FIGURE 1

CCR9 and 47 induction requires signaling via RA receptors. (a) Experimental setup to investigate the nature of CCR9-inducing signals. (II–IV) Purified CD11c⁺ splenic DCs (2.5 10⁵) were pulsed with pOVA(1 nM) and incubated with 5 10⁵ CFSE-labeled OT-I cells in the lower chamber of a 0.4 m transwell. (I, III, and IV) Purified CD11c⁺ MLN DCs (10⁵) were pulsed with 1 nM pOVA and added (III) alone or (I and IV) together with 2 10⁵ CFSE-labeled OT-I cells to the upper insert. After 4 days of culture, OT-I cells cultured with (I) MLN DCs or (II–IV) splenic DCs were analyzed for CCR9 expression by flow cytometry. Results are from one representative experiment of three performed. (b and c) RAR signaling is required for induction of CCR9 and 47 expression on OT-I cells. Antigen-loaded (20 pM) MLN DCs (10⁵) were co-cultured with OT-I cells (2 10⁵) in the presence or absence of pan-RAR inhibitor AGN194310. Following 4 days of culture, CCR9 and 47 expression was determined on the OT-I cells by flow cytometry. (c) Bars represent means.e.m. from three to four experiments. *P<0.05 compared to no added inhibitor. (d) RAR signaling is required for splenic DC-mediated induction of 47 expression on OT-I cells. Antigen-loaded (20 pM) splenic DCs (10⁵) were co-cultured with OT-I cells (2 10⁵) in the presence or absence of pan-RAR inhibitor AGN194310. Following 4 days of culture, 47 expression was determined on the OT-I cells by flow cytometry. One representative experiment of four performed. CCR, CC chemokine receptor; CXCR, CXC chemokine receptor; DC, dendritic cell; MLN, mesenteric lymph node; pOVA, ovalbumin peptide; RA, retinoic acid; RAR, retinoic acid receptor; SPL, splenic.