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Lamin A is a component of the inner nuclear membrane that, together with epigenetic factors, organizes the genome in higher order structures required for transcriptional control. Mutations in the lamin A/C gene cause several diseases belonging to the class of laminopathies, including muscular dystrophies. Nevertheless, molecular mechanisms involved in the pathogenesis of lamin A-dependent dystrophies are still largely unknown. The polycomb group (PcG) of proteins are epigenetic repressors and lamin A interactors, primarily involved in the maintenance of cell identity. Using a murine model of Emery-Dreifuss muscular dystrophy (EDMD), we show here that lamin A loss deregulated PcG positioning in muscle satellite stem cells, leading to derepression of non-muscle-specific genes and [p16.sup.INK4a], a senescence driver encoded in the Cdkn2a locus. This aberrant transcriptional program caused impairment in self-renewal, loss of cell identity, and premature exhaustion of the quiescent satellite cell pool. Genetic ablation of the Cdkn2a locus restored muscle stem cell properties in lamin A/C-null dystrophic mice. Our findings establish a direct link between lamin A and PcG epigenetic silencing and indicate that lamin A-dependent muscular dystrophy can be ascribed to intrinsic epigenetic dysfunctions of muscle stem cells., Introduction The nuclear lamina (NL) is located in the inner part of the nuclear membrane and is composed of a complex network of type V filament proteins, the lamins (1, [...]

Astrocytes are essential players in development and functions, being particularly relevant as regulators of brain energy metabolism, ionic homeostasis and synaptic transmission. They are also the major source of l‐serine in the brain, which is synthesized from the glycolytic intermediate 3‐phosphoglycerate through the phosphorylated pathway. l‐Serine is the precursor of the two main co‐agonists of the N‐methyl‐d‐aspartate receptor, glycine and d‐serine. Strikingly, dysfunctions in both l‐ and d‐serine metabolism are associated with neurological and psychiatric disorders. Here, we exploited a differentiation protocol, based on the generation of human mature astrocytes from neural stem cells, and investigated the modification of the proteomic and metabolomic profile during the differentiation process. We show that differentiated astrocytes are more similar to mature rather than to reactive ones, and that axogenesis and pyrimidine metabolism increase up to 30 days along with the folate cycle and sphingolipid metabolism. Consistent with the proliferation and cellular maturation processes that are taking place, also the intracellular levels of l‐serine, glycine, threonine, l‐ and d‐aspartate (which level is unexpectedly higher than that of d‐serine) show the same biosynthetic time course. A significant utilization of l‐serine from the medium is apparent while glycine is first consumed and then released with a peak at 30 days, parallel to its intracellular level. These results underline how metabolism changes during astrocyte differentiation, highlight that d‐serine synthesis is restricted in differentiated astrocytes and provide a valuable model for developing potential novel therapeutic approaches to address brain diseases, especially the ones related to serine metabolism alterations. [ABSTRACT FROM AUTHOR]

Radiological imaging is currently employed as the most effective technique for screening, diagnosis, and follow up of patients with breast cancer (BC), the most common type of tumor in women worldwide. However, the introduction of the omics sciences such as metabolomics, proteomics, and molecular genomics, have optimized the therapeutic path for patients and implementing novel information parallel to the mutational asset targetable by specific clinical treatments. Parallel to the "omics" clusters, radiological imaging has been gradually employed to generate a specific omics cluster termed "radiomics". Radiomics is a novel advanced approach to imaging, extracting quantitative, and ideally, reproducible data from radiological images using sophisticated mathematical analysis, including disease-specific patterns, that could not be detected by the human eye. Along with radiomics, radiogenomics, defined as the integration of "radiology" and "genomics", is an emerging field exploring the relationship between specific features extracted from radiological images and genetic or molecular traits of a particular disease to construct adequate predictive models. Accordingly, radiological characteristics of the tissue are supposed to mimic a defined genotype and phenotype and to better explore the heterogeneity and the dynamic evolution of the tumor over the time. Despite such improvements, we are still far from achieving approved and standardized protocols in clinical practice. Nevertheless, what can we learn by this emerging multidisciplinary clinical approach? This minireview provides a focused overview on the significance of radiomics integrated by RNA sequencing in BC. We will also discuss advances and future challenges of such radiomics-based approach. [ABSTRACT FROM AUTHOR]

