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TOLEROGENIC BONE MARROW-DERIVED DENDRITIC CELLS MODULATE ALLERGIC REACTIVITY - PowerPoint PPT Presentation


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BACKGROUND

Asthma (allergic lung disease; ALD) is presently described as a syndrome characterized by: (a) intermittent and reversible airway obstruction; (b) airway hyperresponsiveness (AHR); and (c) airway inflammation. Despite significant pharmacological advances in asthma therapy, the past two decades have seen an alarming increase in the prevalence of asthma world wide. In the United States alone, asthma affects approximately 14 -15 million children and adults.

Dendritic cells (DC) are a family of professional antigen (Ag) presenting cells (APC), considered by many to be the central APC for induction of primary immune responses. Their abilities to process and present various types of antigens are unmatched in this context. The decision of whether or not encounter with an antigen will lead to an immune response is controlled in many respects at the level of APC and is subject to tight regulation. Tolerogenic DC have been implicated as critical in defining immunologic ‘self’ and preventing the induction of both autoimmunity and chronic inflammation against environmental proteins. Recent studies show that different subsets of DC play important roles in central and peripheral tolerance.

It has been reported that treatment of DC with IL-10 inhibits their terminal differentiation, can reduce expression of co-stimulatory molecules, and can lead to suppression of antigen specific responses. We wished to determine whether such “tolerogenic” DC could reverse pre-existing asthma using a standard model of ALD (Schneider et al, 2001).

CD40

CD54

CD86

CD80

MHC-Il

DCGM-CSF

600

400

DCIL-10

200

FIG. 3. DCIL-10 THERAPY REDUCES Th2-CYTOKINES IN THE AIRWAYS.

BAL fluids from ALD mice treated with saline, DCIL-10 , or OVA-pulsed DCIL-10 were assessed on treatment day 28. The levels of IL-4, IL-5, and IL-13 were significantly affected by the DC treatments, with OVA-presenting DC providing additional protective effects over DCIL-10 not exposed to OVA. (These results comprise one representative experiment of eight)

0

sal DC DC/OVA med

sal DC DC/OVA med

sal DC DC/OVA med

sal DC DC/OVA med

sal DC DC/OVA med

sal DC DC/OVA med

sal DC DC/OVA med

sal DC DC/OVA med

DCTNF

IL-1b

IL-6

IL-10

IL-12

IL-12

IFN-g

IL-5

IL-4

DCIL-10

DCTNF

DCGMCSF

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No FITC-dex

FITC-dex:

100 µg/ml 50 µg/ml

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DCIL-10

(at 370C)

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ALD

Norm.

Number of cells

IL-9

IL-13

DCIL-10

(at 40C)

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0 0.1 1 10 100

MIP-1a (ng/ml)

IL-10

TGF-b

300

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TOLEROGENIC BONE MARROW-DERIVED DENDRITIC CELLS MODULATE ALLERGIC REACTIVITY

OF LUNG CELLS FROM MICE WITH SEVERE ALLERGIC LUNG DISEASE.

Aarti Nayyar, Xiaobei Zhang and John R. Gordon.

Immunology Research Group (Dept Vet Microbiology), University of Saskatchewan, Saskatoon.

HYPOTHESIS

TOLEROGENIC BONE MARROW-DERIVED DENDRITIC CELLS CAN BE USED AS AN EFFECTIVE THERAPY FOR SEVERE ALLERGIC LUNG DISEASE

OBJECTIVES

To generate (through culture in IL-10-containing media) and characterize tolerogenic populations of mouse bone marrow-derived dendritic cells (DC)

2. To assess the impact of these DCs, as well as fully mature BMDC, on tolerance induction in a mouse model of allergic lung disease/asthma.

ALLERGIC LUNG DISEASE (ALD) MODEL

BALB/c mice were sensitized with OVA/alum (2 µg/mg, i.p.) on dy 0 & 14, exposed to 1% OVA aerosols on days 28, 30, & 32, then treated with DCIL-10, DCGM-CSF or DCTNF on day 42 (Schneider et al, 2001).

RESULTS

FIG 2. DCIL-10-TREATMENTS ABROGATE AHR IN MICE WITH SEVERE ALD.

FIG. 1. FUNCTIONAL CHARACTERIZATION OF DCIL-10IN VITRO

BALB/c mice with severe ALD (≈60% airway eosinophils on airway allergen challenge) were given 1x106 DCIL-10, DCGM-CSF, or DCTNF transtracheally. Over the next 3 weeks they were assess by head-out body plethysmo-graphy for AHR to methacholine. This experiment is representative of ≈8 others in which we have found that, beginning at 15 -17 days post-transplant, the AHR of DCIL-10-treated mice, but not those treated with either DCGM-CSF or DCTNF, disappears.

Bone marrow cells from BALB/c mice were cultured in high (20 ng/ml) , then low (7.5 ng/ml) dose GM-CSF + IL-10 (50 ng/ml). After 15 dy, the cells were analyzed (A) by FACS for multiple markers (left panel), in vitro cytokine release (right panel), (B) phagocytic capacity (left) and chemokine receptor expression (right). These DCIL-10 were compared in the FACS analysis with immature DC GM-CSF and immunostimulatory, mature DCTNF. DCIL-10 expressed slightly lower levels of cell surface CD40, CD54 and MHC-II. They also released significant amounts of IL-10 and TGFb (A).They possessed functional phagocytosis and strong chemotaxis to the inflammatory chemokine MIP-1a (B).

DAY 21

DCIL-10

DCIL-10

DCIL-10

CYTOKINE SECRETION

A.

TGF-b

FACS analysis also confirmed that the DCIL-10 populations did not express neutrophil, macrophage, B or T cell markers, and that they did express low levels of CD11c, and DEC205, as well as expected levels of CD11b, MHC-I & CD45RB

DCGM-CSF and DCTNF populations did not express appreciable levels of IL-10 or TGFb. DCTNF expressed high levels of TNF and IL-12, while DCGM-CSF expressed substantial amounts of IL-6, but not the other cytokines.

B.

FITC-dextran PHAGOCYTOSIS

CHEMOKINE RECEPTORS

REFERENCES

-Jonuleit H, Schmitt KE, Steinbrink K and Enk AH(2001) Dendritic cells as a tool to induce anergic and regulatory T cells. Trends Immunol 22: 394-400.

-Steinbrink K, Matthias W, Jonuleit H, Jurgen K and Enk AH (1997) Induction of tolerance by IL-10 treated dendritic cells. J Immunol 159: 4772-80.

-Schneider AM, Zhang X, Li F, and Gordon JR. (2001) Differential induction of allergen-specific IgA, AHR, or allergic airway disease following sensitization with limiting doses of ovalbumin-alum. Cell Immunol 212:101-109

CONCLUSION: DELIVERY OF DCIL-10, BUT NOT DCGM-CSF OR DCTNF, INTO THE AIRWAYS OF MICE WITH ALD ABROGATES AHR & SUBSTANTIALLY AMELIORATES Th2 REACTIVITY

In MIP-3a chemotaxis (i.e., CCR7) assays, DCGM-CSF and DCIL-10 were shown to express low, but significant, levels of CCR7, while the DCTNF responded very strongly via this receptor, as expected (data not shown).

Both DCIL-10 and DCGM-CSF avidly phagocytosed FITC-dextran, while the DCTNF did not.

SUPPORTED BY GRANTS FROM THE CANADIAN INSTITUTES FOR HEALTH RESEARCH AND THE SASKATCHEWAN LUNG ASSOCIATION


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