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Comparison between isolation protocols highlights intrinsic variability of human umbilical cord mesenchymal cells

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Abstract

Mesenchymal stem cells (MSCs) though multipotent exhibit limited lifespan in vitro, with progressive reduction in capacity for self-renewal leading to irreversible arrest of cell division, which limits their use for therapeutic purposes. Human umbilical cord wall MSCs are easy to process and proliferate rapidly in culture, but variability of individual samples and impact upon in vitro expansion and aging processes is unknown. We compared isolation protocols to determine which one yields the highest number of viable cells with the best proliferation capacity. Three different protocols were tested: two were enzymatic procedures and one explant method. Isolated cells were evaluated in terms of proliferation, differentiation capacity, and phenotype. All samples were processed using one or more protocols. After passage 2 adherent cells displayed standard phenotypic and differentiation characteristics of MSCs, but our results show that isolating cells directly from Wharton’s jelly is more advantageous. Cells obtained from explants presented similar characteristics to those from enzymatic protocols, but always reached proliferation arrest earlier, irrespective of initial population doubling times. From the same sample, cells obtained with enzymatic protocol ii reached later passages while exhibiting shorter doubling times in culture than cells from other protocols, that is, took longer to reach senescence. More important, each individual MSC sample exhibited different population doubling rates and reached senescence at different passages, irrespective of protocol. Thus, even when in strict conformity with procedures and quality control, each cord sample shows a unique behavior, a finding that should be taken into account when planning for therapeutic approaches.

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Acknowledgments

We would like to acknowledge the contribution of Maria Alves, Isis Mozetic, and Érica Moreira that have helped make this study possible. We thank Andreia Kondo and the team of nurses at our local Public Umbilical Cord Blood Bank for their help to obtain the samples. We thank Dr. Luiz Sardinha, Dr. Luciana Marti, and Dr. Andrea Sertié for expert technical assistance and insightful suggestions. UNIEMP and CAPES have financed graduate fellowships. Anna Carla Goldberg is a recipient of personal fellowship from CNPq. We are indebted to UC donors and acknowledge the generous support by the Ruhman family.

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Authors and Affiliations

  1. Hospital Israelita Albert Einstein, IIEP, São Paulo, Brazil

    Fernanda Vieira Paladino, Joana Silveira Peixoto-Cruz, Carolina Santacruz-Perez & Anna Carla Goldberg

  2. Animal Cell Technology Unit - Instituto de Tecnologia Química e Biológica, Oeiras, Portugal

    Joana Silveira Peixoto-Cruz

Authors
  1. Fernanda Vieira Paladino
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  2. Joana Silveira Peixoto-Cruz
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  3. Carolina Santacruz-Perez
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  4. Anna Carla Goldberg
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Correspondence to Anna Carla Goldberg.

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Paladino, F.V., Peixoto-Cruz, J.S., Santacruz-Perez, C. et al. Comparison between isolation protocols highlights intrinsic variability of human umbilical cord mesenchymal cells. Cell Tissue Bank 17, 123–136 (2016). https://doi.org/10.1007/s10561-015-9525-6

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  • DOI: https://doi.org/10.1007/s10561-015-9525-6

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