Interactions of human mesenchymal stromal cells with peripheral blood mononuclear cells in a Mitogenic proliferation assay

Maryanne C. Herzig, Barbara A. Christy, Robbie K. Montgomery, Christopher P. Delavan, Katherine J. Jensen, Sarah E. Lovelace, Carolina Cantu, Christi L. Salgado, Andrew P. Cap, James A. Bynum

Research output: Contribution to journalArticlepeer-review

Abstract

Background: Immunomodulation by mesenchymal stromal cells (MSCs) is a potentially important therapeutic modality. MSCs suppress peripheral blood mononuclear cell (PBMC) proliferation in vitro, suggesting a mechanism for suppressing inflammatory responses in vivo. This study details the interactions of PBMCs and MSCs. Methods: Pooled human PBMCs and MSCs were co-cultured at different MSC:PBMC ratios and harvested from 0 to 120 h, with and without phytohaemagglutin A (PHA) stimulation. Proliferation of adherent MSCs and non-adherent PBMCs was assessed by quantitation of ATP levels. PBMC surface marker expression was analyzed by flow cytometry. Indoleamine 2,3-dioxygenase (IDO) activity was determined by kynurenine assay and IDO mRNA by RT-PCR. Cytokine release was measured by ELISA. Immunofluorescent microscopy detected MSC, PBMC, monocyte (CD14+) and apoptotic events. Results: PBMC proliferation in response to PHA gave a robust ATP signal by 72 h, which was suppressed by co-culture with densely plated MSCs. Very low level MSC seeding densities relative to PBMC number reproducibly stimulated PBMC proliferation. The CD4+/CD3+ population significantly decreased over time while the CD8+/CD3+ population significantly increased. No change in CD4+/CD8+ ratio is seen with high density MSC co-culture; very low density MSCs augment the changes seen in PHA stimulated PBMCs alone. IDO activity in MSCs co-cultured with PBMCs correlated with PBMC suppression. MSCs increased the secretion of IL-10 and IL-6 from stimulated co-cultures and decreased TNF-α secretion. In stimulated co-culture, low density MSCs decreased in number; fluorescence immunomicroscopy detected association of PBMC with MSC and phosphatidyl serine externalization in both cell populations. Conclusions: A bidirectional interaction between MSCs and PBMCs occurs during co-culture. High numbers of MSCs inhibit PHA-stimulated PBMC proliferation and the PBMC response to stimulation; low numbers of MSCs augment these responses. Low density MSCs are susceptible to attrition, apparently by PBMC-induced apoptosis. These results may have direct application when considering therapeutic dosing of patients; low MSC doses may have unintended detrimental consequences.

Original languageEnglish (US)
Article number113000
JournalJournal of Immunological Methods
Volume492
DOIs
StatePublished - May 2021

Keywords

  • Immunomodulation
  • Interactions
  • Mesenchymal stromal cells
  • PBMC
  • Stem

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

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