Sensitive and quantitative method to evaluate DNA methylation of the positive regulatory domains (PRDI, PRDII) and cAMP response element (CRE) in human endothelial nitric oxide synthase promote

Nitric oxide plays a prominent role in the cardiovascular system and much attention has been devoted in the last years on deciphering the regulation of human endothelial nitric oxide synthase (eNOS) expression. Epigenetic-based mechanisms have a key role in the eNOS expression and their pathologic perturbations may have profound effects on the steady state RNA levels in the endothelium. The human eNOS promoter lacks a canonical TATA box and it does not contain a proximal CpG island. A differentially DNA methylated region (DMR) in the native eNOS proximal promoter is involved in gene expression regulation. Here we describe a quantitative, sensitive and cost-effective method that, relying on a novel normalization strategy, allows the quantification of DNA methylation status of the positive regulatory domains (PRDI, PRDII) and cAMP response element (CRE) in human eNOS promoter. This technique will enable to explore the functional relevance of DNA methylation perturbations of eNOS promoter both under pathological and physiological conditions

Abnormal DNA Methylation Induced by Hyperglycemia Reduces CXCR 4 Gene Expression in CD 34+ Stem Cells

Background CD 34+ stem/progenitor cells are involved in vascular homeostasis and in neovascularization of ischemic tissues. The number of circulating CD 34+ stem cells is a predictive biomarker of adverse cardiovascular outcomes in diabetic patients. Here, we provide evidence that hyperglycemia can be "memorized" by the stem cells through epigenetic changes that contribute to onset and maintenance of their dysfunction in diabetes mellitus. Methods and Results Cord-blood-derived CD 34+ stem cells exposed to high glucose displayed increased reactive oxygen species production, overexpression of p66shc gene, and downregulation of antioxidant genes catalase and manganese superoxide dismutase when compared with normoglycemic cells. This altered oxidative state was associated with impaired migration ability toward stromal-cell-derived factor 1 alpha and reduced protein and mRNA expression of the C-X-C chemokine receptor type 4 ( CXCR 4) receptor. The methylation analysis by bisulfite Sanger sequencing of the CXCR 4 promoter revealed a significant increase in DNA methylation density in high-glucose CD 34+ stem cells that negatively correlated with mRNA expression (Pearson r=-0.76; P=0.004). Consistently, we found, by chromatin immunoprecipitation assay, a more transcriptionally inactive chromatin conformation and reduced RNA polymerase II engagement on the CXCR 4 promoter. Notably, alteration of CXCR 4 DNA methylation, as well as transcriptional and functional defects, persisted in high-glucose CD 34+ stem cells despite recovery in normoglycemic conditions. Importantly, such an epigenetic modification was thoroughly confirmed in bone marrow CD 34+ stem cells isolated from sternal biopsies of diabetic patients undergoing coronary bypass surgery. Conclusions CD 34+ stem cells "memorize" the hyperglycemic environment in the form of epigenetic modifications that collude to alter CXCR 4 receptor expression and migration

P3249The endocannabinoid system is a novel therapeutic target for muscular dystrophy cardiomyopathy

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Persone anziane e cura. Riflessioni pedagogiche e narrazioni nel tempo della pandemia

Esito interdisciplinare di riflessioni e scambi tra professionisti medici, consulenti pedagogici e operatori della cura e dell'assistenza, il volume affronta la condizione della persona anziana nell\u2019attuale quadro pandemico, proponendo una pedagogia degli anziani orientata al compito della cura e della responsabilit\ue0 verso le persone nei diversi ambiti di cura, assistenza e accompagnamento.Interdisciplinary outcome of reflections and exchanges between medical professionals, pedagogical consultants and care and assistance operators, the volume addresses the condition of the elderly person in the current pandemic framework, proposing a pedagogy of the elderly oriented to the task of care and responsibility towards people in the areas of care, assistance and accompaniment