Mechanistic insights into achievement of cardiac allograft long-term survival by treatment with immature dendritic cells and sub-dose sirolimus.
BACKGROUND
Administration of immature dendritic cells (DC) prolongs but does not result in indefinite allograft survival. We attempted to achieve this goal by adding a sub-therapeutic dose of immunosuppression.


METHODS
DC propagated from B10 (H-2(b)) mouse bone marrow (BM) were transfected with nuclear factor-kappaB (NF-kappaB)-binding-site-specific oligodeoxyribonucleotide (ODN). The allostimulatory activity of transfected and normal DC were examined in mixed-lymphocyte reaction (MLR) and cytotoxic T-lymphocyte (CTL) assays in vitro, and their influence on allograft survival by systemic administration of DC in vivo.


RESULTS
Transfection of DC with NF-kappaB ODN resulted in complete abrogation of NF-kappaB activity and inhibition of co-stimulation. Allogeneic (C3H, H-2(k)) T cells stimulated by ODN DC demonstrated impairment in MLR and CTL activity. Administration of ODN DC significantly prolonged B10 allograft survival. In contrast to cyclosporine, which failed to enhance the effect of ODN DC, a combination of ODN DC with sirolimus at 6 mg/kg/day for 6 days achieved long-term survival in all allografts. This was associated with low CTL activity of either graft-infiltrating cells or splenic T cells and increased TUNEL-positive cells in T-cell areas of recipient mesenteric lymph nodes. Analysis of transcription factor nuclear translocation with Cellomics indicated that stimulation with ODN DC showed inhibited T-cell nuclear translocation of signal transducer and activator of transcription (Stat)1 and Stat3, extracellular signal-related kinase (ERK) and activating transcription factor (ATF)-2, but not NF-kappaB and P38, compared with mature DC. The selective inhibition was enhanced by sirolimus, but not cyclosporine.


CONCLUSIONS
Sirolimus enhances immature DC tolerogenicity by induction of T-cell apoptosis, and promotes immature DC-induced inhibition of Stat1, ERK and ATF-2 activation.
Tao
Ran
R
Thomas E. Starzl Transplantation Institute and Department of Surgery, University of Pittsburgh, Pittsburgh, Pennsylvania 15261, USA.
Wang
Lianfu
L
Chen
Cheng-Hsu
CH
Wang
Shi-Ho
SH
Demarco
Richard A
RA
Lotze
Michael T
MT
Thai
Ngoc L
NL
Fung
John J
JJ
Lu
Lina
L
Qian
Shiguang
S
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D003713:Dendritic Cells / Q000187:drug effects / Q000637:transplantation*; D005434:Flow Cytometry; D006085:Graft Survival / Q000187:drug effects* / Q000502:physiology*; D016027:Heart Transplantation; D007166:Immunosuppressive Agents / Q000008:administration & dosage*; D007959:Lymphocyte Culture Test, Mixed; D051379:Mice; D016328:NF-kappa B / Q000187:drug effects / Q000378:metabolism; D009838:Oligodeoxyribonucleotides / Q000494:pharmacology; D020123:Sirolimus / Q000008:administration & dosage*; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects / Q000502:physiology*; D014157:Transcription Factors / Q000187:drug effects; D014162:Transfection
D007166:Immunosuppressive Agents; D016328:NF-kappa B; D009838:Oligodeoxyribonucleotides; D014157:Transcription Factors; D020123:Sirolimus