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Quantum dots for tracking dendritic cells and priming an immune response in vitro and in vivo.

Sen D, Deerinck TJ, Ellisman MH, Parker I, Cahalan MD - PLoS ONE (2008)

Bottom Line: LPS-induced maturation of DCs reduced the rate of endocytosis and the proportion of cells taking up QDs.Antigen-conjugated QDs induced CD69 expression, T cell proliferation, and IFN-gamma production in vivo with greater efficiency than equivalent amounts of free antigen.These results establish QDs as a versatile platform for immunoimaging of dendritic cells and as an efficient nanoparticle-based antigen delivery system for priming an immune response.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, University of California Irvine, Irvine, California, United States of America.

ABSTRACT
Dendritic cells (DCs) play a key role in initiating adaptive immune response by presenting antigen to T cells in lymphoid organs. Here, we investigate the potential of quantum dots (QDs) as fluorescent nanoparticles for in vitro and in vivo imaging of DCs, and as a particle-based antigen-delivery system to enhance DC-mediated immune responses. We used confocal, two-photon, and electron microscopies to visualize QD uptake into DCs and compared CD69 expression, T cell proliferation, and IFN-gamma production by DO11.10 and OT-II T cells in vivo in response to free antigen or antigen-conjugated to QDs. CD11c(+) DCs avidly and preferentially endocytosed QDs, initially into small vesicles near the plasma membrane by an actin-dependent mechanism. Within 10 min DCs contained vesicles of varying size, motion, and brightness distributed throughout the cytoplasm. At later times, endocytosed QDs were compartmentalized inside lysosomes. LPS-induced maturation of DCs reduced the rate of endocytosis and the proportion of cells taking up QDs. Following subcutaneous injection of QDs in an adjuvant depot, DCs that had endocytosed QDs were visualized up to 400 microm deep within draining lymph nodes. When antigen-conjugated QDs were used, T cells formed stable clusters in contact with DCs. Antigen-conjugated QDs induced CD69 expression, T cell proliferation, and IFN-gamma production in vivo with greater efficiency than equivalent amounts of free antigen. These results establish QDs as a versatile platform for immunoimaging of dendritic cells and as an efficient nanoparticle-based antigen delivery system for priming an immune response.

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Activation of DO11.10 T cells in vivo by DCs pulsed with QDova.Overlays of successive two-photon images of draining lymph nodes 8 hr after adoptive transfer of CMAC-labeled ova-specific T cells into mice immunized with QD (A) or QDova (B) included in 50 µl CFA. These images are single frames from Video S10 and S11, respectively. Scale bars = 20 µm. (C–H) Flow cytometry profiles showing activation of naïve T cells in draining lymph nodes of mice, into which ∼4×106 CFSE-labeled ova-specific T cells were adoptively transferred, and which were subsequently s.c. injected in the lower flank with varying doses of free ovalbumin (C–E, light gray), or ovalbumin conjugated to QDs (dark gray) included in 50 µl CFA. (I) Normalized activation index as a function of total amount of ovalbumin delivered either unconjugated (diamonds) or conjugated with QDs (squares). Data are representative of at least 3 independent experiments.
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pone-0003290-g007: Activation of DO11.10 T cells in vivo by DCs pulsed with QDova.Overlays of successive two-photon images of draining lymph nodes 8 hr after adoptive transfer of CMAC-labeled ova-specific T cells into mice immunized with QD (A) or QDova (B) included in 50 µl CFA. These images are single frames from Video S10 and S11, respectively. Scale bars = 20 µm. (C–H) Flow cytometry profiles showing activation of naïve T cells in draining lymph nodes of mice, into which ∼4×106 CFSE-labeled ova-specific T cells were adoptively transferred, and which were subsequently s.c. injected in the lower flank with varying doses of free ovalbumin (C–E, light gray), or ovalbumin conjugated to QDs (dark gray) included in 50 µl CFA. (I) Normalized activation index as a function of total amount of ovalbumin delivered either unconjugated (diamonds) or conjugated with QDs (squares). Data are representative of at least 3 independent experiments.

Mentions: We examined dynamic T cell / DC interactions in mice bearing adoptively transferred ova-specific T cells with and without immunization with QDova. In mice injected with QDs without ova, T cells migrated freely, interacting transiently with DCs and covering a broad territory, as indicated by time overlays of superimposed frames (Figure 7A; Video S10). In contrast, clusters of T cells were observed surrounding QDova-bearing DCs within draining lymph nodes from mice immunized with QDova (Figure 7B; Video S11). Time overlays showed that blue T cell tracks corresponded with red tracks of QDova-bearing DCs. These T cell-DC clusters were observed at depths up to 150 µm. These results show that antigen-conjugated QD-labeled DCs can functionally engage in prolonged interactions with antigen-specific T cells to initiate an immune response.


