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The adapter protein ADAP is required for selected dendritic cell functions.

Togni M, Engelmann S, Reinhold D, Schraven B, Reinhold A - Cell Commun. Signal (2012)

Bottom Line: In contrast, ADAP-deficient BMDCs showed defects in CD11c-mediated cellular responses, with significantly diminished production of IL-6, TNF-∝ and IL-10.Actin polymerization was enhanced after CD11c integrin stimulation.In summary, we propose that the adapter molecule ADAP is critical for selected CD11c integrin-mediated functions of dendritic cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute for Molecular and Clinical Immunology, Otto von Guericke University Magdeburg, Leipziger Strasse 44, 39120 Magdeburg, Germany. annegret.reinhold@med.ovgu.de.

ABSTRACT

Background: The cytosolic adaptor protein ADAP (adhesion and degranulation promoting adapter protein) is expressed by T cells, natural killer cells, myeloid cells and platelets. ADAP is involved in T-cell-receptor-mediated inside-out signaling, which leads to integrin activation, adhesion and reorganization of the actin cytoskeleton. However, little is known about the role of ADAP in myeloid cells. In the present study, we analyzed the function of ADAP in bone-marrow-derived dendritic cells (BMDCs) from ADAP-deficient mice.

Results: ADAP-deficient BMDCs showed almost normal levels of antigen uptake, adhesion, maturation, migration from the periphery to the draining lymph nodes, antigen-specific T-cell activation, and production of the proinflammatory cytokines IL-6 and TNF-∝. Furthermore, we provide evidence that the activation of signaling pathways after lipopolysaccharide (LPS) stimulation are not affected by the loss of ADAP. In contrast, ADAP-deficient BMDCs showed defects in CD11c-mediated cellular responses, with significantly diminished production of IL-6, TNF-∝ and IL-10. Actin polymerization was enhanced after CD11c integrin stimulation.

Conclusions: In summary, we propose that the adapter molecule ADAP is critical for selected CD11c integrin-mediated functions of dendritic cells.

No MeSH data available.


Related in: MedlinePlus

Cytokine production and actin polymerization after CD11c stimulation. (A) Production of cytokines TNF- ∝, IL-6 and IL-10 by BMDCs of wild-type animals (WT) and ADAP-deficient animals (ADAP) was measured in the supernatants 24 h after stimulation with anti-CD11c (mean + SEM, n = 3, *P < 0.05, ns = non significant). (B) BMDCs of WT and ADAP were incubated with anti-CD11c. After incubation, cells were permeabilized and stained with TRITC-phalloidin. The cellular F-actin content was analyzed by FACS. Results are expressed as the percentage increase in mean fluorescence intensity (mean ± SEM, n = 6; P = 0.0031 for the two curves, as assessed by two-way ANOVA). (C) BMDCs of WT and ADAP were stimulated with anti-CD11c for 0, 5, 10, 20, and 30 min. Lysates were separated by SDS-PAGE, and immunoblotted with the indicated antibodies. One representative of three independent experiments is shown.
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Figure 6: Cytokine production and actin polymerization after CD11c stimulation. (A) Production of cytokines TNF- ∝, IL-6 and IL-10 by BMDCs of wild-type animals (WT) and ADAP-deficient animals (ADAP) was measured in the supernatants 24 h after stimulation with anti-CD11c (mean + SEM, n = 3, *P < 0.05, ns = non significant). (B) BMDCs of WT and ADAP were incubated with anti-CD11c. After incubation, cells were permeabilized and stained with TRITC-phalloidin. The cellular F-actin content was analyzed by FACS. Results are expressed as the percentage increase in mean fluorescence intensity (mean ± SEM, n = 6; P = 0.0031 for the two curves, as assessed by two-way ANOVA). (C) BMDCs of WT and ADAP were stimulated with anti-CD11c for 0, 5, 10, 20, and 30 min. Lysates were separated by SDS-PAGE, and immunoblotted with the indicated antibodies. One representative of three independent experiments is shown.

Mentions: In addition to its role in inside-out signaling, ADAP is implicated in integrin-mediated outside-in signaling in T cells [20]. To assess the role of ADAP in outside-in signaling in BMDCs, we first investigated cytokine production following CD11c stimulation. After stimulation with anti-CD11c and in the absence of additional stimuli, the production of IL-6, TNF-∝ and IL-10 was clearly reduced in ADAP-deficient BMDCs compared with that in wild-type BMDCs (2,665 ± 292 and 1,209 ± 322 pg/ml TNF-∝, 5,257 ± 1,627 and 2,428 ± 1,167 pg/ml IL-6, and 25.3 ± 2.2 and 16.3 ± 0.9 pg/ml IL-10, for wild-type and ADAP-deficient BMDCs, respectively; Figure 6A).


