Cytométrie Paris


Année de publication : 2017

Zofia Maciorowski, Pratip K Chattopadhyay, Paresh Jain (2017 Apr 4)

Basic Multicolor Flow Cytometry.

Current protocols in immunology : 5.4.1-5.4.38 : DOI : 10.1002/cpim.26 En savoir plus

Multicolor flow cytometry is a rapidly evolving technology that uses multiple fluorescent markers to identify and characterize cellular subpopulations of interest, allowing rapid analysis on tens of thousands of cells per second, with the possibility of isolating pure, viable populations by cell sorting for further experimentation. This unit covers the tools needed by the beginning immunologist to plan and run multicolor experiments, with information on fluorochromes and their characteristics, spectral spillover, compensation and spread, instrument and reagent variables, and the basic elements of multicolor panel design. Protocols to quantify and maximize sensitivity by titration of reagents and optimization of instrument settings, as well as basic surface and intracellular cell staining, are included. © 2017 by John Wiley & Sons, Inc.

Coralie L Guerin, Elisa Rossi, Bruno Saubamea, Audrey Cras, Virginie Mignon, Jean-Sébastien Silvestre, David M Smadja (2017 Mar 18)

Human very Small Embryonic-like Cells Support Vascular Maturation and Therapeutic Revascularization Induced by Endothelial Progenitor Cells.

Stem cell reviews and reports : 552-560 : DOI : 10.1007/s12015-017-9731-7 En savoir plus

Very small embryonic-like stem cells (VSELs) are major pluripotent stem cells defined as cells of small size being Lineage- negative, CD133-positive, and CD45-negative. We previously described that human bone marrow VSELs were able to differentiate into endothelial cells and promoted post-ischemic revascularization in mice with surgically induced critical limb ischemia. In the present work, we isolated bone marrow VSELs from patients with critical limb ischemia and studied their ability to support endothelial progenitor cells therapeutic capacity and revascularization potential. Sorted bone marrow VSELs cultured in angiogenic media were co-injected with endothelial progenitor cells and have been show to trigger post-ischemic revascularization in immunodeficient mice, and support vessel formation in vivo in Matrigel implants better than human bone marrow mesenchymal stem cells. In conclusion, VSELs are a potential new source of therapeutic cells that may give rise to cells of the endothelial and perivascular lineage in humans. VSELs are the first real vasculogenic stem cells able to differentiate in endothelial and perivascular lineage in human adult described from now. Thus, because VSELs presence have been proposed in adult tissues, we think that VSELs are CD45 negative stem cells able to give rise to vascular regeneration in human tissues and vessels.

Coralie L Guerin, Adeline Blandinières, Benjamin Planquette, Jean-Sébastien Silvestre, Dominique Israel-Biet, Olivier Sanchez, David M Smadja (2017 Mar 13)

Very Small Embryonic-like Stem Cells Are Mobilized in Human Peripheral Blood during Hypoxemic COPD Exacerbations and Pulmonary Hypertension.

Stem cell reviews and reports : 561-566 : DOI : 10.1007/s12015-017-9732-6 En savoir plus

Very small embryonic-like stem cells (VSELs) are major pluripotent stem cells involved in vascular and tissue regeneration and constitute a recruitable pool of stem/progenitor cells with putative instrumental role in organ repair. Here, we hypothesized that VSELs might be mobilized from the bone marrow (BM) to peripheral blood (PB) in patients with hypoxic lung disease or pulmonary hypertension (PH). The objective of the present study was then to investigate the changes in VSELs number in peripheral blood of patients with hypoxic lung disease and PH. We enrolled 26 patients with Chronic Obstructive Pulmonary Disease (COPD) with or without hypoxemia, 13 patients with PH and 20 controls without any respiratory or cardiovascular diseases. In PH patients, VSELs levels have been determined during right heart catheterization in pulmonary blood and PB. For this purpose, mononuclear cells were separated by density gradient and VSELs have been quantified by using a multiparametric flow cytometry approach. The number of PB-VSELs in hypoxic COPD patients was significantly increased compared with non-hypoxic COPD patients or controls (p = 0.0055). In patients with PH, we did not find any difference in VSELs numbers between arterial pulmonary blood and venous PB (p = 0.93). However, we found an increase in VSELs in the peripheral blood of patients with PH (p = 0.03). In conclusion, we unraveled that circulating VSELs were increased in peripheral blood of patients with hypoxic COPD or with PH. Thus, VSELs may serve as a reservoir of pluripotent stem cells that can be recruited into PB and may play an important role in promoting lung repair.

Eléonore Blondiaux, Laetitia Pidial, Gwennhael Autret, Gabriel Rahmi, Daniel Balvay, Etienne Audureau, Claire Wilhelm, Coralie L Guerin, Patrick Bruneval, Jean-Sébastien Silvestre, Philippe Menasché, Olivier Clément (2017 Feb 4)

Bone marrow-derived mesenchymal stem cell-loaded fibrin patches act as a reservoir of paracrine factors in chronic myocardial infarction.

Journal of tissue engineering and regenerative medicine : 3417-3427 : DOI : 10.1002/term.2255 En savoir plus

The combination of mesenchymal stem cells and tissue-engineered fibrin patches improves the therapeutic efficacy of stem cells. In vivo cardiac magnetic resonance (4.7 Tesla) and ex vivo high-spatial resolution CMR were used to track the fate of human bone marrow-derived mesenchymal stem cell (BMSC) delivered on an epicardial scaffold and more specifically assess their potential intramyocardial migration. Fifty-seven nude rats underwent permanent coronary artery ligation. Two months later, those with a left ventricular ejection fraction ≤55% were randomly allocated to receive a patch loaded with human BMSC (BMSC-P, n = 10), a patch loaded with BMSCs labelled with iron oxide nanoparticles (BMSC*-P, n = 12), an acellular patch (A-P, n = 8) or to serve as sham-operated animals (SHAM, n = 7). BMSC secretion of cytokines and growth factors was evaluated with flow-cytometry. Cardiac functional parameters of cell-treated groups (BMSC*-P and BMSC-P) yielded significantly better outcomes than the SHAM group (p = 0.044 and p = 0.026, respectively, for ejection fraction). Angiogenesis was higher in the cell-patch than in control groups (e.g. BMSC*P vs. SHAM: p = 0.007). No BMSCs were identified into the myocardium on cardiac magnetic resonance or histological sections, although persisting BMSCs were identified on the epicardial surface 21 days post-transplantation in 10% of rats hearts (Lamin A/C and CD90 positive). Cytokine and growth factor profiling demonstrated an increase in their release by cells seeded in patches. The absence of stem cell migration into the myocardium and the persistence of stem cells on the epicardial surface suggest that fibrin patches are likely to act predominantly as reservoirs of paracrine factors. Copyright © 2017 John Wiley & Sons, Ltd.