AL amyloidosis is caused by the misfolding of immunoglobulin light chains leading to an impaired function of tissues and organs in which they accumulate. Due to the paucity of -omics profiles from undissected samples, few studies have addressed amyloid-related damage system wide. To fill this gap, we evaluated proteome changes in the abdominal subcutaneous adipose tissue of patients affected by the AL isotypes κ and λ. Through our retrospective analysis based on graph theory, we have herein deduced new insights representing a step forward from the pioneering proteomic investigations previously published by our group. ECM/cytoskeleton, oxidative stress and proteostasis were confirmed as leading processes. In this scenario, some proteins, including glutathione peroxidase 1 (GPX1), tubulins and the TRiC complex, were classified as biologically and topologically relevant. These and other results overlap with those already reported for other amyloidoses, supporting the hypothesis that amyloidogenic proteins could induce similar mechanisms independently of the main fibril precursor and of the target tissues/organs. Of course, further studies based on larger patient cohorts and different tissues/organs will be essential, which would be a key point that would allow for a more robust selection of the main molecular players and a more accurate correlation with clinical aspects. [ABSTRACT FROM AUTHOR]

Objectives: Extracellular vesicles (EVs) are key biological mediators of several physiological functions within the cell microenvironment. Platelets are the most abundant source of EVs in the blood. Similarly, platelet lysate (PL), the best platelet derivative and angiogenic performer for regenerative purposes, is enriched of EVs, but their role is still too poorly discovered to be suitably exploited. Here, we explored the contribution of the EVs in PL, by investigating the angiogenic features extrapolated from that possessed by PL. Methods: We tested angiogenic ability and molecular cargo in 3D bioprinted models and by RNA sequencing analysis of PL‐derived EVs. Results: A subset of small vesicles is highly represented in PL. The EVs do not retain aggregation ability, preserving a low redox state in human umbilical vein endothelial cells (HUVECs) and increasing the angiogenic tubularly‐like structures in 3D endothelial bioprinted constructs. EVs resembled the miRNome profile of PL, mainly enriched with small RNAs and a high amount of miR‐126, the most abundant angiogenic miRNA in platelets. The transfer of miR‐126 by EVs in HUVEC after the in vitro inhibition of the endogenous form, restored angiogenesis, without involving VEGF as a downstream target in this system. Conclusion: PL is a biological source of available EVs with angiogenic effects involving a miRNAs‐based cargo. These properties can be exploited for targeted molecular/biological manipulation of PL, by potentially developing a product exclusively manufactured of EVs. [ABSTRACT FROM AUTHOR]

Health visitors (HVs) and environmental health officers (EHOs) are the healthcare workers (HCWs) who, in the Italian National Health Service, mainly operate in the prevention departments of local health authorities, guaranteeing the territorial activities specifically declared with the respective professional profiles. During the SARS-CoV-2 pandemic, it was necessary to reallocate all HCWs supporting Hygiene and Public Health Services involved on the front lines of the emergency, in order to perform preventive activities and to take immediate action to fight the spread of the virus. By means of an IT survey consisting of three sections, this study investigated how 960 HVs and EHOs dealt with this reallocation, with the shifting in service assignment, and with the perceived level of fatigue and pressure, through the application of skills acquired from university training. The synergy among the preventive health professions, the ability to work in a multi-professional team, and the complementary training of HCWs represent the main strengths for overcoming future public health challenges, aimed at protecting human health. [ABSTRACT FROM AUTHOR]

Abscisic acid (ABA) regulates plant responses to stress, partly via NO. In mammals, ABA stimulates NO production by innate immune cells and keratinocytes, glucose uptake and mitochondrial respiration by skeletal myocytes and improves blood glucose homeostasis through its receptors LANCL1 and LANCL2. We hypothesized a role for the ABA-LANCL1/2 system in cardiomyocyte protection from hypoxia via NO. The effect of ABA and of the silencing or overexpression of LANCL1 and LANCL2 were investigated in H9c2 rat cardiomyoblasts under normoxia or hypoxia/reoxygenation. In H9c2, hypoxia induced ABA release, and ABA stimulated NO production. ABA increased the survival of H9c2 to hypoxia, and L-NAME, an inhibitor of NO synthase (NOS), abrogated this effect. ABA also increased glucose uptake and NADPH levels and increased phosphorylation of Akt, AMPK and eNOS. Overexpression or silencing of LANCL1/2 significantly increased or decreased, respectively, transcription, expression and phosphorylation of AMPK, Akt and eNOS; transcription of NAMPT, Sirt1 and the arginine transporter. The mitochondrial proton gradient and cell vitality increased in LANCL1/2-overexpressing vs. -silenced cells after hypoxia/reoxygenation, and L-NAME abrogated this difference. These results implicate the ABA-LANCL1/2 hormone-receptor system in NO-mediated cardiomyocyte protection against hypoxia. [ABSTRACT FROM AUTHOR]

