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Maturation stages of mouse dendritic cells in growth factor-dependent long-term cultures.

Winzler C, Rovere P, Rescigno M, Granucci F, Penna G, Adorini L, Zimmermann VS, Davoust J, Ricciardi-Castagnoli P - J. Exp. Med. (1997)

Bottom Line: Antigen uptake and presentation of native protein antigen was reduced.In contrast, presentation of immunogenic peptides and allostimulatory activity became very efficient and secretion of IL-12 p75 was detectable after antigen presentation.This functional DC maturation ended by apoptotic cell death, and no reversion to the immature phenotype was observed.

View Article: PubMed Central - PubMed

Affiliation: CNR Centre of Cellular and Molecular Pharmacology, Milan, Italy.

ABSTRACT
The signals controlling the checkpoints of dendritic cells (DC) maturation and the correlation between phenotypical and functional maturational stages were investigated in a defined model system of growth factor-dependent immature mouse DC. Three sequential stages of DC maturation (immature, mature, and apoptotic) were defined and characterized. Immature DC (stage 1) had low expression of costimulatory molecules, highly organized cytoskeleton, focal adhesion plaques, and slow motility; accordingly, they were very efficient in antigen uptake and processing of soluble proteins. Further, at this stage most of major histocompatibility complex class II molecules were within cytoplasmic compartments consistent with a poor allostimulatory capacity. Bacteria or cytokines were very efficient in inducing progression from stage 1 towards stage 2 (mature). Morphological changes were observed by confocal analysis including depolymerization of F-actin and loss of vinculin containing adhesive structures which correlates with acquisition of high motility. Antigen uptake and presentation of native protein antigen was reduced. In contrast, presentation of immunogenic peptides and allostimulatory activity became very efficient and secretion of IL-12 p75 was detectable after antigen presentation. This functional DC maturation ended by apoptotic cell death, and no reversion to the immature phenotype was observed.

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Antigen uptake (FITC-OVA and FITC-DX) by D1 bulk  population in the presence or absence of TNFα was analyzed by doublecolor FACS® analysis. The D1 cells that better internalize soluble antigens  at 37°C expressed comparatively low levels of MHC class II molecules,  whereas class IIbright were less efficient in both FITC-OVA and FITC-DX  uptake. Incubation of the cells with TNFα reduced the protein uptake of  the immature D1 subpopulation and had no effects on the mature D1  subpopulation.
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Figure 7: Antigen uptake (FITC-OVA and FITC-DX) by D1 bulk population in the presence or absence of TNFα was analyzed by doublecolor FACS® analysis. The D1 cells that better internalize soluble antigens at 37°C expressed comparatively low levels of MHC class II molecules, whereas class IIbright were less efficient in both FITC-OVA and FITC-DX uptake. Incubation of the cells with TNFα reduced the protein uptake of the immature D1 subpopulation and had no effects on the mature D1 subpopulation.

Mentions: Unstimulated D1 cultures were very efficient in fluid phase and receptor-mediated antigen uptake, a property largely abolished when the experiments were performed at 0°C. Using double-color FACS® analysis, the uptake at 37°C of FITC-OVA and FITC-DX was analyzed in the MHC class IIintand in the MHC class IIbright D1 cell subpopulations. The highest uptake of FITC-OVA and FITCDX was associated with MHC class II int cells (Fig. 7). In contrast, the MHC class II bright D1 subpopulation was less efficient in FITC-OVA and FITC-DX uptake, thus indicating that macropinocytosis and receptor-mediated endocytosis are highly reduced in mature DC.


Maturation stages of mouse dendritic cells in growth factor-dependent long-term cultures.

Winzler C, Rovere P, Rescigno M, Granucci F, Penna G, Adorini L, Zimmermann VS, Davoust J, Ricciardi-Castagnoli P - J. Exp. Med. (1997)

Antigen uptake (FITC-OVA and FITC-DX) by D1 bulk  population in the presence or absence of TNFα was analyzed by doublecolor FACS® analysis. The D1 cells that better internalize soluble antigens  at 37°C expressed comparatively low levels of MHC class II molecules,  whereas class IIbright were less efficient in both FITC-OVA and FITC-DX  uptake. Incubation of the cells with TNFα reduced the protein uptake of  the immature D1 subpopulation and had no effects on the mature D1  subpopulation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196118&req=5

Figure 7: Antigen uptake (FITC-OVA and FITC-DX) by D1 bulk population in the presence or absence of TNFα was analyzed by doublecolor FACS® analysis. The D1 cells that better internalize soluble antigens at 37°C expressed comparatively low levels of MHC class II molecules, whereas class IIbright were less efficient in both FITC-OVA and FITC-DX uptake. Incubation of the cells with TNFα reduced the protein uptake of the immature D1 subpopulation and had no effects on the mature D1 subpopulation.
Mentions: Unstimulated D1 cultures were very efficient in fluid phase and receptor-mediated antigen uptake, a property largely abolished when the experiments were performed at 0°C. Using double-color FACS® analysis, the uptake at 37°C of FITC-OVA and FITC-DX was analyzed in the MHC class IIintand in the MHC class IIbright D1 cell subpopulations. The highest uptake of FITC-OVA and FITCDX was associated with MHC class II int cells (Fig. 7). In contrast, the MHC class II bright D1 subpopulation was less efficient in FITC-OVA and FITC-DX uptake, thus indicating that macropinocytosis and receptor-mediated endocytosis are highly reduced in mature DC.

Bottom Line: Antigen uptake and presentation of native protein antigen was reduced.In contrast, presentation of immunogenic peptides and allostimulatory activity became very efficient and secretion of IL-12 p75 was detectable after antigen presentation.This functional DC maturation ended by apoptotic cell death, and no reversion to the immature phenotype was observed.

View Article: PubMed Central - PubMed

Affiliation: CNR Centre of Cellular and Molecular Pharmacology, Milan, Italy.

ABSTRACT
The signals controlling the checkpoints of dendritic cells (DC) maturation and the correlation between phenotypical and functional maturational stages were investigated in a defined model system of growth factor-dependent immature mouse DC. Three sequential stages of DC maturation (immature, mature, and apoptotic) were defined and characterized. Immature DC (stage 1) had low expression of costimulatory molecules, highly organized cytoskeleton, focal adhesion plaques, and slow motility; accordingly, they were very efficient in antigen uptake and processing of soluble proteins. Further, at this stage most of major histocompatibility complex class II molecules were within cytoplasmic compartments consistent with a poor allostimulatory capacity. Bacteria or cytokines were very efficient in inducing progression from stage 1 towards stage 2 (mature). Morphological changes were observed by confocal analysis including depolymerization of F-actin and loss of vinculin containing adhesive structures which correlates with acquisition of high motility. Antigen uptake and presentation of native protein antigen was reduced. In contrast, presentation of immunogenic peptides and allostimulatory activity became very efficient and secretion of IL-12 p75 was detectable after antigen presentation. This functional DC maturation ended by apoptotic cell death, and no reversion to the immature phenotype was observed.

Show MeSH
Related in: MedlinePlus