Preparation of cardiac extracts from embryonal hearts to capture RNA–protein interactions by CLIP

The interaction of RNA with specific RNA-binding proteins (RBP) leads to the establishment of complex regulatory networks through which gene expression is controlled. Careful consideration should be given to the exact environment where a given RNA/RBP interplay occurs, as the functional responses might depend on the type of organism as well as the specific cellular or subcellular contexts. This requisite becomes particularly crucial for the study of long non-coding RNAs (lncRNA), as a consequence of their peculiar tissue-specificity and timely regulated expression. The functional characterization of lncRNAs has traditionally relied on the use of established cell lines that, although useful, are unable to fully recapitulate the complexity of a tissue or organ. Here, we detail an optimized protocol, with comments and tips, to identify the RNA interactome of given RBPs by performing cross-linking immunoprecipitation (CLIP) from mouse embryonal hearts. We tested the efficiency of this protocol on the murine pCharme, a muscle-specific lncRNA interacting with Matrin3 (MATR3) and forming RNA-enriched condensates of biological significance in the nucleus

Epigenetic regulation of Wnt7b expression by the cis-acting long noncoding RNA Lnc-Rewind in muscle stem cells

Skeletal muscle possesses an outstanding capacity to regenerate upon injury due to the adult muscle stem cells (MuSCs) activity. This ability requires the proper balance between MuSCs expansion and differentiation which is critical for muscle homeostasis and contributes, if deregulated, to muscle diseases. Here, we functionally characterize a novel chromatin-associated lncRNA, Lnc-Rewind, which is expressed in murine MuSCs and conserved in human. We find that, in mouse, Lnc-Rewind acts as an epigenetic regulator of MuSCs proliferation and expansion by influencing the expression of skeletal muscle genes and several components of the WNT (Wingless-INT) signalling pathway. Among them, we identified the nearby Wnt7b gene as a direct Lnc-Rewind target. We show that Lnc-Rewind interacts with the G9a histone lysine methyltransferase and mediates the in cis repression of Wnt7b by H3K9me2 deposition. Overall, these findings provide novel insights into the epigenetic regulation of adult muscle stem cells fate by lncRNAs

Epigenetic remodelling of Fxyd1 promoters in developing heart and brain tissues

FXYD1 is a key protein controlling ion channel transport. FXYD1 exerts its function by regulating Na+/K+-ATPase activity, mainly in brain and cardiac tissues. Alterations of the expression level of the FXYD1 protein cause diastolic dysfunction and arrhythmias in heart and decreased neuronal dendritic tree and spine formation in brain. Moreover, FXYD1, a target of MeCP2, plays a crucial role in the pathogenesis of the Rett syndrome, a neurodevelopmental disorder. Thus, the amount of FXYD1 must be strictly controlled in a tissue specific manner and, likely, during development. Epigenetic modifications, particularly DNA methylation, represent the major candidate mechanism that may regulate Fxyd1 expression. In the present study, we performed a comprehensive DNA methylation analysis and mRNA expression level measurement of the two Fxyd1 transcripts, Fxyd1a and Fxyd1b, in brain and heart tissues during mouse development. We found that DNA methylation at Fxyd1a increased during brain development and decreased during heart development along with coherent changes in mRNA expression levels. We also applied ultra-deep methylation analysis to detect cell to cell methylation differences and to identify possible distinct methylation profile (epialleles) distribution between heart and brain and in different developmental stages. Our data indicate that the expression of Fxyd1 transcript isoforms inversely correlates with DNA methylation in developing brain and cardiac tissues suggesting the existence of a temporal-specific epigenetic program. Moreover, we identified a clear remodeling of epiallele profiles which were distinctive for single developmental stage both in brain and heart tissues

ARIA digital anamorphosis : Digital transformation of health and care in airway diseases from research to practice

