IgG-mediated suppression of primary IgM- and IgG-responses in FcγRIIB KO and FcRγ KO mice.

<p>FcγRIIB KO, FcRγ KO, and BALB/c mice were immunized with 10 μg IgG^b anti-SRBC and 5x10⁶ SRBC, SRBC alone, or IgG alone. (A,C) Five days after immunization, the number of spleen cells producing IgM anti-SRBC was assayed. Responses are shown as percentage of the direct PFC response/spleen in mice given SRBC alone (100%, open bars); black bars show responses in mice given IgG and SRBC. Direct PFC/spleen in the respective control groups (receiving antigen alone) were in A: BALB/c, 12,218; FcγRIIB KO, 36,391 (p<0.05) and in C: BALB/c, 57,279; FcRγ KO 40,831. (B,D) Seven to 49 days later, serum levels of IgG anti-SRBC were assayed in ELISA on sera diluted 1:625. Data are representative of one (A, C) or pooled from three (B, D) experiments with each KO strain. p-values denote comparisons between mice immunized with IgG anti-SRBC together with SRBC and mice immunized with SRBC alone. **, p < 0.01; ***, p < 0.001.</p

IgE immune complex challenge induces similar lung MCp levels in CD23^−/− as in wild type.

<p>(A–B) A representative experiment showing the quantification of MCp from wild type (WT) or CD23^−/− mice sensitized with OVA/alum and challenged with IgE-anti-TNP/OVA-TNP. The error bars shown are SEM. (C) A summary of all the four experiments performed expressed as a stimulation index (S.I.) where the numbers represent the mean number of MCp/lung or MCp/10⁶MNC per experiment of sensitized WT mice challenged with IgE-anti-TNP/OVA-TNP divided by the mean number obtained from CD23^−/− mice treated in parallel. The mean of the S.I. from all experiments is given in bold text at the bottom. The representative experiment shown in (A–B) is indicated in bold italics. There was no statistical difference (ns) in the tested parameters using a two-way ANOVA from a comparison of all individual mice from each group from the four experiments.</p

Challenge with IgE immune complex enhance lung MCp numbers compared to antigen alone.

<p>(A–C) A representative experiment showing the quantification of MCp and MNC from wild type (WT) mice sensitized with OVA/alum and challenged with OVA-TNP, IgE-anti-TNP/OVA-TNP or IgE-anti-TNP alone. The error bars shown are SEM. (D) A summary of all nine experiments performed expressed as a stimulation index (S.I.) where the numbers represent the mean number of MCp/lung, MNC/lung or MCp/10⁶ MNC per experiment of sensitized mice challenged with IgE-anti-TNP/OVA-TNP divided by the mean number obtained from sensitized mice challenged with OVA-TNP alone for each experiment. (E) A summary of all the experiments where B-cells, T-cells and dendritic cells (DC) were analyzed with flow cytometry. In some experiments flow cytometry was not done (nd). The mean of the S.I. from all experiments is given in bold text at the bottom. The experiment shown in (A–C) is indicated in bold italics. The p-values shown are derived from two-way ANOVA from a comparison of all individual mice in each group from the nine experiments.</p

IgE immune complex-induced enhancement of lung MCp is dependent on an FcRγ chain associated receptor.

<p>(A–B) A representative experiment showing the quantification of MCp from or FcRγ^−/− sensitized with OVA/alum and challenged with either IgE-anti-TNP/OVA-TNP or OVA-TNP alone (C) A summary of all the three experiments performed expressed as a stimulation index (S.I.) where the numbers represent the mean number of MCp/lung or MCp/10⁶MNC of sensitized FcRγ^−/− mice challenged with IgE-anti-TNP/OVA-TNP divided by the mean number obtained in FcRγ^−/− mice challenged with OVA-TNP alone. The mean of the S.I. from all experiments is given in bold text at the bottom. The representative experiment shown in (A–B) is indicated in bold italics. There was no statistical difference (ns) in the tested parameters using a two-way ANOVA from a comparison of all individual mice from each group from the three experiments.</p

FcRγ-chain deficient mice have a reduced number of lung MCp after IgE immune complex challenge.

