Targeting acute allograft rejection by immunotherapy with ex vivo-expanded natural CD4+ CD25+ regulatory T cells.
BACKGROUND
Natural CD4CD25 regulatory T (Treg) cells have been implicated in suppressing alloreactivity in vitro and in vivo. We hypothesized that immunotherapy using ex vivo-expanded natural Treg could prevent acute allograft rejection in mice.


METHODS
Natural CD4+ CD25+ Treg were freshly purified from naive mice via automated magnetic cell sorter and expanded ex vivo by anti-CD3/CD28 monoclonal antibody (mAb)-coated Dynabeads. Suppression was assayed in vitro by mixed lymphocyte reaction and in vivo by targeting cardiac allograft rejection. Survival of Treg or effector T (Teff) cells after adoptive transfer in vivo was tracked by flow cytometry and all allografts were examined by histology and immunohistochemistry.


RESULTS
By day nine in culture, 26.6+/-5.3-fold of expansion was achieved by co-culture of fresh natural Treg with anti-CD3/CD28 mAb-coated Dynabeads and interleukin-2. Ex vivo-expanded Treg exerted stronger suppression than fresh ones towards alloantigens in vitro and prevented CD4 Teff-mediated but only delayed CD4+/CD8+ Teff-mediated heart allograft rejection in Rag-/- mice. Long-term surviving allografts showed no signs of acute or chronic rejection with graft-infiltrating Treg expressing CD25 and FoxP3. Infused Treg persisted and expanded long-term in vivo and trafficked through the peripheral lymphoid tissues. CD25 expression was dynamic in vivo: maintained CD25 expression on Treg was indicative for the preservation of allosuppression, while significantly enhanced CD25 expression on CD4+ effector T cells was most likely associated with T-cell expansion and graft rejection.


CONCLUSIONS
Therapeutic use of ex vivo-expanded natural CD4+ CD25+ Treg may be a feasible and nontoxic modality for controlling allograft rejection or perhaps inducing allograft tolerance.
Xia
Guliang
G
Department of Surgery-Organ Transplantation, Northwestern University Feinberg School of Medicine, Chicago, IL 60611, USA.
He
Jie
J
Zhang
Zheng
Z
Leventhal
Joseph R
JR
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D018106:CD28 Antigens / Q000276:immunology; D017252:CD3 Complex / Q000276:immunology; D015704:CD4 Antigens / Q000032:analysis; D018920:Coculture Techniques; D006084:Graft Rejection / Q000517:prevention & control / Q000628:therapy*; D016027:Heart Transplantation / Q000276:immunology; D016219:Immunotherapy, Adoptive; D053645:Interleukin-2 Receptor alpha Subunit / Q000032:analysis; D051379:Mice; D008863:Microspheres; D050378:T-Lymphocytes, Regulatory / Q000187:drug effects / Q000276:immunology / Q000637:transplantation*; D014184:Transplantation, Homologous
D000911:Antibodies, Monoclonal; D018106:CD28 Antigens; D017252:CD3 Complex; D015704:CD4 Antigens; D053645:Interleukin-2 Receptor alpha Subunit