Before entering human clinical studies to evaluate their safety and effectiveness, new drugs and novel medical treatments are subject to extensive animal testing that are expensive and time-consuming. By contrast, advanced technologies enable the development of animal-free models that allow the efficacy of innovative therapies to be studied without sacrificing animals, while providing helpful information and details. We report on the powerful combination of 3D bioprinting (3DB) and photo-thermal therapy (PTT) applications. To this end, we realize a 3DB construct consisting of glioblastoma U87-MG cells in a 3D geometry, incorporating biomimetic keratin-coated gold nanoparticles (Ker-AuNPs) as a photo-thermal agent. The resulting plasmonic 3DB structures exhibit a homogeneous cell distribution throughout the entire volume while promoting the localization of Ker-AuNPs within the cells. A 3D immunofluorescence assay and transmission electron microscopy (TEM) confirm the uniform distribution of fluorescent-labeled Ker-AuNPs in the volume and their capability to enter the cells. Laser-assisted (λ = 532 nm) PTT experiments demonstrate the extraordinary ability of Ker-AuNPs to generate heating, producing the highest temperature rise of about 16 °C in less than 2 min. [ABSTRACT FROM AUTHOR]

The cellular heterogeneity of the tumor environment of breast cancer (BC) is extremely complex and includes different actors such as neoplastic, stromal, and immunosuppressive cells, which contribute to the chemical and mechanical modification of the environment surrounding the tumor-exasperating immune-escaping mechanisms. In addition to molecular signals that make the tumor microenvironment (TME) unacceptable for the penetrance of the immune system, the physical properties of tumoral extracellular matrix (tECM) also have carved out a fundamental role in the processes of the protection of the tumor niche. Tumor-associated macrophages (TAMs), with an M2 immunosuppressive phenotype, are important determinants for the establishment of a tumor phenotype excluded from T cells. NF-κB transcription factors orchestrate innate immunity and represent the common thread between inflammation and cancer. Many studies have focused on canonical activation of NF-κB; however, activation of non-canonical signaling predicts poor survival and resistance to therapy. In this scenario, we demonstrated the existence of an unusual association of NF-κB components in TAMs that determines the deposition of HSPG2 that affects the stiffness of tECM. These results highlight a new mechanism counterbalanced between physical factors and a new perspective of mechano-pathology to be targeted to counteract immune evasion in BC. [ABSTRACT FROM AUTHOR]

The Neuromuscular Junction (NMJ) is a chemical synapse localized between the terminal branches of the spinal motor neurons and myofibers. In the past two decades coculture systems to generate the NMJ in culture are developed to address concerns about animal models, despite the complexity of its highly specialized structure makes the in vitro modeling a challenging task. Microfluidics, unlike mass cocultures, allow spatial and temporal control over different microenvironment by manipulating either neural cells or muscle cell populations independently. Therefore, exploiting an organ‐on‐a‐chip approach, the aim is to obtain a reliable and predictive in vitro human model of NMJ in physiological and pathological conditions, to investigate the occurrence of synapse detriment in α‐sarcoglycanopathy, a subtype of limb‐girdle muscular dystrophy. For this purpose, motor neurons derived from human induced pluripotent stem cells (hiPSCs) and either healthy or α‐sarcoglycan mutant human myogenic progenitors are seeded in two separated chambers of a microfluidic device. Differentiated myotubes and hiPSCs‐derived motor neurons on‐chip are able to establish points of interaction where pre‐ and postsynaptic structures colocalize. Moreover, the attraction of motor neurons axons by muscle fibers and the NMJ maturation appear to be affected by the muscular compartment, being impaired by the dystrophic cellular component. [ABSTRACT FROM AUTHOR]

Mechanistic target of rapamycin (MTOR) plays a critical role in the regulation of cell growth and in the response to energy state changes. Drugs inhibiting MTOR are increasingly used in antineoplastic therapies. Myocardial MTOR activity changes during hypertrophy and heart failure (HF). However, whether MTOR exerts a positive or a negative effect on myocardial function remains to be fully elucidated. Here, we show that ablation of Mtor in the adult mouse myocardium results in a fatal, dilated cardiomyopathy that is characterized by apoptosis, autophagy, altered mitochondrial structure, and accumulation of eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1). 4E-BP1 is an MTOR-containing multi-protein complex-1 (MTORC1) substrate that inhibits translation initiation. When subjected to pressure overload, Mtor-ablated mice demonstrated an impaired hypertrophic response and accelerated HF progression. When the gene encoding 4E-BP1 was ablated together with Mtor, marked improvements were observed in apoptosis, heart function, and survival. Our results demonstrate a role for the MTORC1 signaling network in the myocardial response to stress. In particular, they highlight the role of 4E-BP1 in regulating cardiomyocyte viability and in HF. Because the effects of reduced MTOR activity were mediated through increased 4E-BP1 inhibitory activity, blunting this mechanism may represent a novel therapeutic strategy for improving cardiac function in clinical HF., Introduction Mechanistic target of rapamycin (MTOR) is a key regulator of protein synthesis in the cardiomyocyte (1, 2). Increased protein synthesis underpins hypertrophic growth, a salient feature of the heart [...]

