Apoptosis-inducing factor (AIF) is a mitochondrial flavoprotein that, beyond its apoptotic function, is required for the normal expression of major respiratory chain complexes. Here we identified an ...AIF-interacting protein, CHCHD4, which is the central component of a redox-sensitive mitochondrial intermembrane space import machinery. Depletion or hypomorphic mutation of AIF caused a downregulation of CHCHD4 protein by diminishing its mitochondrial import. CHCHD4 depletion sufficed to induce a respiratory defect that mimicked that observed in AIF-deficient cells. CHCHD4 levels could be restored in AIF-deficient cells by enforcing its AIF-independent mitochondrial localization. This modified CHCHD4 protein reestablished respiratory function in AIF-deficient cells and enabled AIF-deficient embryoid bodies to undergo cavitation, a process of programmed cell death required for embryonic morphogenesis. These findings explain how AIF contributes to the biogenesis of respiratory chain complexes, and they establish an unexpected link between the vital function of AIF and the propensity of cells to undergo apoptosis.
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•AIF interacts with CHCHD4, a regulator of the intermembrane space import machinery•AIF regulates specific respiratory chain complexes by acting upstream of CHCHD4•AIF is indispensable for translation-coupled mitochondrial import of CHCHD4•Restoring CHCHD4 reverses the metabolic and cell death phenotypes of Aif−/y ESCs
Hangen et al. show that the mitochondrial protein AIF regulates the biogenesis of respiratory chain complexes by interacting with, and by controlling the mitochondrial import of the mammalian homolog of yeast MIA40, CHCHD4, which is the central component of a redox-sensitive mitochondrial intermembrane space import machinery.
This article reviews our current knowledge about cell-derived extracellular vesicles (EVs), including microparticles and exosomes, and their emergence as mediators of a new important mechanism of ...cell-to-cell communication. Particular emphasis has been given to the increasing involvement of EVs in the field of radiation-induced vascular injury. Although EVs have been considered for a long time as cell “dust”, they in fact precisely reflect the physiological state of the cells. The role of microparticles and exosomes in mediating vascular dysfunction suggests that they may represent novel pathways in short- or long-distance paracrine intercellular signaling in vascular environment. In this article, the mechanisms involved in the biogenesis of microparticles and exosomes, their composition and participation in the pathogenesis of vascular dysfunction are discussed. Furthermore, this article highlights the concept of EVs as potent vectors of biological information and protagonists of an intercellular communication network. Special emphasis is made on EV-mediated microRNA transfer and on the principal consequences of such signal exchange on vascular injury and radiation-induced nontargeted effect. The recent progress in elucidating the biology of EVs has provided new insights for the field of radiation, advancing their use as diagnostic biomarkers or in therapeutic interventions.
A radiological accident, whether from industrial, medical, or malicious origin, may result in localized exposure to high doses of ionizing radiations, leading to the development of local radiation ...injury (LRI), that may evolve toward deep ulceration and necrosis of the skin and underlying tissues. Early diagnosis is therefore crucial to facilitate identification and management of LRI victims. Circulating microRNAs (miRNA) have been studied as potential diagnostic biomarkers of several diseases including hematological defects following whole-body irradiation (WBI). This study aims to identify a blood miRNA signature associated with LRI in a preclinical C57BL/6J mouse model of hindlimb irradiation using different 10-MV X-ray doses that lead to injuries of different severities. To this end, we first performed broad-spectrum plasma miRNA profiling, followed by a targeted validation step, on two independent animal cohorts. Using a multivariate sparse partial least square discriminant analysis, we identified a panel of eight circulating miRNAs able to segregate mice according to LRI severity. Interestingly, these miRNAs were previously associated with WBI (miR-150-5p, miR-342-3p, miR-146a-5p), inflammation (miR-18a-5p, miR-148b-3p, miR-532-5p) and skin diseases (miR-139-5p, miR-195-5p). Our results suggest the use of circulating miRNAs as suitable molecular biomarkers for LRI prognosis and diagnosis.
