Immature and Maturation-Resistant Human Dendritic Cells Generated from Bone Marrow Require Two Stimulations to Induce T Cell Anergy In Vitro

Derya Unutmaz,Thomas G Berger,Hendrik Schulze-Koops,Ester Müller,Michaela Schäfer,Manfred B Lutz

doi:10.1371/journal.pone.0006645

Abstract

Immature dendritic cells (DC) represent potential clinical tools for tolerogenic cellular immunotherapy in both transplantation and autoimmunity. A major drawback in vivo is their potential to mature during infections or inflammation, which would convert their tolerogenicity into immunogenicity. The generation of immature DC from human bone marrow (BM) by low doses of GM-CSF (lowGM) in the absence of IL-4 under GMP conditions create DC resistant to maturation, detected by surface marker expression and primary stimulation by allogeneic T cells. This resistence could not be observed for BM-derived DC generated with high doses of GM-CSF plus IL-4 (highGM/4), although both DC types induced primary allogeneic T cell anergy in vitro. The estabishment of the anergic state requires two subsequent stimulations by immature DC. Anergy induction was more profound with lowGM-DC due to their maturation resistance. Together, we show the generation of immature, maturation-resistant lowGM-DC for potential clinical use in transplant rejection and propose a two-step-model of T cell anergy induction by immature DC.

Highlights

Establishment of immunological tolerance is the ultimate goal for causative treatments of autoimmune diseases, allergies and transplant rejections
Human bone marrow (BM)-dendritic cells (DC) generated under low doses of GM-CSF (lowGM) conditions are maturation resistant
We described a method how murine DC can be generated with low doses of GM-CSF from BM cells without undergoing spontaneous maturation

Summary

Introduction

Establishment of immunological tolerance is the ultimate goal for causative treatments of autoimmune diseases, allergies and transplant rejections. With the diversity of T cell tolerance mechanisms, there might not be a single tolerogenic DC type able to control all of these mechanisms. The various mechanisms of T cell tolerance can be subdivided into T cell-intrinsic mechanisms, such as T cell anergy, deletion, immune deviation and T cell extrinsic control by induction of regulatory T cells (T regs) [4,5,6,7]. T cell anergy induction has been originally described with T cell clones, i.e. cells that have undergone at least one stimulation before employed in the anergy experiments [8]. Subsequent exposure of such primed, but resting CD4+ T cells cross-linked to CD3 resulted in an anergic state of these cells. Previous studies indicate T cell anergy could be induced by a single application of orally applied ovalbumin antigen [9] or superantigen [10]

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Results

Conclusion