Alterations in the DMD gene, which codes for the protein dystrophin, cause forms of dystrophinopathies such as Duchenne muscular dystrophy, an X-linked disease. Cardiomyopathy linked to DMD mutations is becoming the leading cause of death in patients with dystrophinopathy. Since phenotypic pathophysiological mechanisms are not fully understood, the improvement and development of new disease models, considering their relative advantages and disadvantages, is essential. The application of genetic engineering approaches on induced pluripotent stem cells, such as gene-editing technology, enables the development of physiologically relevant human cell models for in vitro dystrophinopathy studies. The combination of induced pluripotent stem cells-derived cardiovascular cell types and 3D bioprinting technologies hold great promise for the study of dystrophin-linked cardiomyopathy. This combined approach enables the assessment of responses to physical or chemical stimuli, and the influence of pharmaceutical approaches. The critical objective of in vitro microphysiological systems is to more accurately reproduce the microenvironment observed in vivo. Ground-breaking methodology involving the connection of multiple microphysiological systems comprised of different tissues would represent a move toward precision body-on-chip disease modelling could lead to a critical expansion in what is known about inter-organ responses to disease and novel therapies that have the potential to replace animal models. In this review, we will focus on the generation, development, and application of current cellular, animal, and potential for bio-printed models, in the study of the pathophysiological mechanisms underlying dystrophin-linked cardiomyopathy in the direction of personalized medicine. [ABSTRACT FROM AUTHOR]

In the central nervous system (CNS), oligodendrocytes are the glial element in charge of myelin formation. Obtaining an overall presence of oligodendrocyte precursor cells/oligodendrocytes (OPCs/OLs) in culture from different sources of NSCs is an important research area, because OPCs/OLs may provide a promising therapeutic strategy for diseases affecting myelination of axons. The present study was designed to differentiate human olfactory bulb NSCs (OBNSCs) into OPCs/OLs and using expression profiling (RT-qPCR) gene, immunocytochemistry, and specific protein expression to highlight molecular mechanism(s) underlying differentiation of human OBNSCs into OPCs/OLs. The differentiation of OBNSCs was characterized by a simultaneous appearance of neurons and glial cells. The differentiation medium, containing cAMP, PDGFA, T3, and all-trans-retinoic acid (ATRA), promotes OBNSCs to generate mostly oligodendrocytes (OLs) displaying morphological changes, and appearance of long cytoplasmic processes. OBNSCs showed, after 5 days in OLs differentiation medium, a considerable decrease in the number of nestin positive cells, which was associated with a concomitant increase of NG2 immunoreactive cells and few O4(+)-OPCs. In addition, a significant up regulation in gene and protein expression profile of stage specific cell markers for OPCs/OLs (CNPase, Galc, NG2, MOG, OLIG1, OLIG2, MBP), neurons, and astrocytes (MAP2, β-TubulinIII, GFAP) and concomitant decrease of OBNSCs pluripotency markers (Oct4, Sox2, Nestin), was demonstrated following induction of OBNSCs differentiation. Taken together, the present study demonstrate the marked ability of a cocktail of factors containing PDGFA, T3, cAMP, and ATRA, to induce OBNSCs differentiation into OPCs/OLs and shed light on the key genes and pathological pathways involved in this process. [ABSTRACT FROM AUTHOR]

Facioscapulohumeral dystrophy (FSHD) is an autosomal dominant muscular dystrophy and one of the more frequent hereditary myopathy. The pathology shows a wide range of clinical signs, with modifying factors contributing to this variability. Among these factors, the beneficial activity of estrogen hormones is still controversial. We investigated the effect of the estrogens 17β-estradiol (E2) and the 5α-dihydrotestosterone-derived 3β-androstenediol (3β-diol) on muscle regeneration. To recapitulate human cell hormone sensitivity, we exploited a humanized heterokaryon FSHD mouse model, constituted by engrafting of human primary muscle mesenchymal stroma cells with perivascular cells (PVCs)-like phenotype in surgerytreated murine muscle. Lentiviral expression of the pathogenic FSHD gene, DUX4, in these cells impaired muscle structural and functional recovery. Notably, both hormones counteracted DUX4 activity and rescued structural and functional muscle performance impaired by DUX4 expression. Interestingly, E2 and 3β-diol act differently on muscle recovery by reducing muscle fibrosis and improving muscle differentiation, respectively. These results demonstrate that estrogens recover murine muscle regeneration reduced by DUX4 expression and support the hypothesis of their beneficial activity on human FSHD muscle. [ABSTRACT FROM AUTHOR]

