Mast cell anaphylatoxin receptor expression can enhance IgE-dependent skin inflammation in mice

Abstract

Background: Mast cells express receptors for complement anaphylatoxins C3a and C5a (ie, C3a receptor [C3aR] and C5a receptor [C5aR]), and C3a and C5a are generated during various IgE-dependent immediate hypersensitivity reactions in vivo. However, it is not clear to what extent mast cell expression of C3aR or C5aR influences C3a- or C5a-induced cutaneous responses or IgE-dependent mast cell activation and passive cutaneous anaphylaxis (PCA) in vivo.

Objective: We sought to assess whether mouse skin mast cell expression of C3aR or C5aR influences (1) the cells' responsiveness to intradermal injections of C3a or C5a or (2) the extent of IgE-dependent mast cell degranulation and PCA in vivo.

Methods: We measured the magnitude of cutaneous responses to intradermal injections of C3a or C5a and the extent of IgE-dependent mast cell degranulation and PCA responses in mice containing mast cells that did or did not express C3aR or C5aR.

Results: The majority of the skin swelling induced by means of intradermal injection of C3a or C5a required that mast cells at the site expressed C3aR or C5aR, respectively, and the extent of IgE-dependent degranulation of skin mast cells and IgE-dependent PCA was significantly reduced when mast cells lacked either C3aR or C5aR. IgE-dependent PCA responses associated with local increases in C3a levels occurred in antibody-deficient mice but not in mice deficient in FcɛRIγ.

Conclusion: Expression of C3aR and C5aR by skin mast cells contributes importantly to the ability of C3a and C5a to induce skin swelling and can enhance mast cell degranulation and inflammation during IgE-dependent PCA in vivo.

FIG 1. Mast cells enhance C3a- or C5a- induced tissue swelling in a C3aR- or C5aR-dependent manner, respectively

Changes (Δ) in ear thickness in mice after injection (i.d.) of 100 ng C3a (A) or C5a (B). **P < .01, ***P < .001 vs. Kit^W-sh/W-sh mice, and +++ P < .001 vs. C3aR^-/- BMCMCs→Kit^W-sh/W-sh mice (A) or C5aR^-/- BMCMCs→Kit^W-sh/W-sh mice (B).

FIG 2. Ear swelling and C3a production during IgE-dependent PCA reactions are not dependent on the presence of native antibodies

(A) Changes (Δ) in ear thickness after injection (i.v.) of 200 mg DNP-HSA into WT (C57BL/6) and Ab-deficient (B6.129S2-Igh-6^tm1Cgn/J) mice 24 h after i.d. injection of vehicle (left ears) or 20 ng anti-DNP IgE (right ears). *** P < .001 vs. corresponding vehicle control; +++ P < .001 vs. IgE/DNP-HSA treated WT group. (B) Percentage of mast cells (MCs) in vehicle- (V) or IgE-injected pinnae exhibiting extensive (ext), moderate (mod) or no (none) degranulation 6 h after i.v. DNP-HSA challenge. *** P < .001 vs. corresponding vehicle group. P = 0.16 for IgE/DNP-HSA treated WT vs. IgE/DNP-HSA treated Ab-deficient group. (C) C3a in ear lysates 2 h after injection (i.v.) of 200 μg DNP-HSA into WT or Ab-deficient mice 24 h after i.d. injection of vehicle (V; left ears) or 20 ng anti-DNP IgE (right ears). *** P < .001 vs. vehicle; +++ P < .001 vs. IgE/DNP-HSA treated WT group.

FIG 3. IgE-dependent PCA reactions require the Fc receptor gamma chain

(A) Changes (Δ) in ear thickness after injection (i.v.) of 200 μg DNP-HSA into mice 24 h after i.d. injection of vehicle (left ears) or 20 ng anti-DNP IgE (right ears). *** P < .001 vs. corresponding vehicle injected group; +++ P < .001 vs. IgE/DNP-HSA treated Fcer1g^-/- group. (B) Numbers of mast cells per mm of ear cartilage were counted in toluidine blue-stained ear pinnae sections prepared from WT or Fcer1g^-/- mice (data for vehicle- (V) or IgE-injected ears). * P < .05. (C) Percentage of mast cells (MCs) in vehicle- (V) or IgE-injected pinnae exhibiting extensive (ext), moderate (mod) or no (none) degranulation 6 h after i.v. DNP-HSA challenge. *** P < .001 vs. corresponding vehicle group, +++ P < .001 vs. IgE/DNP-HSA treated Fcer1g^-/- group. (D) C3a in ear lysates 2 h after injection (i.v.) of 200 μg DNP-HSA into WT or Fcer1g^-/- mice 24 h after i.d. injection of vehicle (V; left ears) or 20 ng anti-DNP IgE (right ears). *** P < .001 vs. corresponding vehicle injected group; +++ P < .001 vs. IgE/DNP-HSA treated Fcer1g^-/- group. n.s. = not significant (P > .05).

FIG 4. C3aR and C5aR enhance ear swelling during IgE-dependent PCA reactions

Changes (Δ) in ear thickness after injection (i.v.) of 200 μg DNP-HSA into mice 24 h after i.d. injection of 20 ng anti-DNP IgE (right ears). * P < .05, *** P < .001 vs. C3aR^-/- mice, and +++ P < .001 vs. C5aR^-/- mice.

FIG 5. Enhancement of IgE-dependent PCA reactions by mast cell expression of C3aR and C5aR

(A) Changes (Δ) in ear thickness after injection (i.v.) of 200 μg DNP-HSA into mice 24 h after i.d. injection of vehicle (left ears) or 20 ng anti-DNP IgE (right ears). *** P < .001 vs. Kit^W-sh/W-sh mice; ++ P < .01, +++ P < .001 vs. C3aR^-/- BMCMCs→Kit^W-sh/W-sh mice, and ### P< .001 vs. C5aR^-/- BMCMCs→Kit^W-sh/W-sh mice. No swelling was detected in any of the left (vehicle-injected) ears. (B) Numbers of leukocytes in dermis of ear pinnae 6 h after i.v. DNP-HSA challenge. *** P < .001 for the indicated comparisons; n.s. = not significant (P > .05). (C) Percentage of mast cells (MCs) in vehicle- (V) or IgE-injected pinnae exhibiting extensive (ext), moderate (mod) or no (none) degranulation 6 h after i.v. DNP-HSA challenge (insets show MCs in Giemsa-stained, plastic-embedded 1 μm sections of ear pinnae; scale bar = 10 μm). ** P < .01, *** P < .001 vs. corresponding vehicle group, +++ P< .001 vs. IgE/DNP-HSA treated WT BMCMCs→Kit^W-sh/W-sh mice, # ## P < .001 vs. IgE/DNP-HSA treated C3aR^-/- BMCMCs→Kit^W-sh/W-sh mice.