MicroRNA target prediction and validation Ritchie, William; Rasko, John E J; Flamant, Stéphane
Advances in experimental medicine and biology,
01/2013, Letnik:
774
Journal Article
Recenzirano
The accurate prediction and validation of microRNA targets is essential to understanding the function of microRNAs. Computational predictions indicate that all human genes may be regulated by ...microRNAs, with each microRNA possibly targeting thousands of genes. Here we discuss computational and experimental methods for identifying mammalian microRNA targets. We describe microRNA target prediction resources and procedures that are suitable for experiments where more accurate prediction of microRNA targets is more important than detecting all putative targets. We then discuss experimental methods for identifying and validating microRNA target genes, with an emphasis on the target reporter assay as the method of choice for specifically testing functional microRNA target sites.
Micro-RNAs (miRNAs) control gene expression by destabilizing targeted transcripts and inhibiting their translation. Aberrant expression of miRNAs has been described in many human cancers, including ...chronic myeloid leukemia. Current first-line therapy for newly diagnosed chronic myeloid leukemia is imatinib mesylate, which typically produces a rapid hematologic response. However the effect of imatinib on miRNA expression in vivo has not been thoroughly examined.
Using a TaqMan Low-Density Array system, we analyzed miRNA expression in blood samples from newly diagnosed chronic myeloid leukemia patients before and within the first two weeks of imatinib therapy. Quantitative real-time PCR was used to validate imatinib-modulated miRNAs in sequential primary chronic myeloid leukemia samples (n=11, plus 12 additional validation patients). Bioinformatic target gene prediction analysis was performed based on changes in miRNA expression.
We observed increased expression of miR-150 and miR-146a, and reduced expression of miR-142-3p and miR-199b-5p (3-fold median change) after two weeks of imatinib therapy. A significant correlation (P<0.05) between the Sokal score and pre-treatment miR-142-3p levels was noted. Expression changes in the same miRNAs were consistently found in an additional cohort of chronic myeloid leukemia patients, as compared to healthy subjects. Peripheral blood cells from chronic phase and blast crisis patients displayed a 30-fold lower expression of miR-150 compared to normal samples, which is of particular interest since c-Myb, a known target of miR-150, was recently shown to be necessary for Bcr-Abl-mediated transformation.
We found that imatinib treatment of chronic myeloid leukemia patients rapidly normalizes the characteristic miRNA expression profile, suggesting that miRNAs may serve as a novel clinically useful biomarker in this disease.
Local overexposure to ionizing radiation leads to chronic inflammation, vascular damage and cachexia. Here we investigate the kinetics of inflammatory cells from day (D)1 to D180 after mouse hindlimb ...irradiation and analyze the role of monocyte (Mo) subsets in tissue revascularization. At D1, we find that Mo and T cells are mobilized from spleen and bone marrow to the blood. New vessel formation during early phase, as demonstrated by ~1.4- and 2-fold increased angiographic score and capillary density, respectively, correlates with an increase of circulating T cells, and Mo
and type 1-like macrophages in irradiated muscle. At D90 vascular rarefaction and cachexia are observed, associated with decreased numbers of circulating Mo
and Type 2-like macrophages in irradiated tissue. Moreover, CCR2- and CX3CR1-deficency negatively influences neovascularization. However adoptive transfer of Mo
enhances vessel growth. Our data demonstrate the radiation-induced dynamic inflammatory waves and the major role of inflammatory cells in neovascularization.
