The recruitment of selected dendritic cell (DC) subtypes conditions the class from the immune response. pDCs either in lymph nodes or at peripheral sites of swelling. = 3). (B) Enriched circulating bloodstream DC subsets had been rested for 2 h at 37C and researched in transwell (5-m pore size) migration assay. Types of Compact disc11c FACS? information of gated HLA-DR+ lineage marker? migrating cells (representative of 10). (C) Reactions of blood Compact disc11c? pDCs and Compact disc11c+ myeloid DCs to different chemokines. Each chemokine was examined over an array of concentrations (1C1,000 ng/ml) in support of the perfect response is demonstrated ( 5). Open up in another window Shape 2. Powerful activity of the constitutive chemokine SDF-1/CXCL12 and buy H 89 dihydrochloride high CXCR4, CXCR3, and l-selectin manifestation on pDCs. CXCR4 (A), CXCR3 (B), and l-selectin (C) cell surface area manifestation on DC human population (representative of 3). For pDC CXCR4, manifestation was also examined after 2 h of preincubation at 37C (A, dashed range). mRNA manifestation of CXCR4 and CXCR3 was also dependant on quantitative PCR (D) on a single populations (outcomes indicated as pg/50 ng total RNA had been normalized using 18S RNA, suggest of = 3). Finally, ideal response to CXCR4 and CXCR3 ligands can be shown for every DC human population (E, mean of 5). The constitutive manifestation of SDF-1/CXCL12 and of l-selectin ligands on HEVs may be in charge of the trafficking of pDCs through the blood towards the lymph nodes in homeostatic circumstances. This hypothesis can be supported from the noticed pDC insufficiency in lymph nodes from l-selectinCdeficient mice (19). Therefore, pDC migration could possibly be similar to that of naive T cells through the blood towards the lymph nodes through l-selectin ligands as well as the CCR7 ligand 6Ckine/CCL21 expressed by HEV (11). However, CCR7 ligands are unlikely to be involved for resting pDC recruitment as they lack CCR7 expression. The model proposed here would imply that unlike myeloid DCs that capture pathogens in peripheral tissues and consequently migrate in lymph nodes, pDCs are constitutively recruited in lymph nodes, where they might be exposed to infectious agents. The Inducible CXCR3 Ligands Regulate pDC Responsiveness to the Constitutive Chemokine SDF-1/CXCL12. Among chemokine receptors in pDCs, CXCR3 was expressed at the highest level (Fig. 1 A) and was absent on all other DC populations as detected by immunofluorescence at buy H 89 dihydrochloride cell surface, or by quantitative RT-PCR (Fig. 2, B and D). However, only high concentrations (1C5 g/ml) of the CXCR3 ligands IP-10/CXCL10, Mig/CXCL9, and I-TAC/ CXCL11 induced significant but modest migration buy H 89 dihydrochloride of pDCs (Figs. 1 C and 3, A and C). In the presence of a suboptimal dose of Rabbit polyclonal to SORL1 SDF-1/CXCL12 (10 ng/ml), IP-10/CXCL10 at a lower concentration (100C1,000 ng/ml) induced brisk migration of pDCs (Fig. 3 A). buy H 89 dihydrochloride In combination with SDF-1/CXCL12, all CXCR3 ligands dramatically decreased the threshold of SDF-1/CXCL12 sensitivity by 20C50-fold (Fig. 3, B and C). These observations were confirmed on purified FACS?-sorted pDCs (Fig. 3 D) and extend a very recent study (17). Open in a separate window Figure 3. CXCR3 ligands selectively induce pDCs to respond to low SDF-1/CXCL12 concentration. (A) Dose response to IP-10/CXCL10 of pDCs in the presence or absence of a low dose of SDF-1/CXCL12 (10 ng/ml). (B) Dose response to SDF-1/CXCL12 of pDCs in the presence or absence of 1 g/ml IP-10/CXCL10. (C) Response of pDCs to all CXCR3 ligands (1 g/ml), tested individually, in the presence or absence of a low dose of SDF-1/CXCL12 (10 ng/ml). (D) Dose response to SDF-1/CXCL12 of FACS?-sorted CD11c? pDCs and CD11c+ myeloid DCs in the presence or absence of 1 g/ml IP-10/CXCL10. (E) Effects of 1 g/ml IP-10/CXCL10 on the dose response to SDF-1/CXCL12 of various DC and T cell populations. Results are expressed as folds of IP-10Cinduced migration over that of SDF-1/CXCL12 alone (ratio migration index in SDF-1/CXCL12 + IP-10/CXCL10/migration index in SDF-1/CXCL12 alone). Results from 3. With FACS?-sorted CD11c+ circulating blood myeloid DCs, no synergistic activity was observed (Fig. 3 D). Also, neither the other DC populations tested (monocyte-derived DCs and CD34-derived DC subsets) nor naive T cells displayed synergistic response (Fig. 3 E), in agreement with the lack of CXCR3 expression on these cells..