The incidence of diabetes mellitus is rapidly increasing throughout the world. Although the exact cause of the disease is not fully clear, perhaps, genetics, ethnic origin, obesity, age, and lifestyle are considered as few of many contributory factors for the disease pathogenesis. In recent years, the disease progression is particularly linked with functional and taxonomic alterations in the gastrointestinal tract microbiome. A change in microbial diversity, referred as microbial dysbiosis, alters the gut fermentation profile and intestinal wall integrity and causes metabolic endotoxemia, low-grade inflammation, autoimmunity, and other affiliated metabolic disorders. This article aims to summarize the role of the gut microbiome in the pathogenesis of diabetes. Additionally, we summarize gut microbial dysbiosis in preclinical and clinical diabetes cases reported in literature in the recent years. [ABSTRACT FROM AUTHOR]

Background and Purpose: Propranolol is a vasoactive drug that shows antiangiogenic and antitumour activities in melanoma. However, it is unknown whether these activities are dose-dependent and whether there is a relationship between systemic vascular effects of propranolol and anti-melanoma activity.Experimental Approach: Effects of increasing doses of propranolol (10, 20, 30 and 40 mg·kg-1 ·day-1 ) on tumour growth were studied in B16F10 melanoma-bearing mice. Histological and biochemical analyses were used to assess propranolol effects on angiogenesis and cancer cell proliferation. Systemic vascular resistance (SVR) was evaluated by measuring cardiac output and arterial BP.Key Results: In vitro analyses revealed that B16F10 cells expressed β-adrenoceptors, but neither isoprenaline, a β-adrenoceptor agonist, nor the β-blocker propranolol affected cancer cell proliferation. In vivo studies showed that the antitumour efficacy of propranolol follows a U-shaped biphasic dose-response curve. Low doses (10 and 20 mg·kg-1 ·day-1 ) significantly inhibit tumour growth, whereas higher doses are progressively less effective. We also found that high-dose propranolol stimulates tumour arteriogenesis whereas no effect on angiogenesis was observed at any dose. Based on these data and considering that propranolol is a vasoactive drug, we hypothesized that it causes systemic vasoconstriction or vasodilation depending on the dose and thus alters tumour perfusion and growth. Consistent with this hypothesis, we found that propranolol has a biphasic effect on SVR with low and high doses producing vasoconstriction and vasodilation respectively.Conclusions and Implications: Propranolol inhibits melanoma growth in a U-shaped biphasic manner. A direct relationship exists between SVR and anti-melanoma activity. [ABSTRACT FROM AUTHOR]

Critical limb ischemia (CLI), foot ulcers, former amputation, and impaired regeneration are independent risk factors for limb amputation in subjects with diabetes. The present work investigates whether and by which mechanism diabetes negatively impacts on functional properties of muscular pericytes (MPs), which are resident stem cells committed to reparative angiomyogenesis. We obtained muscle biopsy samples from patients with diabetes who were undergoing major limb amputation and control subjects. Diabetic muscles collected at the rim of normal tissue surrounding the plane of dissection showed myofiber degeneration, fat deposition, and reduction of MP vascular coverage. Diabetic MPs (D-MPs) display ultrastructural alterations, a differentiation bias toward adipogenesis at the detriment of myogenesis and an inhibitory activity on angiogenesis. Furthermore, they have an imbalanced redox state, with downregulation of the antioxidant enzymes superoxide dismutase 1 and catalase, and activation of the pro-oxidant protein kinase C isoform β-II (PKCβII)-dependent p66Shc signaling pathway. A reactive oxygen species scavenger or, even more effectively, clinically approved PKCβII inhibitors restore D-MP angiomyogenic activity. Inhibition of the PKCβII-dependent p66Shc signaling pathway could represent a novel therapeutic approach for the promotion of muscle repair in individuals with diabetes. [ABSTRACT FROM AUTHOR]

Skeletal muscle tissue engineering is a promising approach for the treatment of muscular disorders. However, the complex organization of muscle, combined with the difficulty in finding an appropriate source of regenerative cells and in providing an adequate blood supply to the engineered tissue, makes this a hard task to face. In the present work, we describe an innovative approach to rejuvenate adult skeletal muscle-derived pericytes (MP) based on the use of a PEG-based hydrogel scaffold. MP were isolated from young (piglet) and adult (boar) pigs to assess whether aging affects tissue regeneration efficiency. In vitro, MP from boars had similar morphology and colony forming capacity to piglet MP, but an impaired ability to form myotubes and capillary-like structures. However, the use of a PEG-based hydrogel to support adult MP significantly improved their myogenic differentiation and angiogenic potentials in vitro and in vivo. Thus, PEG-based hydrogel scaffolds may provide a progenitor cell "niche" that promotes skeletal muscle regeneration and blood vessel growth, and together with pericytes may be developed for use in regenerative applications. [ABSTRACT FROM AUTHOR]