Quantum dots for tracking dendritic cells and priming an immune response in vitro and in vivo.

Sen D, Deerinck TJ, Ellisman MH, Parker I, Cahalan MD - PLoS ONE (2008)

Activation of DO11.10 T cells in vivo by DCs pulsed with QDova.Overlays of successive two-photon images of draining lymph nodes 8 hr after adoptive transfer of CMAC-labeled ova-specific T cells into mice immunized with QD (A) or QDova (B) included in 50 µl CFA. These images are single frames from Video S10 and S11, respectively. Scale bars = 20 µm. (C–H) Flow cytometry profiles showing activation of naïve T cells in draining lymph nodes of mice, into which ∼4×106 CFSE-labeled ova-specific T cells were adoptively transferred, and which were subsequently s.c. injected in the lower flank with varying doses of free ovalbumin (C–E, light gray), or ovalbumin conjugated to QDs (dark gray) included in 50 µl CFA. (I) Normalized activation index as a function of total amount of ovalbumin delivered either unconjugated (diamonds) or conjugated with QDs (squares). Data are representative of at least 3 independent experiments.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2538605&req=5

pone-0003290-g007: Activation of DO11.10 T cells in vivo by DCs pulsed with QDova.Overlays of successive two-photon images of draining lymph nodes 8 hr after adoptive transfer of CMAC-labeled ova-specific T cells into mice immunized with QD (A) or QDova (B) included in 50 µl CFA. These images are single frames from Video S10 and S11, respectively. Scale bars = 20 µm. (C–H) Flow cytometry profiles showing activation of naïve T cells in draining lymph nodes of mice, into which ∼4×106 CFSE-labeled ova-specific T cells were adoptively transferred, and which were subsequently s.c. injected in the lower flank with varying doses of free ovalbumin (C–E, light gray), or ovalbumin conjugated to QDs (dark gray) included in 50 µl CFA. (I) Normalized activation index as a function of total amount of ovalbumin delivered either unconjugated (diamonds) or conjugated with QDs (squares). Data are representative of at least 3 independent experiments.
Mentions: We examined dynamic T cell / DC interactions in mice bearing adoptively transferred ova-specific T cells with and without immunization with QDova. In mice injected with QDs without ova, T cells migrated freely, interacting transiently with DCs and covering a broad territory, as indicated by time overlays of superimposed frames (Figure 7A; Video S10). In contrast, clusters of T cells were observed surrounding QDova-bearing DCs within draining lymph nodes from mice immunized with QDova (Figure 7B; Video S11). Time overlays showed that blue T cell tracks corresponded with red tracks of QDova-bearing DCs. These T cell-DC clusters were observed at depths up to 150 µm. These results show that antigen-conjugated QD-labeled DCs can functionally engage in prolonged interactions with antigen-specific T cells to initiate an immune response.

Bottom Line: LPS-induced maturation of DCs reduced the rate of endocytosis and the proportion of cells taking up QDs.Antigen-conjugated QDs induced CD69 expression, T cell proliferation, and IFN-gamma production in vivo with greater efficiency than equivalent amounts of free antigen.These results establish QDs as a versatile platform for immunoimaging of dendritic cells and as an efficient nanoparticle-based antigen delivery system for priming an immune response.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, University of California Irvine, Irvine, California, United States of America.

ABSTRACT
Dendritic cells (DCs) play a key role in initiating adaptive immune response by presenting antigen to T cells in lymphoid organs. Here, we investigate the potential of quantum dots (QDs) as fluorescent nanoparticles for in vitro and in vivo imaging of DCs, and as a particle-based antigen-delivery system to enhance DC-mediated immune responses. We used confocal, two-photon, and electron microscopies to visualize QD uptake into DCs and compared CD69 expression, T cell proliferation, and IFN-gamma production by DO11.10 and OT-II T cells in vivo in response to free antigen or antigen-conjugated to QDs. CD11c(+) DCs avidly and preferentially endocytosed QDs, initially into small vesicles near the plasma membrane by an actin-dependent mechanism. Within 10 min DCs contained vesicles of varying size, motion, and brightness distributed throughout the cytoplasm. At later times, endocytosed QDs were compartmentalized inside lysosomes. LPS-induced maturation of DCs reduced the rate of endocytosis and the proportion of cells taking up QDs. Following subcutaneous injection of QDs in an adjuvant depot, DCs that had endocytosed QDs were visualized up to 400 microm deep within draining lymph nodes. When antigen-conjugated QDs were used, T cells formed stable clusters in contact with DCs. Antigen-conjugated QDs induced CD69 expression, T cell proliferation, and IFN-gamma production in vivo with greater efficiency than equivalent amounts of free antigen. These results establish QDs as a versatile platform for immunoimaging of dendritic cells and as an efficient nanoparticle-based antigen delivery system for priming an immune response.

Show MeSH
Related in: MedlinePlus