Ex Vivo Expansion of Human Alloantigen-Reactive Regulatory T Cells Using Monocyte-Derived Dendritic Cells versus CD40L-Stimulated B Cells.
1Thomas E. Starzl Transplantation Institute Department of Surgery, University of Pittsburgh school of medicine, Pittsburgh, PA
2Department of Surgery, UCSF, San Francisco, CA.
Meeting: 2016 American Transplant Congress
Abstract number: A35
Keywords: T cells
Session Information
Session Type: Poster Session
Date: Saturday, June 11, 2016
Session Time: 5:30pm-7:30pm
Presentation Time: 5:30pm-7:30pm
Location: Halls C&D
Background:
Alloantigen-reactive murine and human CD4+CD25+FOXP3+ regulatory T cells (arTreg) are more potent in promoting long-term allograft survival than polyclonal Treg in experimental models. Dendritic cells (DCs) are well known professional antigen-presenting cells (APC) specialized to initiate and regulate immunity. CD40L-stimulated B cells (CD40L-sBc) have been reported to expand human arTreg. However, no direct comparison of these APCs in their ability to expand arTreg has been reported. In order to establish a protocol to generate large numbers of human arTreg with potent specific suppressive capacity for therapeutic application, we compared immature monocyte-derived DC (mono-DC), LPS-matured DC, and CD40L-sBc in their ability to expand arTreg.
Methods & Results:
To generate mono-DCs, immunobead-selected blood CD14+ cells were cultured with GM-CSF and IL-4 for 7 days. To mature mono-DCs, LPS was added for the last day to the mono-DC differentiation cultures; to make CD40L-sBc, flow-sorted CD20+ cells were cultured with irradiated 3T3-CD40L cells, IL-4 and cyclosporine A for 7 days before restimulation with CD40L-3T3 and IL-4 for an additional 3 days. Then these APCs were irradiated and co-cultured with flow-sorted CD4+CD25+CD127– Tregs and IL-2 for 9 to 11 days. We observed that LPS-matured DC expressed higher levels of CD80 and CD86 than immature DC and CD40L-sBc (figure A). Treg cultured with LPS-matured DCs were expanded ~40 fold, while those with immature DC and CD40L-sBc were expanded 10- to 30- fold (figureB). Expression of FOXP3 and CTLA-4 was higher in Treg expanded by either DCs or LPS-matured DCs. In addition, Treg expanded by DCs were superior suppressors of T cell proliferation stimulated by either anti-CD3/CD28 beads or allo-sBc. 
Conclusions:
LPS-matured human mono-DCs are superior to immature DC and CD40L-sBc for expansion of functionally suppressive FOXP3+ Treg.
CITATION INFORMATION: Zhang H, Zhao Y, Lu L, Tang Q, Ezzelarab M, Thomson A. Ex Vivo Expansion of Human Alloantigen-Reactive Regulatory T Cells Using Monocyte-Derived Dendritic Cells versus CD40L-Stimulated B Cells. Am J Transplant. 2016;16 (suppl 3).
To cite this abstract in AMA style:
Zhang H, Zhao Y, Lu L, Tang Q, Ezzelarab M, Thomson A. Ex Vivo Expansion of Human Alloantigen-Reactive Regulatory T Cells Using Monocyte-Derived Dendritic Cells versus CD40L-Stimulated B Cells. [abstract]. Am J Transplant. 2016; 16 (suppl 3). https://atcmeetingabstracts.com/abstract/ex-vivo-expansion-of-human-alloantigen-reactive-regulatory-t-cells-using-monocyte-derived-dendritic-cells-versus-cd40l-stimulated-b-cells/. Accessed May 11, 2025.« Back to 2016 American Transplant Congress