naa a22 4500 510739237 CHVBK 20180411083022.0 cr unu---uuuuu 180411e20130601xx s 000 0 eng 10.1007/s12015-011-9316-9 doi (NATIONALLICENCE)springer-10.1007/s12015-011-9316-9 A New Methodological Sequence to Expand and Transdifferentiate Human Umbilical Cord Blood Derived CD133+ Cells into a Cardiomyocyte-like Phenotype [Elektronische Daten] [Yu-Xin Cui, Wael Kafienah, M-S Suleiman, Raimondo Ascione] Transplantation of antigenic-separated stem cells for human cardiovascular diseases such as myocardial infarction needs to be supported by experimental studies that allow refinement of the procedure. In this study we investigated optimising a protocol for the expansion and subsequent differentiation of human umbilical cord blood (HUCB) derived CD133+ stem cells into a cardiomyocyte-like lineage. CD133+ cells from HUCB were selected first by immunomagnetic separation and their purity was confirmed by flow cytometry analysis. For expansion and differentiation we developed a novel culture medium recipe that involves sequential signalling factors. Briefly, CD133+ cells were expanded for 6days under optimal serum-free conditions in combination with fibronectin and assessed by microscopy and AlamarBlue proliferation assay. Expanded CD133+ cells were then plated in a cardiac differentiation promoting medium and cultured up to 4weeks. With this protocol HUCB-CD133+ cells can be regularly expanded in serum-free medium to obtain recovery and growth in vitro up to 6 folds. The addition of recombinant human thrombopoietin to the remaining factors of the expanding medium was associated with larger cell expansion. Expanded UCB CD133+ cells showed a cardiomyocyte-like phenotype following differentiation in vitro through expressing intracellular cardiac specific markers including cardiac-specific α-actin, myosin heavy chain and troponin I. This change in phenotype was associated with the expression of cardiac-specific transcription factors Gata-4 and MEF2C. In addition, the change in phenotype was associated with an upregulation of nuclear receptor transcription factors including PPAR α, PPARγ, RXR α and RXRβ. We believe our protocol represents a significant advancement and overcome the technical hurdle of deriving cardiomyogenic-like cells from HUCB CD133+ stem cells. In addition, it has the required attributes of simplicity and consistency. This will permit more robust manipulation of these cells towards better engraftment and repair in patients with myocardial infarction. Springer Science+Business Media, LLC, 2011 Human cord blood nationallicence CD133 nationallicence Isolation nationallicence Cardiac transdifferentiation nationallicence Nuclear receptor transcription factors nationallicence Cui Yu-Xin Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut Kafienah Wael Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut Suleiman M-S Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut Ascione Raimondo Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut Stem Cell Reviews and Reports Humana Press Inc 9/3(2013-06-01), 350-359 1550-8943 9:3<350 2013 9 12015 https://doi.org/10.1007/s12015-011-9316-9 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s12015-011-9316-9 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Cui Yu-Xin Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut NATIONALLICENCE 700 1- Kafienah Wael Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut NATIONALLICENCE 700 1- Suleiman M-S Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut NATIONALLICENCE 700 1- Ascione Raimondo Bristol Heart Institute, University of Bristol, Level 7, Bristol Royal Infirmary, Upper Maudlin Street, BS2 8HW, Bristol, UK aut NATIONALLICENCE 773 0- Stem Cell Reviews and Reports Humana Press Inc 9/3(2013-06-01), 350-359 1550-8943 9:3<350 2013 9 12015 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer