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Hyper immunoglobulin E response in mice with monoclonal populations of B and T lymphocytes.

Curotto de Lafaille MA, Muriglan S, Sunshine MJ, Lei Y, Kutchukhidze N, Furtado GC, Wensky AK, Olivares-Villagómez D, Lafaille JJ - J. Exp. Med. (2001)

Bottom Line: This unusually high IgE response was prevented by the infusion of regulatory alpha/beta CD4(+) T cells belonging to both CD25(+) and CD25(-) subpopulations.The regulation by the infused T cells impeded the development of fully competent OVA-specific effector/memory Th2 lymphocytes without inhibiting the initial proliferative response of T cells or promoting activation-induced cell death.Our results indicate that hyper IgE responses do not occur in normal individuals due to the presence of regulatory T cells, and imply that the induction of regulatory CD4(+) T cells could be used for the prevention of atopy.

View Article: PubMed Central - PubMed

Affiliation: Program of Molecular Pathogenesis, Skirball Institute for Biomolecular Medicine, New York University School of Medicine, New York, NY 10016, USA.

ABSTRACT
A key event in the pathogenesis of allergies is the production of antibodies of the immunoglobulin (Ig)E class. In normal individuals the levels of IgE are tightly regulated, as illustrated by the low serum IgE concentration. In addition, multiple immunizations are usually required to generate detectable IgE responses in normal experimental animals. To define the parameters that regulate IgE production in vivo, we generated mice bearing monoclonal populations of B and T lymphocytes specific for influenza virus hemagglutinin (HA) and chicken ovalbumin (OVA), respectively. A single immunization of the monoclonal mice with the cross-linked OVA-HA antigen led to serum IgE levels that reached 30-200 microg/ml. This unusually high IgE response was prevented by the infusion of regulatory alpha/beta CD4(+) T cells belonging to both CD25(+) and CD25(-) subpopulations. The regulation by the infused T cells impeded the development of fully competent OVA-specific effector/memory Th2 lymphocytes without inhibiting the initial proliferative response of T cells or promoting activation-induced cell death. Our results indicate that hyper IgE responses do not occur in normal individuals due to the presence of regulatory T cells, and imply that the induction of regulatory CD4(+) T cells could be used for the prevention of atopy.

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Generation of 17/9 homologous replacement mice. (A and B) Schematic representation of the targeting of the 17/9 antibody genes into A: the heavy chain, and B: the κ light chain immunoglobulin loci. 1: germline loci; 2: targeting vectors; 3: targeted loci. J genes (JH and Jκ), μ switch region (Sμ), μ enhancer (Eμ), and κ and μ constant regions (Cκ and Cμ, respectively) are indicated. Neo-loxP indicates the lox-P flanked neomycin resistance cassette. Restriction sites: B, BamHI; C, Cla I; E, EcoRI; H, HindIII; M, MfeI; S, SmaI; X, XhoI. (C) Southern analysis of Mfe I-digested DNA from 17/9 VDJ heavy chain gene targeted ES clones before (R11 clone) and after (F3 and A9 clones) cre-mediated deletion of the neo-loxP cassette. The filter was hybridized to probe A in panel A. DNA from the untargeted J1 ES cell line is also shown. (D) Southern analysis of VJ 17/9 κ light chain targeted ES clones. BamHI/EcoRI double-digested DNA was hybridized to probe D in B. Targeted clones before (V11) and after (B1) deletion of the neo-loxP cassette. J1: untargeted ES cell line.
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fig1: Generation of 17/9 homologous replacement mice. (A and B) Schematic representation of the targeting of the 17/9 antibody genes into A: the heavy chain, and B: the κ light chain immunoglobulin loci. 1: germline loci; 2: targeting vectors; 3: targeted loci. J genes (JH and Jκ), μ switch region (Sμ), μ enhancer (Eμ), and κ and μ constant regions (Cκ and Cμ, respectively) are indicated. Neo-loxP indicates the lox-P flanked neomycin resistance cassette. Restriction sites: B, BamHI; C, Cla I; E, EcoRI; H, HindIII; M, MfeI; S, SmaI; X, XhoI. (C) Southern analysis of Mfe I-digested DNA from 17/9 VDJ heavy chain gene targeted ES clones before (R11 clone) and after (F3 and A9 clones) cre-mediated deletion of the neo-loxP cassette. The filter was hybridized to probe A in panel A. DNA from the untargeted J1 ES cell line is also shown. (D) Southern analysis of VJ 17/9 κ light chain targeted ES clones. BamHI/EcoRI double-digested DNA was hybridized to probe D in B. Targeted clones before (V11) and after (B1) deletion of the neo-loxP cassette. J1: untargeted ES cell line.