Introduction The cancer stem cell model links neoplastic cells with normal stem cell biology, but little is known on how normal stem cells are transformed into cancer stem cells. Methods To investigate the processes underlying the transformation of normal stem cells we developed in vitro a cancer stem cell model from human amniotic and chorionic placenta membranes. In this model we studied the expression of specific stem cell molecules by flow cytometry, and genes, by real time RT-PCR. Microscopy immunfluorescence was employed to investigate the proliferative and differentiation patterns. Fluorescence microscopy and FACS were employed to investigate the proliferative and differentiation patterns. To evaluate the tumorigenic potential of our model we injected the cells into NOD.CB17-Prkdcscid/ NCrHsd mice. Results Normal human stem cells from amniotic and chorionic placenta membranes were converted into neural cell lineages, under specific conditions, to form secondary neurospheres with a capacity for self-renewal. After extensive in vitro culture, these cells underwent spontaneous transformations and acquired a neuroblastoma (NB)-like phenotype with an elevated proliferative potential that is comparable to established neuroblastoma cell lines. The ability of these cells to transform their phenotype was evidenced by increased clonogenic ability in vitro; by augmented expression level of certain proliferation- and transformation-related genes (e.g., CCNA2, MYCN, ENPP2, GRIA3, and KIT); by the presence of multinucleated and hyperdiploid cells. We further demonstrated that the transformed phenotype is an NB by measuring the expression of NB-specific markers, disialoganglioside GD2 and N-Myc proteins. Conclusions We have developed a cancer stem cell model starting from normal human stem cells derived from amniotic and chorionic placenta membranes. These cells are able to differentiate into neural cell lineages and to undergo spontaneous transformations and acquire an NB-like phenotype. [ABSTRACT FROM AUTHOR]

In pluripotent stem cells, there is increasing evidence for crosstalk between post-transcriptional and transcriptional networks, offering multifold steps at which pluripotency can be controlled. In addition to well-studied transcription factors, chromatin modifiers and miRNAs, RNA-binding proteins are emerging as fundamental players in pluripotency regulation. Here, we report a new role for the RNA-binding protein ESRP1 in the control of pluripotency. Knockdown of Esrp1 in mouse embryonic stem cells induces, other than the well-documented epithelial to mesenchymal-like state, also an increase in expression of the core transcription factors Oct4, Nanog and Sox2, thereby enhancing self-renewal of these cells. Esrp1-depleted embryonic stem cells displayed impaired early differentiation in vitro and formed larger teratomas in vivo when compared to control embryonic stem cells. We also show that ESRP1 binds to Oct4 and Sox2 mRNAs and decreases their polysomal loading. ESRP1 thus acts as a physiological regulator of the finely-tuned balance between self-renewal and commitment to a restricted developmental fate. Importantly, both mouse and human epithelial stem cells highly express ESRP1, pinpointing the importance of this RNA-binding protein in stem cell biology. [ABSTRACT FROM AUTHOR]

Stem cells and regenerative medicine have obtained a remarkable consent from the scientific community for their promising ability to recover aged, injured and diseased tissue. However, despite the noteworthy potential, hurdles currently hinder their use and clinical application: cell survival, immune response, tissue engraftment and efficient differentiation. Hence a new interdisciplinary scientific approach, such as tissue engineering, is going deep attempts to mimic neo-tissue-genesis as well as stem cell engraftment amelioration. Skeletal muscle tissue engineering represents a great potentiality in medicine for muscle regeneration exploiting new generation injectable hydrogel as scaffold supporting progenitor/stem cells for muscle differentiation reconstructing the natural skeletal muscle tissue architecture influenced by matrix mechanical and physical property and by a dynamic environment. [ABSTRACT FROM AUTHOR]

The article focuses on a research that aims to identify the pathways that regulate intracellular Ca2+ in human cardiac progenitor cells (hCPCs) and to evaluate whether Ca2+ are triggered by hCPCs and cardiomyocytes' interaction. It states that Ca2+ indicator and Fluo-3 and 2-photon microscopy were used to measure cytosolic Ca2+ levels. Results show that hCPCs show spontaneous elevations in intracellular Ca2+.