Digital anamorphosis is used to define a distorted image of health and care that may be viewed correctly using digital tools and strategies. MASK digital anamorphosis represents the process used by MASK to develop the digital transformation of health and care in rhinitis. It strengthens the ARIA change management strategy in the prevention and management of airway disease. The MASK strategy is based on validated digital tools. Using the MASK digital tool and the CARAT online enhanced clinical framework, solutions for practical steps of digital enhancement of care are proposed.Peer reviewe

Adherence to treatment in allergic rhinitis using mobile technology. The MASK Study

Background: Mobile technology may help to better understand the adherence to treatment. MASK-rhinitis (Mobile Airways Sentinel NetworK for allergic rhinitis) is a patient-centred ICT system. A mobile phone app (the Allergy Diary) central to MASK is available in 22 countries. Objectives: To assess the adherence to treatment in allergic rhinitis patients using the Allergy Diary App. Methods: An observational cross-sectional study was carried out on all users who filled in the Allergy Diary from 1 January 2016 to 1 August 2017. Secondary adherence was assessed by using the modified Medication Possession Ratio (MPR) and the Proportion of days covered (PDC) approach. Results: A total of 12143 users were registered. A total of 6949 users reported at least one VAS data recording. Among them, 1887 users reported >= 7 VAS data. About 1195 subjects were included in the analysis of adherence. One hundred and thirty-six (11.28%) users were adherent (MPR >= 70% and PDC = 70% and PDC = 1.50) and 176 (14.60%) were switchers. On the other hand, 832 (69.05%) users were non-adherent to medications (MPR Conclusion and clinical relevance: Adherence to treatment is low. The relative efficacy of continuous vs on-demand treatment for allergic rhinitis symptoms is still a matter of debate. This study shows an approach for measuring retrospective adherence based on a mobile app. This also represents a novel approach for analysing medication-taking behaviour in a real-world setting.Peer reviewe

Allergic Rhinitis and its Impact on Asthma (ARIA) Phase 4 (2018) : Change management in allergic rhinitis and asthma multimorbidity using mobile technology

Allergic Rhinitis and its Impact on Asthma (ARIA) has evolved from a guideline by using the best approach to integrated care pathways using mobile technology in patients with allergic rhinitis (AR) and asthma multimorbidity. The proposed next phase of ARIA is change management, with the aim of providing an active and healthy life to patients with rhinitis and to those with asthma multimorbidity across the lifecycle irrespective of their sex or socioeconomic status to reduce health and social inequities incurred by the disease. ARIA has followed the 8-step model of Kotter to assess and implement the effect of rhinitis on asthma multimorbidity and to propose multimorbid guidelines. A second change management strategy is proposed by ARIA Phase 4 to increase self-medication and shared decision making in rhinitis and asthma multimorbidity. An innovation of ARIA has been the development and validation of information technology evidence-based tools (Mobile Airways Sentinel Network [MASK]) that can inform patient decisions on the basis of a self-care plan proposed by the health care professional.Peer reviewe

ARIA digital anamorphosis: Digital transformation of health and care in airway diseases from research to practice

Digital anamorphosis is used to define a distorted image of health and care that may be viewed correctly using digital tools and strategies. MASK digital anamorphosis represents the process used by MASK to develop the digital transformation of health and care in rhinitis. It strengthens the ARIA change management strategy in the prevention and management of airway disease. The MASK strategy is based on validated digital tools. Using the MASK digital tool and the CARAT online enhanced clinical framework, solutions for practical steps of digital enhancement of care are proposed

Muscle regeneration and RNA: new perspectives for ancient molecules

The ability of the ribonucleic acid (RNA) to self-replicate, combined with a unique cocktail of chemical properties, suggested the existence of an RNA world at the origin of life. Nowadays, this hypothesis is supported by innovative high-throughput and biochemical approaches, which definitively revealed the essential contribution of RNA-mediated mechanisms to the regulation of fundamental processes of life. With the recent development of SARS-CoV-2 mRNA-based vaccines, the potential of RNA as a therapeutic tool has received public attention. Due to its intrinsic singlestranded nature and the ease with which it is synthesized in vitro, RNA indeed represents the most suitable tool for the development of drugs encompassing every type of human pathology. The maximum effectiveness and biochemical versatility is achieved in the guise of non-coding RNAs (ncRNAs), which are emerging as multifaceted regulators of tissue specification and homeostasis. Here, we report examples of coding and ncRNAs involved in muscle regeneration and discuss their potential as therapeutic tools. Small ncRNAs, such as miRNA and siRNA, have been successfully applied in the treatment of several diseases. The use of longer molecules, such as lncRNA and circRNA, is less advanced. However, based on the peculiar properties discussed below, they represent an innovative pool of RNA biomarkers and possible targets of clinical value