Although it is generally acknowledged that cytokines regulate normal hematopoiesis in an autocrine/paracrine fashion, their possible role in chronic myelogenous leukemia (CML) and resistance to ...imatinib mesylate treatment remain poorly investigated. Here, we report that CD34(+) progenitors from patients with CML at diagnosis are selectively targeted by the cytokine/alarmin interleukin (IL)-33. Indeed, CML CD34(+) progenitors upregulate their cell surface expression of the IL-33-specific receptor chain ST2, proliferate and produce cytokines in response to IL-33, conversely to CD34(+) cells from healthy individuals. Moreover, ST2 overexpression is normalized following imatinib mesylate therapy, whereas IL-33 counteracts in vitro imatinib mesylate-induced growth arrest in CML CD34(+) progenitors via reactivation of the STAT5 pathway, thus supporting the notion that IL-33 may impede the antiproliferative effects of imatinib mesylate on CD34(+) progenitors in CML. Clinically, the levels of circulating soluble ST2, commonly considered a functional signature of IL-33 signaling in vivo, correlate with disease burden. Indeed, these elevated peripheral concentrations associated with a high Sokal score predictive of therapeutic outcome are normalized in patients in molecular remission. Finally, we evidenced a facilitating effect of IL-33 on in vivo maintenance of CD34(+) progenitors from patients with CML by using xenotransplant experiments in immunodeficient NOG mice, and we showed that engraftment of mouse BCR-ABL-transfected bone marrow progenitors was less efficient in IL-33-deficient mice compared with wild-type recipients. Taken together, our results provide evidence that IL-33/ST2 signaling may represent a novel cytokine-mediated mechanism contributing to CML progenitor growth and support a role for this pathway in CML maintenance and imatinib mesylate resistance.
This study aims to investigate the mechanisms of human mesenchymal stem cell-derived extracellular vesicles (HuMSC-EV)-induced proangiogenic paracrine effects after radiation injury. HuMSC-EV were ...locally administered in mice hindlimb following 80-Gy X-ray irradiation and animals were monitored at different time points. HuMSC-EV improved neovascularization of the irradiated tissue, by stimulating angiogenesis, normalizing cutaneous blood perfusion, and increasing capillary density and production of proangiogenic factors. HuMSC-EV also stimulated vasculogenesis by promoting the recruitment and differentiation of bone marrow progenitors. Moreover, HuMSC-EV improved arteriogenesis by increasing the mobilization of monocytes from the spleen and the bone marrow and their recruitment into the muscle, with a pro-inflammatory potential. Importantly, monocyte depletion by clodronate treatment abolished the proangiogenic effect of HuMSC-EV. The critical role of Ly6C(hi) monocyte subset in HuMSC-EV-induced neovascularization process was further confirmed using Ccr2
mice. This study demonstrates that HuMSC-derived EV enhances the neovascularization process in the irradiated tissue by increasing the production of proangiogenic factors, promoting the recruitment of vascular progenitor cells, and the mobilization of innate cells to the injured site. These results support the concept that HuMSC-EV might represent a suitable alternative to stem cells for therapeutic neovascularization in tissue repair.
Skin lesions caused by accidental exposure to radiation or by radiotherapy are a major clinical challenge. We evaluated the effect of bone marrow mononuclear cells (BMMNC) on collagen remodeling and ...vascular function in radiation-induced skin lesions in the acute and late phases in mice. We studied the effect of BMMNC transplantation in a mouse model of cutaneous radiation injury combining local skin gamma-irradiation and biopsy punch wound. Mice were first irradiated, punched and then BMMNC were intramuscularly administered. Seven days after injury, BMMNC promoted wound healing by (i) increasing re-epithelialization, tissue collagen density and mRNA levels of collagens 1A1, 1A2, and 3A1, and (ii) inhibiting the radiation-induced vascular activation and limiting interactions between leukocytes and the vascular endothelium compared with control. Importantly, BMMNC did not amplify the inflammatory response despite the infiltration of neutrophils and macrophages associated with the expression of IL-6 and MCP-1 mRNAs in the tissue. Remarkably, the beneficial effects of BMMNC therapy on matrix remodeling were maintained for 2 months. Furthermore, BMMNC injection restored vascular function in skin tissue by increasing vascular density and vascular permeability. This therapeutic strategy based on BMMNC injection protects against radiation-induced skin lesions by preventing vascular dysfunction and unfavorable remodeling in the acute and late phases.
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