<p>(A–B) A representative experiment showing the quantification of MCp from wild type (WT) or FcRγ^−/− sensitized with OVA/alum and challenged with IgE-anti-TNP/OVA-TNP (C) A summary of all the four experiments performed expressed as a stimulation index (S.I.) where the numbers represent the mean number of MCp/lung or MCp/10⁶ MNC of sensitized WT mice challenged with IgE-anti-TNP/OVA-TNP divided by the mean number obtained in FcRγ^−/− mice treated in parallel. The mean of the S.I. from all experiments is given in bold text at the bottom. The representative experiment shown in (A–B) is indicated in bold italics. The p-values shown are derived using a two-way ANOVA from a comparison of all individual mice from each group from the four experiments.</p

Intranasal challenge with OVA-TNP does not enhance the lung MCp number over basal levels.

<p>(A–C) One experiment showing the quantification of MCp and MNC from wild type (WT) mice sensitized with OVA/alum and challenged with either OVA-TNP alone, OVA aerosol or left unchallenged. The error bars shown are SEM. (D) A comparison of the OVA-TNP challenged and the unchallenged group from the two experiments performed expressed as a stimulation index (S.I.) where the numbers represent the mean number of MCp/lung, MNC/lung or MCp/10⁶ MNC per experiment of sensitized mice challenged with OVA-TNP divided by the mean number obtained from sensitized mice left unchallenged. The mean of the S.I. from the experiments is given in bold text at the bottom. The experiment shown in (A–C) is indicated in bold italics. There was no statistical difference (ns) between the OVA-TNP challenged and the unchallenged mice in the tested parameters using a two-way ANOVA from a comparison of all individual mice from each group from the two experiments.</p

01_A_S3A_069

Κωδικός τεκμηρίου: 01_A_S3A_069-019. Είδος τεκμηρίου: SLIDES 10Χ12,5 Χ 1 ΚΑΡΕ - ΕΓΧΡΩΜΑ. Ανήκει σε: 01_A_S3A_069 - ΚΟΥΤΙ "ΔΙΑΦΑΝΕΙΕΣ ΜΕΓ.

CR1/2 on FDCs are required for a robust IgG anti-SRBC response to SRBC.

<p>BALB/c and <i>Cr2^−/−</i> mice were irradiated and reconstituted with either BALB/c or <i>Cr2^−/−</i> bone marrow. Six weeks after reconstitution, mice (n = 6/group) were immunized i.v. with 5×10⁶, 5×10⁷, or 5×10⁸ SRBC. All mice were bled at indicated time points. Sera were diluted 1∶125 (A) or 1∶625 (B and C) and screened for IgG anti-SRBC in ELISA. P-values represent comparisons between the responses in recipients with the same background; ns = p>0.05; * = p<0.05; ** = p<0.01; *** = p<0.001. Representative of two (A) or one (B, C) experiments.</p

IgE and CD23 are associated with bexosomes.

<p>(A) Bexosomes were isolated from B cell cultures stimulated with anti-CD40± IL-4 or IgE as indicated. 1.2511 fold increase ±0.177SD between lanes three and four. Equal bexosome protein was loaded on each lane and the blot was probed with polyclonal anti-CD23. Blot was probed with anti-MHC class II or anti-CD9and the result is shown at the bottom of (A). (B) Exosomes from B cell cultures stimulated with anti-CD40 and IgE (All lanes). IL-4 stimulation is as indicated. Lanes 1 and 2 are bexosomes from WT mice, Lanes, 3 and 4 are bexosomes from CD23^−/− mice. Blots are representative of at least three independent experiments. (C) Graph represents fold difference in densitometry between anti-CD23 blots for anti-CD40/IL4 and anti-CD40/IL4/IgE.</p

Bexosome-induced antigen specific T cell proliferation is enhanced by IgE, only in the presence of CD23and in vivo IgE immune complex T proliferation is enhanced in ADAM10B^−/− mice.

<p>(A) B cell cultures were stimulated as in <i>Methods</i>. Stimulated B cells from either WT or CD23^−/− mice were pre-incubated ±IgE for 24 hours and then IgE/Ag ICs were added for an additional 24 hours. Bexosomes were isolated and cultured with purified Ag-specific DO11 T cells for 3 days. Proliferation was determined using a [3H]-thymidine pulse (<i>Methods</i>). (B,C) On day −1 WT or ADAM10^B−/− mice were adoptively transferred with Ag-specific DO11 T cells and on day 0 immunized with IgE/Ag ICs (<i>Methods</i>); on day 3, spleens were removed and examined for expanded Ag specific DO11⁺ CD4⁺ T cells by flow cytometry (B) or immunohistochemistry (C). *p>0.05, ***p>0.0005; n = at least five per group.</p