrial function and network morphology. We unveiled that the mitochondrial membrane potential was reduced in PARK2-mutated patients, without inducing PINK1 accumulation, even when triggered with the ionophore carbonyl cyanide m-chlorophenylhydrazone (CCCP). Lastly, the analysis of the mitochondrial network morphology did not reveal any significant alterations in PARK2-mutated patients compared to control subjects. Thus, our results suggested that the network morphology was not influenced by the mitochondrial depolarization and by the lack of Parkin, revealing a possible impairment of fission and, more in general, of mitochondrial dynamics. In conclusion, the present work highlighted new molecular factors and pathways altered by PARK2 mutations, which will unravel possible biochemical pathways altered in the sporadic form of PD.

Parkinson’s disease (PD) is the second most frequent neurodegenerative disease worldwide and the availability of early biomarkers and novel biotargets represents an urgent medical need. The main pathogenetic hallmark of PD is the specific loss of nigral dopaminergic neurons, in which mitochondrial dysfunction plays a crucial role. Mitochondrial proteases are central to the maintenance of healthy mitochondria and they have recently emerged as drug targets. However, an exhaustive characterization of these enzymes and their targets is still lacking, due to difficulties in analyzing proteolytic fragments by bottom-up proteomics approaches. Here, we propose the “mitochondrial dimethylation-TAILS” strategy, which combines the isolation of mitochondria with the enrichment of N-terminal peptides to analyze the mitochondrial N-terminome. We applied this method in a cellular model of altered dopamine homeostasis in neuroblastoma SH-SY5Y cells, which recapitulates early steps of PD pathogenesis. The main aim was to identify candidate mitochondrial proteases aberrantly activated by dopamine dysregulation and their cleaved targets. The proposed degradomics workflow was able to improve the identification of mitochondrial proteins if compared to classical shotgun analysis. In detail, 40% coverage of the mitochondrial proteome was obtained, the sequences of the transit peptides of two mitochondrial proteins were unveiled, and a consensus cleavage sequence for proteases involved in the processing of mitochondrial proteins was depicted. Mass spectrometry proteomics data have been submitted to ProteomeXchange with the identifier PXD013900. Moreover, sixty-one N-terminal peptides whose levels were affected by dopamine treatment were identified. By an in-depth analysis of the proteolytic peptides included in this list, eleven mitochondrial proteins showed altered proteolytic processing. One of these proteins (i.e., the 39S ribosomal protein L49 – MRPL49) was cleaved by the neprilysin protease, already exploited in clinics as a biotarget. We eventually demonstrated a mitochondrial subcellular localization of neprilysin in human cells for the first time. Collectively, these results shed new light on mitochondrial dysfunction linked to dopamine imbalance in PD and opened up the possibility to explore the mitochondrial targets of neprilysin as candidate biomarkers.

Background: Sleep-Disordered Breathing (SDB) varies from habitual snoring to partial or complete obstruction of the upper airway and can be found in up to 10% of children. SDB can significantly affect children’s wellbeing, as it can cause growth disorders, educational and behavioral problems, and even life-threatening conditions, such as cardiorespiratory failure. Adenotonsillectomy represents the primary treatment for pediatric SDB where adeno-tonsillar hypertrophy is indicated. For those with craniofacial anomalies, or for whom adenotonsillectomy or other treatment modalities have failed, or surgery is contra-indicated, mandibular advancement splints (MAS) may represent a viable treatment option. Whilst the efficacy of these appliances has been consistently demonstrated in adults, there is little information about their effectiveness in children.Aims: To determine the efficacy of mandibular advancement appliances for the management of SDB and related health problems in children. Methods/design: The study will be designed as a single-blind crossover randomized controlled trial with administration of both an ‘Active MAS’ (Twin-block) and a ‘Sham MAS’. Eligible participants will be children aged 8 to 12 years whose parents report they snore ≥ 3 nights per week. Sixteen children will enter the full study after confirming other inclusion criteria, particularly Skeletal class I or class II confirmed by lateral cephalometric radiograph. Each child will be randomly assigned to either a treatment sequence starting with the Active or the Sham MAS. Participants will wear the appliances for three weeks separated by a two-week washout period. For each participant, home-based polysomnographic data will be collected four times; once before and once after each treatment period. The Apnea Hypopnea Index (AHI) will represent the main outcome variable. Secondary outcomes will include, snoring frequency, masseter muscle activity, sleep symptoms, quality of life, daytime sleepiness, children behavior, and nocturnal enuresis. In addition, blood samples will be collected to assess growth hormone changes.Trial registration: This study was registered in the Australian New Zealand Clinical Trials Registry (ANZCTR): [ACTRN12614001013651]