Cyclosporine A inhibits the expression of costimulatory molecules on in vitro-generated dendritic cells: association with reduced nuclear translocation of nuclear factor kappa B.
BACKGROUND
The maturation of dendritic cells (DC) is influenced by various factors, in particular cytokine-mediated signaling events. These include modulation of the activation of nuclear factor kappa B (NF-kappaB), which controls the transcription of genes encoding major histocompatibility complex (MHC) antigens, and costimulatory/accessory molecules for T-cell activation. Here, we investigated the influence of cyclosporine A (CsA) on the in vitro maturation of DC, and on the nuclear translocation and DNA binding of NF-kappaB.


METHODS
DC progenitors were propagated from mouse bone marrow in granulocyte-macrophage colony-stimulating factor (GM-CSF) or in GM-CSF plus either transforming growth factor (TGF)-beta or interleukin (IL)-4, in the presence or absence of CsA (1 microg/ml). After 5 days of culture, cell surface expression of MHC class I/II, CD40, CD80, and CD86 was analyzed by flow cytometry, and nuclear NF-kappaB proteins by electrophoretic mobility shift, antibody supershift, and Western blot assays. The antigen-presenting function of DC was determined in one-way mixed leukocyte reactions.


RESULTS
Exposure of replicating DC progenitors propagated in GM-CSF or GM-CSF+TGF-beta to CsA reduced costimulatory molecule expression, without affecting MHC antigen expression. Nuclear extracts from the CsA-treated DC revealed a decrease in nuclear translocation of NF-kappaB (p50). Mixed leukocyte reaction data were consistent with the flow cytometry and gel shift assay results, and showed reduced allostimulatory ability of the CsA-treated cells compared with untreated controls. Addition of IL-4 from the start of DC cultures conferred resistance to CsA-induced inhibition of NF-kappaB nuclear translocation and DC maturation.


CONCLUSIONS
CsA differentially inhibits the expression of key cell surface costimulatory molecules by in vitro-generated DC. This effect can be overcome, at least in part, by IL-4 and augmented by TGF-beta. The inhibition is linked to a decrease in nuclear translocation/DNA binding of NF-kappaB. Thus, CsA can alter the antigen-presenting function of DC for T-cell activation.
Lee
J I
JI
Thomas E. Starzl Transplantation Institute, University of Pittsburgh, Pennsylvania 15213, USA.
Ganster
R W
RW
Geller
D A
DA
Burckart
G J
GJ
Thomson
A W
AW
Lu
L
L
D000818:Animals; D015703:Antigens, CD / Q000096:biosynthesis; D051940:B7-2 Antigen; D001692:Biological Transport; D002467:Cell Nucleus / Q000378:metabolism*; D016572:Cyclosporine / Q000494:pharmacology*; D003713:Dendritic Cells / Q000187:drug effects* / Q000378:metabolism; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D007166:Immunosuppressive Agents / Q000494:pharmacology*; D015847:Interleukin-4 / Q000494:pharmacology; D008297:Male; D008562:Membrane Glycoproteins / Q000096:biosynthesis; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D016328:NF-kappa B / Q000378:metabolism*; D052002:NF-kappa B p50 Subunit; D016212:Transforming Growth Factor beta / Q000494:pharmacology
D015703:Antigens, CD; D051940:B7-2 Antigen; C497965:Cd86 protein, mouse; D007166:Immunosuppressive Agents; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D052002:NF-kappa B p50 Subunit; D016212:Transforming Growth Factor beta; D015847:Interleukin-4; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D016572:Cyclosporine