The adapter protein ADAP is required for selected dendritic cell functions.

Togni M, Engelmann S, Reinhold D, Schraven B, Reinhold A - Cell Commun. Signal (2012)

Cytokine production and actin polymerization after CD11c stimulation. (A) Production of cytokines TNF- ∝, IL-6 and IL-10 by BMDCs of wild-type animals (WT) and ADAP-deficient animals (ADAP) was measured in the supernatants 24 h after stimulation with anti-CD11c (mean + SEM, n = 3, *P < 0.05, ns = non significant). (B) BMDCs of WT and ADAP were incubated with anti-CD11c. After incubation, cells were permeabilized and stained with TRITC-phalloidin. The cellular F-actin content was analyzed by FACS. Results are expressed as the percentage increase in mean fluorescence intensity (mean ± SEM, n = 6; P = 0.0031 for the two curves, as assessed by two-way ANOVA). (C) BMDCs of WT and ADAP were stimulated with anti-CD11c for 0, 5, 10, 20, and 30 min. Lysates were separated by SDS-PAGE, and immunoblotted with the indicated antibodies. One representative of three independent experiments is shown.
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Figure 6: Cytokine production and actin polymerization after CD11c stimulation. (A) Production of cytokines TNF- ∝, IL-6 and IL-10 by BMDCs of wild-type animals (WT) and ADAP-deficient animals (ADAP) was measured in the supernatants 24 h after stimulation with anti-CD11c (mean + SEM, n = 3, *P < 0.05, ns = non significant). (B) BMDCs of WT and ADAP were incubated with anti-CD11c. After incubation, cells were permeabilized and stained with TRITC-phalloidin. The cellular F-actin content was analyzed by FACS. Results are expressed as the percentage increase in mean fluorescence intensity (mean ± SEM, n = 6; P = 0.0031 for the two curves, as assessed by two-way ANOVA). (C) BMDCs of WT and ADAP were stimulated with anti-CD11c for 0, 5, 10, 20, and 30 min. Lysates were separated by SDS-PAGE, and immunoblotted with the indicated antibodies. One representative of three independent experiments is shown.
Mentions: In addition to its role in inside-out signaling, ADAP is implicated in integrin-mediated outside-in signaling in T cells [20]. To assess the role of ADAP in outside-in signaling in BMDCs, we first investigated cytokine production following CD11c stimulation. After stimulation with anti-CD11c and in the absence of additional stimuli, the production of IL-6, TNF-∝ and IL-10 was clearly reduced in ADAP-deficient BMDCs compared with that in wild-type BMDCs (2,665 ± 292 and 1,209 ± 322 pg/ml TNF-∝, 5,257 ± 1,627 and 2,428 ± 1,167 pg/ml IL-6, and 25.3 ± 2.2 and 16.3 ± 0.9 pg/ml IL-10, for wild-type and ADAP-deficient BMDCs, respectively; Figure 6A).

Bottom Line: In contrast, ADAP-deficient BMDCs showed defects in CD11c-mediated cellular responses, with significantly diminished production of IL-6, TNF-∝ and IL-10.Actin polymerization was enhanced after CD11c integrin stimulation.In summary, we propose that the adapter molecule ADAP is critical for selected CD11c integrin-mediated functions of dendritic cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institute for Molecular and Clinical Immunology, Otto von Guericke University Magdeburg, Leipziger Strasse 44, 39120 Magdeburg, Germany. annegret.reinhold@med.ovgu.de.

ABSTRACT

Background: The cytosolic adaptor protein ADAP (adhesion and degranulation promoting adapter protein) is expressed by T cells, natural killer cells, myeloid cells and platelets. ADAP is involved in T-cell-receptor-mediated inside-out signaling, which leads to integrin activation, adhesion and reorganization of the actin cytoskeleton. However, little is known about the role of ADAP in myeloid cells. In the present study, we analyzed the function of ADAP in bone-marrow-derived dendritic cells (BMDCs) from ADAP-deficient mice.

Results: ADAP-deficient BMDCs showed almost normal levels of antigen uptake, adhesion, maturation, migration from the periphery to the draining lymph nodes, antigen-specific T-cell activation, and production of the proinflammatory cytokines IL-6 and TNF-∝. Furthermore, we provide evidence that the activation of signaling pathways after lipopolysaccharide (LPS) stimulation are not affected by the loss of ADAP. In contrast, ADAP-deficient BMDCs showed defects in CD11c-mediated cellular responses, with significantly diminished production of IL-6, TNF-∝ and IL-10. Actin polymerization was enhanced after CD11c integrin stimulation.

Conclusions: In summary, we propose that the adapter molecule ADAP is critical for selected CD11c integrin-mediated functions of dendritic cells.

No MeSH data available.


Related in: MedlinePlus