Single-cell RNA analysis to investigate the role of phenotypic modulation of smooth muscle cells in the onset of atherosclerosis

Motivazione: l'aterosclerosi è una malattia infiammatoria cronica responsabile di quasi il 50% di tutti i decessi nei paesi occidentali. È caratterizzata dalla formazione di placche lipidiche nella parete arteriosa che riducono il lume, diminuiscono l'elasticità delle pareti e provocano molte malattie cardiovascolari. In risposta a segnali ambientali, le cellule muscolari lisce (SMC) dei vasi possono de-differenziarsi, proliferare e migrare in un processo noto come modulazione fenotipica. Recentemente è stato dimostrato che possono transdifferenziarsi in altri tipi cellulari che sono importanti per la formazione delle placche aterosclerotiche e che determinano la loro capacità di rompersi e indurre trombosi. Obiettivo: Lo scopo di questo lavoro è di utilizzare le più recenti tecnologie nel campo della trascrittomica, come l'scRNAseq e la trascrittomica spaziale, per identificare nuovi fenotipi derivati ​​dalle SMC indotti da un ambiente pro-aterosclerotico e per caratterizzare il loro ruolo nella patologia, ottenendo una firma genica specifica che aiuterà la loro identificazione nei pazienti e, conseguentemente, a bersagliarli nel contesto di un approccio di medicina di precisione. Metodi e risultati: Le cellule derivate dalle SMC dissociate dall'arco aortico e dall'arteria toracica di una coorte di topi knockout per ApoE prona a sviluppare l'aterosclerosi e di una di controllo, sono state analizzate con al tecnica del scRNAseq utilizzando la piattaforma Chromium della 10x Genomics. Successivamente è stata eseguita l'analisi bioinformatica dei dati individuando un nuovo cluster di cellule derivato dalle SMC, caratterizzato dall'espressione del gene Adamtsl1, non ancora descritto in letteratura. Inoltre, abbiamo scoperto che questo tipo cellulare è metabolicamente attivo, produce ECM, ha un fenotipo proinfiammatorio ed è uno stato di transizione tra le SMC contrattili e altri fenotipi cellulari il cui ruolo all'interno delle placche aterosclerotiche è già noto. Successivamente, la presenza di questo cluster è stata dimostrata anche in set di dati di scRNAseq umani e murini provenienti da un altro lavoro. Infine, due campioni aterosclerotici umani sono stati analizzati utilizzando la Visium Spatial transcriptomics (10x Genomics) evidenziando la presenza di questo fenotipo delle SMC all'interno delle placche umane. Conclusioni: Il tipo cellulare Adamtsl1+ derivato dalle SMC identificato potrebbe essere un buon bersaglio per bloccare la transdifferenziazione delle SMC verso i fenotipi coinvolti nella formazione della placca e per migliorare la prognosi dei pazienti con aterosclerosi.Rationale: Atherosclerosis is a chronic inflammatory disease responsible for nearly 50% of all deaths in western countries. It is characterized by the formation of lipid-laden plaques in the arterial wall reducing the lumen, decreasing the elasticity of the walls and causing many cardiovascular diseases. In response to environmental cues, vascular smooth muscle cells (SMCs) can de-differentiate, proliferate and migrate in a process known as phenotypic modulation. Recently, it has been shown that they can transdifferentiate into other cell types which are important for the formation of atherosclerotic plaques and which determine their ability to rupture and induce thrombosis. Objective: The purpose of this work is to use the most recent technologies in the field of transcriptomics, such as scRNAseq and spatial transcriptomics, to identify new SMCs-derived phenotypes induced by a pro-atherosclerotic environment and to characterize their role in the pathology, obtaining a specific gene signature that will help their identification in patients and consequentially to target them in the context of a precision medicine approach. Methods and Results: The SMC-deriving cells dissociated from the aortic arch and thoracic artery of a cohort of atheroprone ApoE knockout mice and of a control one, were analyzed by scRNAseq with 10x Genomics Chromium platform. Then, the bioinformatic analysis of the data was performed identifying a new SMC-derived cell cluster, characterized by the expression of the Adamtsl1 gene, not yet described in the literature. Furthermore, we found that this cell type is metabolically active, produces ECM, has a proinflammatory phenotype, and is a transitional state between contractile SMCs and other cellular phenotypes whose role within atherosclerotic plaques is already known. Subsequently, the presence of this cluster was also demonstrated in human and murine scRNAseq datasets from another work. Finally, two human atherosclerotic samples were analyzed using the Visium Spatial transcriptomics (10x genomics) highlighting the presence of this SMC phenotype within human plaques. Conclusions: The identified SMC-derived Adamtsl1+ cell type could be a good target to block the transdifferentiation of SMCs towards phenotypes involved in plaque formation and to improve the prognosis of patients with atherosclerosis