Mentions: The rearranged V(D)J genes of the 17/9 antibody were cloned from the 17/9 hybridoma (26) and used to construct targeting vectors for the heavy and κ chain Ig loci. Targeting constructs containing a neo-loxP cassette were transfected into the J1 embryonic stem (ES) cell line and G418 resistant colonies carrying homologous integrations were identified by Southern analysis. Probes A, B, C, and D in Fig. 1 were used to characterize the 5′ and 3′ targeting sites. The neo-loxP cassette was removed by cre-mediated recombination. Mice carrying both rearranged receptor genes were generated by crosses between single targeted mice, and are referred to as 17/9 mice. All mice were backcrossed onto a BALB/c background and were housed under specific pathogen-free conditions at the Skirball Institute, New York University School of Medicine. All 17/9 DO11.10 RAG−/− animals in experiments that involved transfer of CD4+CD25− splenocytes (Fig. 3 B), were given sulfamethoxazole (400 mg/liter)/trimethoprim (80 mg/liter) in the drinking water for the duration of the experiment. This treatment prevented wasting of mice transferred with CD4+CD25− cells.


Hyper immunoglobulin E response in mice with monoclonal populations of B and T lymphocytes.

Curotto de Lafaille MA, Muriglan S, Sunshine MJ, Lei Y, Kutchukhidze N, Furtado GC, Wensky AK, Olivares-Villagómez D, Lafaille JJ - J. Exp. Med. (2001)

Generation of 17/9 homologous replacement mice. (A and B) Schematic representation of the targeting of the 17/9 antibody genes into A: the heavy chain, and B: the κ light chain immunoglobulin loci. 1: germline loci; 2: targeting vectors; 3: targeted loci. J genes (JH and Jκ), μ switch region (Sμ), μ enhancer (Eμ), and κ and μ constant regions (Cκ and Cμ, respectively) are indicated. Neo-loxP indicates the lox-P flanked neomycin resistance cassette. Restriction sites: B, BamHI; C, Cla I; E, EcoRI; H, HindIII; M, MfeI; S, SmaI; X, XhoI. (C) Southern analysis of Mfe I-digested DNA from 17/9 VDJ heavy chain gene targeted ES clones before (R11 clone) and after (F3 and A9 clones) cre-mediated deletion of the neo-loxP cassette. The filter was hybridized to probe A in panel A. DNA from the untargeted J1 ES cell line is also shown. (D) Southern analysis of VJ 17/9 κ light chain targeted ES clones. BamHI/EcoRI double-digested DNA was hybridized to probe D in B. Targeted clones before (V11) and after (B1) deletion of the neo-loxP cassette. J1: untargeted ES cell line.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2195981&req=5