Non-coding regulation of myogenic chromatin

Advances in sequencing technologies have revealed that more than half of the transcriptome encodes for several classes of RNA molecules lacking coding capacity. The impact of these species on the control of cell growth, differentiation and development, has been well established, and it is clear that they exert their functions in both nuclear and cytoplasmic compartments (Ballarino et al, JCI, 2015). Through RNA-sequencing of murine myoblasts and differentiated myotubes, we recently identified novel long non-coding RNAs (lncRNAs) which expression is dynamically ordered during myogenesis (Ballarino et al, MCB, 2015). These transcripts were classified on the basis of their expression in proliferating versus differentiated conditions, muscle-restricted activation and link with muscle dystrophy. I will describe the characterization of a murine chromatin associated muscle-specific long noncoding RNA, named as Charme (for Chromatin architect of muscle expression), which contributes to the robustness of the myogenic programme both in vitro and in vivo. In myocytes, Charme knock-down produced the disassembly of specific chromosomal domains and the down-regulation of myogenic genes therein contained. Notably, several Charme-affected genes are associated to human cardiomyopathies, and Charme depletion in mice produces a pathologic phenotype consisting in a global alteration of heart architecture. The existence of a homologous human counterpart, which regulates the same subset of genes, suggests an important and evolutionary conserved function for Charme. Altogether, our study identifies a novel example of chromatin-associated lncRNA involved in the acquisition of muscle cell identit

Safetycracy e regime terapeutico. Le politiche di sospensione delle soggettività

La safetycracy è una forma dello stato di eccezione e un paradigma del potere giustificato da una grave emergenza sanitaria, fondato sulla protezione della vita quale mera sopravvivenza biologica come priorità assoluta rispetto ad ogni altro valore o diritto, e sull’uso strumentale della scienza in campo medico e delle tecnologie di controllo, che mettono i corpi dei cittadini al centro della decisione e dell’azione politica, che diviene così a tutti gli effetti biopolitica. Nell’emergenza Covid-19 in Italia ha avuto attuazione in due diverse politiche: in quella di safetycracy in senso stretto nel 2020 e in quella di regime terapeutico nel biennio 2021-22. La prima è messa in atto quando il potere dichiara di non disporre della soluzione idonea a risolvere l’emergenza sanitaria, e prevede la sospensione della generalità delle soggettività, fatta eccezione per quelle che di chi è direttamente coinvolto o è ritenuto idoneo a dare un proprio contributo contro la minaccia alla salute pubblica. La seconda è attuata quando le autorità dichiarano di disporre della soluzione appropriata a porre termine all’emergenza sanitaria, che consiste in uno specifico trattamento terapeutico o profilattico ritenuto adeguato allo scopo, il quale viene “fortemente consigliato” con un vasto repertorio di strumenti comunicativi e persuasivi e meccanismi psico-sociali e, per alcune categorie, normativamente imposto. In tal caso, la sospensione della soggettività è selettiva e limitata a quella delle sole persone che non intendono conformarsi alle pressioni del potere. Le politiche in questione sono il filo conduttore di questa raccolta di saggi e rapporti di ricerca, alcuni del curatore ed altri tratti dalle tesi di laureandi in Organizzazione e Marketing per la Comunicazione d’Impresa redatte nell’ambito del Laboratorio Rimoves.com del Dipartimento di Comunicazione e Ricerca Sociale dell’Università di Roma “Sapienza” negli AA 2020-21 e 2021-22