Clinical isolation and functional characterization of cord blood CD133+ hematopoietic progenitor cells

Authors

  • Giuseppina Bonanno,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Alessandro Perillo,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Sergio Rutella,

    Corresponding author
    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
      Sergio Rutella, MD, PhD, Department of Hematology, UNICATT Cord Blood Bank, Catholic University Medical School, Largo A.Gemelli 8, 00168, Rome, Italy; e-mail: srutella@rm.unicatt.it.
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  • Daniela Giovanna De Ritis,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Andrea Mariotti,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Maria Marone,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Franco Meoni,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Giovanni Scambia,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Giuseppe Leone,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Salvatore Mancuso,

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • Luca Pierelli

    1. 1From the Department of Gynecology and Obstetrics, the Department of Hematology and Blood Transfusion, and UNICATT Cord Blood Bank, Catholic University Medical School, Rome; the Department of Immunohematology and Blood Transfusion, ASL Viterbo, Viterbo; and the Mediterranean Institute of Neurology (NEUROMED), Pozzilli, Isernia, Italy.
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  • This study was supported in part by the Cord Blood Stem Cell Project (Fondazione Cassa di Risparmio di Roma, Rome, Italy) and by Ministero dell’Università e della Ricerca Scientifica e Tecnologica (MURST, Rome, Italy).

  • TRANSFUSION 2004;44:1087-1097.

Sergio Rutella, MD, PhD, Department of Hematology, UNICATT Cord Blood Bank, Catholic University Medical School, Largo A.Gemelli 8, 00168, Rome, Italy; e-mail: srutella@rm.unicatt.it.

Abstract

BACKGROUND:  Human cord blood is a relevant source of CD133+ HPCs. Clinical-scale isolation of human umbi-lical cord blood (UCB) CD133+ HPCs using immunomag-netic microbeads and the CliniMACS clinical cell isolator is reported. CD133+ HPCs isolated after large-scale pro-cessing were functionally characterized.

STUDY DESIGN AND METHODS:  Closed disposable sets were used to process nine different samples of RBC-reduced UCB nucleated cells. In-vitro hematopoietic assays and human xenografts in NOD/SCID mice were performed to assess the functional properties of isolated CD133+ cells. Different mixtures of human cytokines were tested for the ability to expand nascent CD133+ HPCs. Furthermore, freshly isolated CD133+ cells were conditioned in culture medium specifically tested to support in-vitro myogenesis or osteogenesis.

RESULTS:  Isolation procedures yielded the recovery of an average of 2.53 × 106 CD133+ HPCs with a mean recovery of 96 percent (referred to as RBC-reduced samples) and a final sample purity of 82 percent. Purified CD133+ cells had high cloning efficiency, had relevant long-term activity, and were capable of repopulating irradiated NOD/SCID mice. In 10-day stroma-free cultures, a 2-fold and 8.3-fold expansion of colony-forming cells (CFCs) and extended long-term culture-initiating cells, respectively, was obtained. Freshly isolated CD133+ cells differen-tiated into large nucleated cells expressing either myosin D or osteopontin (as revealed by RT-PCR and immuno-cytochemistry), with a protein/mRNA expression compar-able to or even higher than that observed in UCB CD133– nucleated cells in identical culture conditions.

CONCLUSION:  Collectively, clinical-scale isolation of UCB CD133+ cells provides a relevant amount of primi-tive HPCs with high hematopoietic activity and in-vitro mesenchymal potential.

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