Human Cytokine Expression Profile in Various Conditioned Media for In Vitro Expansion Bone Marrow and Umbilical Cord Blood Immunophenotyped Mesenchymal Stem Cells F. Jenhani, V. Durand, N. Ben Azouna, S. Thallet, T. Ben Othmen, M. Bejaoui, and J. Domenech ABSTRACT Introduction. Bone marrow (BM) represents the major source of mesenchymal stem cells (MSCs); however, umbilical cord blood (UCB) MSCs have some advantages over BM, such as a higher differentiation capability and noninvasive collection methods. Objectives. We compared antigen expression and cytokine-secretion by MSC from BM and UCB expanded with media supplemented with fetal bovine serum (FBS) or human platelet lysate (HPL). Materials and Methods. We compared protocols for the expansion of hMSC starting from samples of BM or UCB by morphological analysis, calculation of population doubling numbers, and cytometry techniques using monoclonal antibodies (BD Biosciences). Using the last technique, cytokines were detected in brain homogenate supernatant fluids of MSC cultured in various media, using the Bio-Plex cytokine assay system (BD Biosciences). Results. Calculating the number population doubling (PD) and colony-forming unit- 1 fibroblast (CFU-F) assays showed significantly better expansion with HPL compared with a selected batch of FBS and within fewer days: PD about 5 for 10%HPL versus 25 for fibroblast growth factor2 (FGF2) medium. By flow cytometry, we observed a greater number of BM MSCs compared with UCB MSCs, as well as differences in the expression of some MSC antigens, particularly CD105, CD90, and CD31. Analysis of cytokines: FGFb, RANTES, VEGF, IL-6, IL-8, G-CSF, and GM-CSF showed only some of them to be expressed: namely, IL-6, IL-8, and VEGF. MSCs derived from UCB showed low concentrations of these cytokines compared with MSCs derived from BM. M ESENCHYMAL stem cells (MSCs), which were initially described in bone marrow (BM), represent a rare population of multipotent progenitors that can differentiate to adipocytes, osteoblasts, chondrocytes, and vascular-smooth muscle–like hematopoietic supportive stromal cells. 1–4 Single cells have been shown to be capable of multilineage differentiation 5,6 and in vivo functional reconstitution of injured tissues in preclinical 7 as well as in clinical 8,9 settings. Accordingly, to culture MSCs, some trials have used serum- free animal media containing combinations of growth fac- tors. 23 Other alternatives supplement the media with human blood– derived platelet lysates or sera from autologous or allogenic donors 24 –28 to create culture conditions that more closely mimic the human environment. However, it is not yet clear how these culture conditions influence the coloning forming unit–fibroblasts (CFu-F), the population doubling time (PDT), the phenotype, and the cytokines secreted into the supernates by MSC populations. From the Cell Immunology, Cytometry and Therapy Cell Laboratory (F.J.), Tunisian National Blood Center (N.B.A.), Tunis, Tunisia; Becton Dickinson Rungis (V.D., S.T.), Paris, France; Tunisian National Bone Marrow Center (T.B.O., M.B.), Tunis, Tunisia; Faculty of Medicine (J.D.), Tours France. Address reprint requests to Prof. Faouzi Jenhani, Laboratoire d’Immunologie Cellulaire et de Cytométrie, Centre National de Transfusion Sanguine, Bab Saadoun, PB 294, El Manar II, 2092, Tunisia. E-mail: faouzi.janhani@rns.tn or Faouzi.jenhani@yahoo.fr © 2011 Published by Elsevier Inc. 0041-1345/–see front matter 360 Park Avenue South, New York, NY 10010-1710 doi:10.1016/j.transproceed.2011.01.021 Transplantation Proceedings, 43, 639 – 643 (2011) 639