In diabetic patients and animal models of diabetes mellitus (DM), circulating endothelial progenitor cell (EPC) number is lower than in normoglycaemic conditions and EPC angiogenic properties are inhibited. Stromal cell derived factor-1 (SDF-1) plays a key role in bone marrow (BM) c-kit+ stem cell mobilization into peripheral blood (PB), recruitment from PB into ischemic tissues and differentiation into endothelial cells. The aim of the present study was to examine the effect of DM in vivo and in vitro, on murine BM-derived c-kit+ cells and on their response to SDF-1. Acute hindlimb ischemia was induced in streptozotocin-treated DM and control mice; circulating c-kit+ cells exhibited a rapid increase followed by a return to control levels which was significantly faster in DM than in control mice. CXCR4 expression by BM c-kit+ cells as well as SDF-1 protein levels in the plasma and in the skeletal muscle, both before and after the induction of ischemia, were similar between normoglycaemic and DM mice. However, BM-derived c-kit+ cells from DM mice exhibited an impaired differentiation towards the endothelial phenotype in response to SDF-1; this effect was associated with diminished protein kinase phosphorylation. Interestingly, SDF-1 ability to induce differentiation of c-kit+ cells from DM mice was restored when cells were cultured under normoglycaemic conditions whereas c-kit+ cells from normoglycaemic mice failed to differentiate in response to SDF-1 when they were cultured in hyperglycaemic conditions. These results show that DM diminishes circulating c-kit+ cell number following hindlimb ischemia and inhibits SDF-1-mediated AKT phosphorylation and differentiation towards the endothelial phenotype of BM-derived c-kit+ cells. [ABSTRACT FROM AUTHOR]

Abstract: Vgf, is a neuro-endocrine specific gene encoding for a large protein precursor of different peptides. A role for VGF in pain modulation has been suggested from immunohistochemical studies showing VGF mRNA widely expressed in primary sensory neurons. In this study, the presence of VGF on the primary sensory afferents in mice was confirmed by showing its immunostaining in cultured neurons of dorsal root ganglia in secretory granule varicosities colocalized with Substance P. Moreover, the functional role of a C-terminal internal VGF-derived peptide, i.e. TLQP-21, was assessed by investigating its peripheral (1, 2, 4, 8mM) and central (1, 2, 4mM) effects on inflammatory pain in the formalin test. A significant increase of pain-related licking response following peripheral injection of TLQP-21 (4 and 8mM) was observed in the second inflammatory phase of the test. In addition, an increase in licking response was detected when 4mM of the peptide was injected alone without formalin. On the other hand, the central administration of TLQP-21 induced an U-shaped curve, with the dose of 2mM being analgesic during the second phase. This study shows for the first time that a VGF-derived peptide may be involved in inflammatory pain in vivo and demonstrates a different action for TLQP21 at the peripheral and central levels of the nociceptive pathways. [Copyright &y& Elsevier]

The immune system is a fine modulator of the tumor biology supporting or inhibiting its progression, growth, invasion and conveys the pharmacological treatment effect. Tumors, on their side, have developed escaping mechanisms from the immune system action ranging from the direct secretion of biochemical signals to an indirect reaction, in which the cellular actors of the tumor microenvironment (TME) collaborate to mechanically condition the extracellular matrix (ECM) making it inhospitable to immune cells. TME is composed of several cell lines besides cancer cells, including tumor-associated macrophages, cancer-associated fibroblasts, CD4+ and CD8+ lymphocytes, and innate immunity cells. These populations interface with each other to prepare a conservative response, capable of evading the defense mechanisms implemented by the host's immune system. The presence or absence, in particular, of cytotoxic CD8+ cells in the vicinity of the main tumor mass, is able to predict, respectively, the success or failure of drug therapy. Among various mechanisms of immunescaping, in this study, we characterized the modulation of the phenotypic profile of CD4+ and CD8+ cells in resting and activated states, in response to the mechanical pressure exerted by a three-dimensional in vitro system, able to recapitulate the rheological and stiffness properties of the tumor ECM. [ABSTRACT FROM AUTHOR]

The Cdkn2a locus is one of the most studied tumor suppressor loci in the context of several cancer types. However, in the last years, its expression has also been linked to terminal differentiation and the activation of the senescence program in different cellular subtypes. Knock-out (KO) of the entire locus enhances the capability of stem cells to proliferate in some tissues and respond to severe physiological and non-physiological damages in different organs, including the heart. Emery–Dreifuss muscular dystrophy (EDMD) is characterized by severe contractures and muscle loss at the level of skeletal muscles of the elbows, ankles and neck, and by dilated cardiomyopathy. We have recently demonstrated, using the LMNA Δ8–11 murine model of Emery–Dreifuss muscular dystrophy (EDMD), that dystrophic muscle stem cells prematurely express non-lineage-specific genes early on during postnatal growth, leading to rapid exhaustion of the muscle stem cell pool. Knock-out of the Cdkn2a locus in EDMD dystrophic mice partially restores muscle stem cell properties. In the present study, we describe the cardiac phenotype of the LMNA Δ8–11 mouse model and functionally characterize the effects of KO of the Cdkn2a locus on heart functions and life expectancy. [ABSTRACT FROM AUTHOR]