rs41291957 controls miR‐143 and miR‐145 expression and impacts coronary artery disease risk

Abstract The role of single nucleotide polymorphisms (SNPs) in the etiopathogenesis of cardiovascular diseases is well known. The effect of SNPs on disease predisposition has been established not only for protein coding genes but also for genes encoding microRNAs (miRNAs). The miR‐143/145 cluster is smooth muscle cell‐specific and implicated in the pathogenesis of atherosclerosis. Whether SNPs within the genomic sequence of the miR‐143/145 cluster are involved in cardiovascular disease development is not known. We thus searched annotated sequence databases for possible SNPs associated with miR‐143/145. We identified one SNP, rs41291957 (G > A), located −91 bp from the mature miR‐143 sequence, as the nearest genetic variation to this miRNA cluster, with a minor allele frequency > 10%. In silico and in vitro approaches determined that rs41291957 (A) upregulates miR‐143 and miR‐145, modulating phenotypic switching of vascular smooth cells towards a differentiated/contractile phenotype. Finally, we analysed association between rs41291957 and CAD in two cohorts of patients, finding that the SNP was a protective factor. In conclusion, our study links a genetic variation to a pathological outcome through involvement of miRNAs

Dataset related to article "Spatial resolution of cellular senescence dynamics in human colorectal liver metastasis"

<p>Dataset related to article "Spatial resolution of cellular senescence dynamics in human colorectal liver metastasis"</p><p><strong>Abstract</strong></p><p>Hepatic metastasis is a clinical challenge for colorectal cancer (CRC). Senescent cancer cells accumulate in CRC favoring tumor dissemination. Whether this mechanism progresses also in metastasis is unexplored. Here, we integrated spatial transcriptomics, 3D-microscopy, and multicellular transcriptomics to study the role of cellular senescence in human colorectal liver metastasis (CRLM). We discovered two distinct senescent metastatic cancer cell (SMCC) subtypes, transcriptionally located at the opposite pole of epithelial (e) to mesenchymal (m) transition. SMCCs differ in chemotherapy susceptibility, biological program, and prognostic roles. Mechanistically, epithelial (e)SMCC initiation relies on nucleolar stress, whereby c- myc dependent oncogene hyperactivation induces ribosomal RPL11 accumulation and DNA damage response. In a 2D pre-clinical model, we demonstrated that RPL11 co-localized with HDM2, a p53-specific ubiquitin ligase, leading to senescence activation in (e)SMCCs. On the contrary, mesenchymal (m)SMCCs undergo TGFβ paracrine activation of NOX4-p15 effectors. SMCCs display opposing effects also in the immune regulation of neighboring cells, establishing an immunosuppressive environment or leading to an active immune workflow. Both SMCC signatures are predictive biomarkers whose unbalanced ratio determined the clinical outcome in CRLM and CRC patients. Altogether, we provide a comprehensive new understanding of the role of SMCCs in CRLM and highlight their potential as new therapeutic targets to limit CRLM progression.</p><p> </p&gt