Dendritic cells genetically engineered with adenoviral vector encoding dnIKK2 induce the formation of potent CD4+ T-regulatory cells.
BACKGROUND
Immature dendritic cells (DC), characterized by low expression of both major histocompatibility complex class II antigens and co-stimulatory molecules, can be instrumental in the induction of peripheral tolerance. Because nuclear factor (NF)-kappa B is central to DC maturation, the authors engineered DC with an adenoviral vector (Adv) encoding for a kinase-defective dominant negative form of IKK2 (dnIKK2) to block NF-kappa B activation and inhibit DC maturation.


METHODS
DC were obtained by culturing bone marrow from Brown Norway (BN) rats with granulocyte-macrophage colony-stimulating factor and interleukin-4 for 11 days. To block NF-kappa B activation, at day 9, cells were transfected with AdV-dnIKK2. At day 11, cells were used as stimulators in primary mixed leukocyte reaction (MLR) with naive Lewis rat lymphocytes as responders. CD4+ T cells were purified from primary MLR and tested in secondary MLR with allogeneic mature DC and in co-culture MLR with naive lymphocytes. The tolerogenic potential of dnIKK2-DC was evaluated in vivo in a model of rat kidney allotransplantation.


RESULTS
DnIKK2-DC were immature and lacked any allostimulatory activity. T cells preexposed to allogeneic dnIKK2-DC were hyporesponsive to a secondary stimulation with mature DC and acquired potent regulatory properties, inhibiting naive T-cell proliferation toward allogeneic stimuli. Pretransplant infusion of allogeneic donor dnIKK2-DC prolonged the survival of a kidney allograft from the same allogeneic donor, without the need for immunosuppressive therapy.


CONCLUSIONS
Allogeneic DC, rendered immature by dnIKK2 transfection, induce in vitro differentiation of naive T cells into CD4+ T-regulatory cells, effective at low ratios with target cells, rendering them applicable for cellular therapy of immune-mediated abnormalities and for preventing transplant rejection.
Tomasoni
Susanna
S
Transplant Research Center Chiara Cucchi de Alessandri e Gilberto Crespi, Mario Negri Institute for Pharmacological Research, Bergamo, Italy.
Aiello
Sistiana
S
Cassis
Linda
L
Noris
Marina
M
Longaretti
Lorena
L
Cavinato
Regiane A
RA
Azzollini
Nadia
N
Pezzotta
Anna
A
Remuzzi
Giuseppe
G
Benigni
Ariela
A
D000256:Adenoviridae; D000818:Animals; D001854:Bone Marrow Cells / Q000276:immunology; D016026:Bone Marrow Transplantation / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology*; D002454:Cell Differentiation; D003713:Dendritic Cells / Q000276:immunology*; D005818:Genetic Engineering / Q000379:methods; D005822:Genetic Vectors; D051550:I-kappa B Kinase; D016030:Kidney Transplantation / Q000276:immunology*; D007959:Lymphocyte Culture Test, Mixed; D023421:Models, Animal; D017346:Protein Serine-Threonine Kinases / Q000235:genetics*; D051381:Rats; D011914:Rats, Inbred BN; D011917:Rats, Inbred Lew; D013601:T-Lymphocytes / Q000166:cytology / Q000276:immunology; D014162:Transfection / Q000379:methods; D014184:Transplantation, Homologous
D017346:Protein Serine-Threonine Kinases; D051550:I-kappa B Kinase