The recent advances, offered by cell therapy in the regenerative medicine field, offer a revolutionary potential for the development of innovative cures to restore compromised physiological functions or organs. Adult myogenic precursors, such as myoblasts or satellite cells, possess a marked regenerative capacity, but the exploitation of this potential still encounters significant challenges in clinical application, due to low rate of proliferation in vitro, as well as a reduced self-renewal capacity. In this scenario, induced pluripotent stem cells (iPSCs) can offer not only an inexhaustible source of cells for regenerative therapeutic approaches, but also a valuable alternative for in vitro modeling of patient-specific diseases. In this study we established a reliable protocol to induce the myogenic differentiation of iPSCs, generated from pericytes and fibroblasts, exploiting skeletal muscle-derived extracellular vesicles (EVs), in combination with chemically defined factors. This genetic integration-free approach generates functional skeletal myotubes maintaining the engraftment ability in vivo. Our results demonstrate evidence that EVs can act as biological "shuttles" to deliver specific bioactive molecules for a successful transgene-free differentiation offering new opportunities for disease modeling and regenerative approaches. [ABSTRACT FROM AUTHOR]

Mechanistic target of rapamycin (MTOR) plays a critical role in the regulation of cell growth and in the response to energy state changes. Drugs inhibiting MTOR are increasingly used in antineoplastic therapies. Myocardial MTOR activity changes during hypertrophy and heart failure (HF). However, whether MTOR exerts a positive or a negative effect on myocardial function remains to be fully elucidated. Here, we show that ablation of Mtor in the adult mouse myocardium results in a fatal, dilated cardiomyopathy that is characterized by apoptosis, autophagy, altered mitochondrial structure, and accumulation of eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1). 4E-BP1 is an MTOR-containing multiprotein complex-1 (MTORC1) substrate that inhibits translation initiation. When subjected to pressure overload, Mtor-ablated mice demonstrated an impaired hypertrophic response and accelerated HF progression. When the gene encoding 4E-BP1 was ablated together with Mtor, marked improvements were observed in apoptosis, heart function, and survival. Our results demonstrate a role for the MTORC1 signaling network in the myocardial response to stress. In particular, they highlight the role of 4E-BP1 in regulating cardiomyocyte viability and in HF. Because the effects of reduced MTOR activity were mediated through increased 4E-BP1 inhibitory activity, blunting this mechanism may represent a novel therapeutic strategy for improving cardiac function in clinical HF. [ABSTRACT FROM AUTHOR]

Expression of the β-myosin heavy chain (β-MHC), a major component of the cardiac contractile apparatus, is tightly regulated as even modest increases can be detrimental to heart under stress. In healthy hearts, continuous inhibition of β-adrenergic tone upregulates β-MHC expression. However, it is unknown whether the duration of the β-adrenergic inhibition and β-MHC expression are related. Here, we evaluated the effects of intermittent β-blockade on cardiac β-MHC expression. To this end, the β-blocker propranolol, at the dose of 15mg/kg, was administered once a day in mice for 14 days. This dosing schedule caused daily drug-free periods of at least 6 h as evidenced by propranolol plasma concentrations and cardiac β-adrenergic responsiveness. Under these conditions, β-MHC expression decreased by about 75% compared to controls. This effect was abolished in mice lacking β1- but not β2-adrenergic receptors (β-AR) indicating that β-MHC expression is regulated in a β1-AR-dependent manner. In β1-AR knockout mice, the baseline β-MHC expression was fourfold higher than in wild-type mice. Also, we evaluated the impact of intermittent β-blockade on β-MHC expression in mice with systolic dysfunction, in which an increased β-MHC expression occurs. At 3 weeks after myocardial infarction, mice showed systolic dysfunction and upregulation of β-MHC expression. Intermittent β-blockade decreased β-MHC expression while attenuating cardiac dysfunction. In vitro studies showed that propranolol does not affect β-MHC expression on its own but antagonizes catecholamine effects on β-MHC expression. In conclusion, a direct relationship occurs between the duration of the β-adrenergic inhibition and β-MHC expression through the β1-AR. [ABSTRACT FROM AUTHOR]