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, 116 (9), 1454-9

PGE(2) Transiently Enhances DC Expression of CCR7 but Inhibits the Ability of DCs to Produce CCL19 and Attract Naive T Cells

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PGE(2) Transiently Enhances DC Expression of CCR7 but Inhibits the Ability of DCs to Produce CCL19 and Attract Naive T Cells

Ravikumar Muthuswamy et al. Blood.

Abstract

Prostaglandin E(2) (PGE(2)) is an inflammatory mediator often used to increase CCR7 expression in the dendritic cells (DCs) used as cancer vaccines and to enhance their responsiveness to lymph node-associated chemokines. Here, we show that high surface expression of CCR7 on PGE(2)-matured DCs is associated with their suppressed production of the endogenous CCR7 ligand, CCL19, and is reversible by exogenous CCL19. In contrast to the PGE(2)-matured DCs, DCs matured in the presence of toll-like receptor (TLR) ligands and interferons produce high levels of both CCL19 and CCR7 mRNA/protein, but show selectively reduced expression of surface CCR7, which is compensated after DC removal from the CCL19-rich maturation environment. In accordance with these findings, PGE(2)-matured DCs show significantly higher in vitro migratory responsiveness to lymph node-associated chemokines directly after DC generation, but not after additional short-term culture in vitro, nor in vivo in patients injected with (111)indium-labeled DCs. The differences in CCL19-producing ability imprinted during DC maturation result in their different abilities to attract CCR7(+) naive T cells. Our data help to explain the impact of PGE(2) on CCR7 expression in maturing DCs and demonstrate a novel mechanism of regulatory activity of PGE(2), mediated by the inhibition of DCs ability to attract naive T cells.

Figures

Figure 1
Figure 1
Transient elevation of CCR7 expression and in vitro migratory responsiveness to lymph node–associated chemokines in PGE2-matured DCs. (A left) Surface CCR7 expression in DCs matured into αDC1s or sDCs in 9 different donors. (A right) In vitro migratory response of αDC1s and sDCs to CCL21, a secondary lymphoid organ chemokine (data from 6 different donors shown as mean ± SD). (B) Surface expression of CCR7 protein on αDC1s and sDCs at 0 (left) and 24 hours after completion of maturation and replating in the absence of the maturation-inducing factors (right). (C) In vitro migratory response to CCL21 of αDC1s and sDCs, directly (left) and 24 hours (right) after removal from the maturation cultures. The numbers in gray boxes represent the ratios between the numbers of sDCs and αDC1 that migrated to each individual concentration of CCL21. Similar data were obtained in 2 independent experiments. (D, left) In vivo migration of αDC1s and sDCs to the lymph nodes. Scintigraphic image of 111indium-labeled αDC1 and sDC at intradermal injection site and draining lymph nodes at 48 hours in a single representative patient. (D, right) In vivo migration of αDC1s or sDCs in 6 patients (each pair of dots represents each individual patient).
Figure 2
Figure 2
Selectively elevated CCR7 expression on the surface of PGE2-matured DCs occurs despite reduced levels of CCR7 mRNA and is suppressed by the exposure to CCL19. (A) Directly after harvesting from maturation cultures, αDC1s and sDCs were analyzed for the expression of surface CCR7 (left panel) and total (surface and intracellular; middle panel) CCR7 protein and for CCR7 mRNA expression (right panel). (B) Surface (top) and total (bottom) expression of CCR7 protein was analyzed in αDC1s, and sDCs matured in the absence or presence of exogenous CCL19 (100 ng/mL). Similar data were obtained in 3 independent experiments (3 different donors).
Figure 3
Figure 3
PGE2 suppresses the production of endogenous CCL19 in DCs induced to mature by TNFα, IFNα, or TLR ligands; poly-I:C or LPS: stability of the maturation-imprinted ability to produce CCL19. (A) CCL19 mRNA (left) and CCL19 protein expression (right) in αDC1s and sDCs. Representative data from one of 6 different donors. (B) CCL19 secretion by DCs exposed for 48 hours to IFNα or PGE2 (or both) in absence or presence of maturation factors, TNFα, poly-I:C (TLR3-ligand), or LPS (TLR4 ligand). Cumulative data (mean ± SEM) from 3 different donors. (C) CCL19 levels in αDC1s or sDCs during maturation or after maturation with or without CD40L. Cumulative data (mean ± SEM) from 3 different donors.
Figure 4
Figure 4
PGE2-matured DCs show suppressed ability to attract naive CD4+ T cells. Negative isolated naive CD4+ T cells were allowed to migrate toward 24-hour culture supernatants from αDC1s and sDCs in transwell migration chambers (3 hours), in the absence (▭) or presence (▬) of CCR7-blocking antibody. The migrated T cells were collected from the bottom chamber and counted. The dotted line represents the average spontaneous migration of T cells in the absence of DC supernatants. Cumulative data (mean ± SEM) from 4 different donors.

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