Non-enzymatic dissociation of human mesenchymal stromal cells improves chemokine-dependent migration and maintains immunosuppressive function

Abhilok Garg, Diarmaid D Houlihan, Victoria Aldridge, Shankar Suresh, Ka Kit Li, Andrew L King, Rupesh Sutaria, Janine Fear, Ricky H Bhogal, Patricia F Lalor, Philip N Newsome

Research output: Contribution to journalArticlepeer-review

25 Citations (Scopus)


BACKGROUND AIMS: Human bone marrow-derived mesenchymal stromal cells (MSC) can suppress inflammation; therefore their therapeutic potential is being explored in clinical trials. Poor engraftment of infused MSC limits their therapeutic utility; this may be caused by MSC processing before infusion, in particular the method of their detachment from culture.

METHODS: Enzymatic methods of detaching MSC (Accutase and TrypLE) were compared with non-enzymatic methods (Cell Dissociation Buffer [CDB], ethylenediamine tetra-acetic acid and scraping) for their effect on MSC viability, chemokine receptor expression, multi-potency, immunomodulation and chemokine-dependent migration.

RESULTS: TrypLE detachment preserved MSC viability and tri-lineage potential compared with non-enzymatic methods; however, this resulted in near complete loss of surface chemokine receptor expression. Of the non-enzymatic methods, CDB detachment preserved the highest viability while retaining significant tri-lineage differentiation potential. Once re-plated, CDB-detached MSC regained their original morphology and reached confluence, unlike with the use of other non-enzymatic methods. Viability was significantly reduced with the use of ethylenediamine tetra-acetic acid and further reduced with the use of cell scraping. Addition of 1% serum during CDB detachment led to higher MSC numbers entering autophagy and increased MSC recovery after re-plating. TrypLE and CDB-detached MSC suppressed CD3(+)CD4(+)CD25(-) T-cell proliferation, although TrypLE-detached MSC exhibited superior suppression at 1:20 ratio. CDB detachment retained surface chemokine receptor expression and consequently increased migration to CCL22, CXCL12 and CCL4, in contrast with TrypLE-detached MSC.

CONCLUSIONS: This study demonstrates that non-enzymatic detachment of MSC with the use of CDB minimizes the negative impact on cell viability, multipotency and immunomodulation while retaining chemokine-dependent migration, which may be of importance in MSC delivery and engraftment in sites of injury.

Original languageEnglish
Pages (from-to)545-59
Number of pages15
Issue number4
Publication statusPublished - Apr 2014

Bibliographical note

Crown Copyright © 2014. Published by Elsevier Inc. All rights reserved.


  • Bone Marrow Cells
  • Cell Culture Techniques
  • Cell Differentiation
  • Cell Movement
  • Cell Proliferation
  • Cell Survival
  • Chemokines
  • Collagenases
  • Ethylenediamines
  • Humans
  • Immunosuppression
  • Mesenchymal Stromal Cells
  • Peptide Hydrolases


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