fig1: Generation of 17/9 homologous replacement mice. (A and B) Schematic representation of the targeting of the 17/9 antibody genes into A: the heavy chain, and B: the κ light chain immunoglobulin loci. 1: germline loci; 2: targeting vectors; 3: targeted loci. J genes (JH and Jκ), μ switch region (Sμ), μ enhancer (Eμ), and κ and μ constant regions (Cκ and Cμ, respectively) are indicated. Neo-loxP indicates the lox-P flanked neomycin resistance cassette. Restriction sites: B, BamHI; C, Cla I; E, EcoRI; H, HindIII; M, MfeI; S, SmaI; X, XhoI. (C) Southern analysis of Mfe I-digested DNA from 17/9 VDJ heavy chain gene targeted ES clones before (R11 clone) and after (F3 and A9 clones) cre-mediated deletion of the neo-loxP cassette. The filter was hybridized to probe A in panel A. DNA from the untargeted J1 ES cell line is also shown. (D) Southern analysis of VJ 17/9 κ light chain targeted ES clones. BamHI/EcoRI double-digested DNA was hybridized to probe D in B. Targeted clones before (V11) and after (B1) deletion of the neo-loxP cassette. J1: untargeted ES cell line.
Mentions: The rearranged V(D)J genes of the 17/9 antibody were cloned from the 17/9 hybridoma (26) and used to construct targeting vectors for the heavy and κ chain Ig loci. Targeting constructs containing a neo-loxP cassette were transfected into the J1 embryonic stem (ES) cell line and G418 resistant colonies carrying homologous integrations were identified by Southern analysis. Probes A, B, C, and D in Fig. 1 were used to characterize the 5′ and 3′ targeting sites. The neo-loxP cassette was removed by cre-mediated recombination. Mice carrying both rearranged receptor genes were generated by crosses between single targeted mice, and are referred to as 17/9 mice. All mice were backcrossed onto a BALB/c background and were housed under specific pathogen-free conditions at the Skirball Institute, New York University School of Medicine. All 17/9 DO11.10 RAG−/− animals in experiments that involved transfer of CD4+CD25− splenocytes (Fig. 3 B), were given sulfamethoxazole (400 mg/liter)/trimethoprim (80 mg/liter) in the drinking water for the duration of the experiment. This treatment prevented wasting of mice transferred with CD4+CD25− cells.

Bottom Line: This unusually high IgE response was prevented by the infusion of regulatory alpha/beta CD4(+) T cells belonging to both CD25(+) and CD25(-) subpopulations.The regulation by the infused T cells impeded the development of fully competent OVA-specific effector/memory Th2 lymphocytes without inhibiting the initial proliferative response of T cells or promoting activation-induced cell death.Our results indicate that hyper IgE responses do not occur in normal individuals due to the presence of regulatory T cells, and imply that the induction of regulatory CD4(+) T cells could be used for the prevention of atopy.

View Article: PubMed Central - PubMed

Affiliation: Program of Molecular Pathogenesis, Skirball Institute for Biomolecular Medicine, New York University School of Medicine, New York, NY 10016, USA.

ABSTRACT
A key event in the pathogenesis of allergies is the production of antibodies of the immunoglobulin (Ig)E class. In normal individuals the levels of IgE are tightly regulated, as illustrated by the low serum IgE concentration. In addition, multiple immunizations are usually required to generate detectable IgE responses in normal experimental animals. To define the parameters that regulate IgE production in vivo, we generated mice bearing monoclonal populations of B and T lymphocytes specific for influenza virus hemagglutinin (HA) and chicken ovalbumin (OVA), respectively. A single immunization of the monoclonal mice with the cross-linked OVA-HA antigen led to serum IgE levels that reached 30-200 microg/ml. This unusually high IgE response was prevented by the infusion of regulatory alpha/beta CD4(+) T cells belonging to both CD25(+) and CD25(-) subpopulations. The regulation by the infused T cells impeded the development of fully competent OVA-specific effector/memory Th2 lymphocytes without inhibiting the initial proliferative response of T cells or promoting activation-induced cell death. Our results indicate that hyper IgE responses do not occur in normal individuals due to the presence of regulatory T cells, and imply that the induction of regulatory CD4(+) T cells could be used for the prevention of atopy.

Show MeSH
Related in: MedlinePlus