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Large-Scale Automated Hollow-Fiber Bioreactor Expansion of Umbilical Cord-Derived Human Mesenchymal Stromal Cells for Neurological Disorders

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Abstract

Neurodegenerative disorders present a broad group of neurological diseases and remain one of the greatest challenges and burdens to mankind. Maladies like amyotrophic lateral sclerosis, Alzheimer’s disease, stroke or spinal cord injury commonly features astroglia involvement (astrogliosis) with signs of inflammation. Regenerative, paracrine and immunomodulatory properties of human mesenchymal stromal cells (hMSCs) could target the above components, thus opening new therapeutic possibilities for regenerative medicine. A special interest should be given to hMSCs derived from the umbilical cord (UC) tissue, due to their origin, properties and lack of ethical paradigms. The aim of this study was to establish standard operating and scale-up good manufacturing practice (GMP) protocols of UC-hMSCs isolation, characterization, expansion and comparison of cells’ properties when harvested on T-flasks versus using a large-scale bioreactor system. Human UC-hMSCs, isolated by tissue explant culture technique from Wharton’s jelly, were harvested after reaching 75% confluence and cultured using tissue culture flasks. Obtained UC-hMSCs prior/after the cryopreservation and after harvesting in a bioreactor, were fully characterized for “mesenchymness” immunomodulatory, tumorigenicity and genetic stability, senescence and cell-doubling properties, as well as gene expression features. Our study demonstrates an efficient and simple technique for large scale UC-hMSCs expansion. Harvesting of UC-hMSCs’ using classic and large scale methods did not alter UC-hMSCs’ senescence, genetic stability or in vitro tumorigenicity features. We observed comparable growth and immunomodulatory capacities of fresh, frozen and expanded UC-hMSCs. We found no difference in the ability to differentiate toward adipogenic, osteogenic and chondrogenic lineages between classic and large scale UC-hMSCs expansion methods. Both, methods enabled derivation of genetically stabile cells with typical mesenchymal features. Interestingly, we found significantly increased mRNA expression levels of neural growth factor (NGF) and downregulated insulin growth factor (IGF) in UC-hMSCs cultured in bioreactor, while IL4, IL6, IL8, TGFb and VEGF expression levels remained at the similar levels. A culturing of UC-hMSCs using a large-scale automated closed bioreactor expansion system under the GMP conditions does not alter basic “mesenchymal” features and quality of the cells. Our study has been designed to pave a road toward translation of basic research data known about human UC-MSCs for the future clinical testing in patients with neurological and immunocompromised disorders. An industrial manufacturing of UC-hMSCs next will undergo regulatory approval following advanced therapy medicinal products (ATMP) criteria prior to clinical application and approval to be used in patients.

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Acknowledgements

Supported by Technology Agency, Czech Republic (TACR) Program DELTA, project TF03000037 and The Ministry of Industry and Trade of the CR (MPO), Aplikace (III. výzva) project CZ.01.1.02/0.0/0.0/16_084/0010317.

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Authors and Affiliations

  1. International Clinical Research Center, St. Anne’s University Hospital, Pekarska 53, 656 91, Brno, Czech Republic

    Ladislava Vymetalova, Tereza Kucirkova, Lucia Knopfova & Petr Benes

  2. PrimeCell Bioscience Inc., Dr. Slabihoudka 6232/11, 708 00, Ostrava-Poruba, Czech Republic

    Veronika Pospisilova, Tomas Kasko, Zdenek Koristek & Serhiy Forostyak

  3. National Tissue Centre Inc., Palachovo náměstí 726/2, 625 00, Brno, Czech Republic

    Veronika Pospisilova, Tomas Kasko & Serhiy Forostyak

  4. Department of Transfusion & Tissue Medicine, University Hospital Brno, Jihlavska 20, 625 00, Brno, Czech Republic

    Hana Lejdarova

  5. Department of Medical Genetics, University Hospital Brno, 625 00, Brno, Czech Republic

    Eva Makaturova & Petr Kuglik

  6. Institute of Experimental Biology, Faculty of Science, Masaryk University, Kotlarska 2, 611 37, Brno, Czech Republic

    Petr Kuglik

  7. Institute of Animal Physiology and Genetics, v.v.i, Czech Academy of Sciences, Veveri 97, 602 00, Brno, Czech Republic

    Veronika Oralova & Eva Matalova

  8. PrimeCell Advanced Therapy Inc., Palachovo nam. 726/2, 625 00, Brno, Czech Republic

    Zdenek Koristek & Serhiy Forostyak

Authors
  1. Ladislava Vymetalova
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  2. Tereza Kucirkova
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  13. Serhiy Forostyak
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Corresponding authors

Correspondence to Zdenek Koristek or Serhiy Forostyak.

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All umbilical cord samples were collected at the Department of Obstetrics and Gynecology, University Hospital Brno with informed consent from mothers and approval from Ethics Committee of University Hospital Brno.

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Special Issue: In Honour of Professor Eva Sykova.

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Vymetalova, L., Kucirkova, T., Knopfova, L. et al. Large-Scale Automated Hollow-Fiber Bioreactor Expansion of Umbilical Cord-Derived Human Mesenchymal Stromal Cells for Neurological Disorders. Neurochem Res 45, 204–214 (2020). https://doi.org/10.1007/s11064-019-02925-y

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  • DOI: https://doi.org/10.1007/s11064-019-02925-y

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