Background: miR-210 hypoxamir is induced by Hif1a in hypoxic cells. Results: miR-210 expression increased during myogenic differentiation in normoxia with a Hif1a-dependent mechanism. Moreover, miR-210 displayed a cytoprotective role in response to mitochondrial dysfunction and oxidative stress. Conclusion: miR-210 regulation and function extend beyond cell response to hypoxia. Significance: Identifying miR-210 as a potential target in skeletal muscle disorders.

In human endothelial cells, nitric oxide (NO) results in class IIa histone deacetylases (HDACs) activation and marked histone deacetylation. It is unknown whether similar epigenetic events occur in embryonic stem cells (ESC) exposed to NO and how this treatment could influence ESC therapeutic potential during tissue regeneration. This study reports that the NO-dependent class IIa HDACs subcellular localization and activity decreases the global acetylation level of H3 histones in ESC and that this phenomenon is associated with the inhibition of Oct4, Nanog, and KLF4 expression. Further, a NO-induced formation of macromolecular complexes including HDAC3, 4, 7, and protein phosphatase 2A (PP2A) have been detected. These processes correlated with the expression of the mesodermalspecific protein brachyury (Bry) and the appearance of several vascular and skeletal muscle differentiation markers. These events were abolished by the class IIa-specific inhibitor MC1568 and by HDAC4 or HDAC7 short interfering RNA (siRNA). The ability of NO to induce mesodermic/ cardiovascular gene expression prompted us to evaluate the regenerative potential of these cells in a mouse model of hindlimb ischemia. We found that NO-treated ESCs injected into the cardiac left ventricle selectively localized in the ischemic hindlimb and contributed to the regeneration of muscular and vascular structures. These findings establish a key role for NO and class IIa HDACs modulation in ESC mesodermal commitment and enhanced regenerative potential in vivo. STEM CELLS 2010;28:431-442 Disclosure of potential conflicts of interest is found at the end of this article.

The CCAAT-binding transcription factor NF-Y plays a central role in regulating cellular proliferation by controlling the expression of genes required for cell-cycle progression such as cyclin A, cyclin B1, cyclin B2, cdc25A, cdc25C, and cdk1. Here we show that unrestricted NF-Y activity leads to apoptosis in an E2F1-and wild-type p53 (wtp53)-dependent manner. Unrestricted NF-Y activity induced an increase in E2F1 mRNA and protein levels. Furthermore, NF-Y directly bound the E2F1 promoter and this correlated with the appearance of open chromatin marks. The ability of NF-Y to induce apoptosis was impaired in cells lacking E2F1 and wtp53. Moreover, NF-Y overexpression elicited phosphorylation of wt p53Ser18 in an E2F1-dependent manner. Our findings establish that NF-Y acts upstream of E2F1 in p53-mediated apoptosis.

Background & Aims: Cholangiocarcinoma (CCA) is a very aggressive cancer showing high cancer stem cells (CSCs) presence. Doublecortin-like kinase1 (DCLK1) has been demonstrated as a CSC marker in different gastroenterological solid tumours. Our aim was to evaluate in vitro the expression and the biological function of DCLK1 in intrahepatic CCA (iCCA) and perihilar CCA (pCCA). Approach & Results: Specimens surgically resected of human CCA were enzymatically digested, submitted to immunosorting for specific CSC markers (LGR5, CD90, EpCAM, CD133, CD13) and primary cell cultures were prepared. DCLK1 expression was analysed in CCA cell cultures by real-time quantitative polymerase chain reaction (RT-qPCR), Western Blot and immunofluorescence. Functional studies have been performed by evaluating the effects of selective DCLK1 inhibitor (LRRK2-IN-1) on cell proliferation (MTS-Assay, cell population doubling time), apoptosis and colony formation capacity. DCLK1 was investigated in situ by immunohistochemistry and RT-qPCR. DCLK1 serum concentration was analysed by enzyme-linked immunosorbent assay (ELISA). We describe DCLK1 in CCA with an increased gene and protein DCLK1 expression in pCCA LGR5+ and in iCCA CD133+ cells compared to unsorted cells. LRRK2-IN-1 showed an anti-proliferative effect in dose-dependent manner. LRRK2-IN-1 markedly impaired cell proliferation, induced apoptosis, decreased colony formation capacity and colony size in both iCCA and pCCA compared to untreated cells. In situ analysis confirm that DCLK1 is present only in tumours, but not Accepted Article This article is protected by copyright. All rights reserved in healthy tissue. Interestingly, DCLK1 was detected in the human serum samples of iCCA (high), pCCA (high), HCC (low) and cirrhotic (low) patients, but it was almost undetectable in healthy controls. Conclusion: DCLK1 characterizes a specific CSC-subpopulation of iCCA CD133+ and pCCA LGR5+ and its inhibition exerts anti-neoplastic effects in primary CCA cell cultures. Human DCLK1 serum might represent a serum biomarker for the early CCA diagnosis.

These results indicate that Axl receptor activation modulates laminar SS anti-apoptotic effects possibly through its association with specific integrin-complexes.

The results of the present study identify CXCR4 as a key player in the EC response to acidic pH and show, for the first time, that HRE may function not only as an effector of hypoxia, but also as an acidosis response element, and raise the possibility that this may constitute a more general mechanism of transcriptional regulation at acidic pH.

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Supplementary Figures 5-6 from Transcription Factor NF-Y Induces Apoptosis in Cells Expressing Wild-Type p53 through E2F1 Upregulation and p53 Activation