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Severe attenuation of the B cell immune response in Msh2-deficient mice.

Vora KA, Tumas-Brundage KM, Lentz VM, Cranston A, Fishel R, Manser T - J. Exp. Med. (1999)

Bottom Line: Recently, results obtained from mice with targeted inactivations of postreplication DNA mismatch repair (MMR) genes have been interpreted to demonstrate a direct role for MMR in antibody variable (V) gene hypermutation.These include lack of progression of the germinal center (GC) reaction associated with increased intra-GC apoptosis, severely diminished antigen-specific immunoglobulin G responses, and near absence of anamnestic responses.Interpretation of the impact of an MMR deficiency on the mechanism of V gene somatic hypermutation could be easily confounded by these perturbations.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology and The Kimmel Cancer Institute, Jefferson Medical College, Philadelphia, Pennsylvania 19107, USA.

ABSTRACT
Recently, results obtained from mice with targeted inactivations of postreplication DNA mismatch repair (MMR) genes have been interpreted to demonstrate a direct role for MMR in antibody variable (V) gene hypermutation. Here we show that mice that do not express the MMR factor Msh2 have wide-ranging defects in antigen-driven B cell responses. These include lack of progression of the germinal center (GC) reaction associated with increased intra-GC apoptosis, severely diminished antigen-specific immunoglobulin G responses, and near absence of anamnestic responses. Mice heterozygous for the Msh2 deficiency display an "intermediate" phenotype in these regards, suggesting that normal levels of Msh2 expression are critical for the B cell response. Interpretation of the impact of an MMR deficiency on the mechanism of V gene somatic hypermutation could be easily confounded by these perturbations.

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Serum antibody responses of Msh2-deficient mice. Immunization was performed with either NP-CGG (A, B, and C) or TNP-Ficoll (D), as described in Materials and Methods, and mice were bled at various times thereafter. Levels of antigen-specific antibody of a particular isotype were determined by  ELISA. In many of these assays, anti-NP mAbs with similar affinities for NP were used as standards, allowing the results to be presented as micrograms per milliliter of these standards. In other assays, relative levels of antigen-binding antibodies in different sera were determined using serial dilutions. The points at which  the resulting OD curves were 50% maximal were then used to calculate the relative dilution factor giving an equivalent OD for each serum sample. The data  illustrated in A and C were obtaining using sera pooled from at least three different animals of the each genotype. The data illustrated in B and D were obtained  from at least three individual sera per time point, and error bars are shown (except for the +/+ anti–TNP-Ficoll data, which were obtained from one animal).
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Figure 3: Serum antibody responses of Msh2-deficient mice. Immunization was performed with either NP-CGG (A, B, and C) or TNP-Ficoll (D), as described in Materials and Methods, and mice were bled at various times thereafter. Levels of antigen-specific antibody of a particular isotype were determined by ELISA. In many of these assays, anti-NP mAbs with similar affinities for NP were used as standards, allowing the results to be presented as micrograms per milliliter of these standards. In other assays, relative levels of antigen-binding antibodies in different sera were determined using serial dilutions. The points at which the resulting OD curves were 50% maximal were then used to calculate the relative dilution factor giving an equivalent OD for each serum sample. The data illustrated in A and C were obtaining using sera pooled from at least three different animals of the each genotype. The data illustrated in B and D were obtained from at least three individual sera per time point, and error bars are shown (except for the +/+ anti–TNP-Ficoll data, which were obtained from one animal).

Mentions: These observations led us to conduct detailed analysis of the anti–NP-CGG serum antibody responses of Msh2-deficient mice. As shown in Fig. 3 A, at early stages of this response, antigen-specific IgM levels were only slightly lower in Msh2 +/− and −/− mice compared with +/+ mice. However, dramatic differences in the levels of NP-specific κ, λ, and IgG isotypes were apparent between Msh2 +/+ and −/− mice at all times in the primary response, with Msh2 +/− mice displaying an intermediate phenotype in most cases (Fig. 3, A and B). Msh2 +/− mice also showed a delayed serum IgG1 response (peaking at day 21 instead of 14), and expressed increased levels of IgG2a, although this isotype was a minor component of the total response in all mice. Also readily apparent was the severely diminished or reduced secondary serum antibody response in Msh2 −/− and +/− mice, respectively. In −/− mice, this secondary response was of a magnitude not noticeably different from peak levels in the primary response in most assays. This result was not idiosyncratic to the anti-NP response, as the secondary response to the carrier, CGG, was also severely blunted in −/− mice and reduced in +/− mice (Fig. 3 C). Analysis of the serum antibody response to TNP-Ficoll, a T cell–independent antigen, revealed analogous IgM responses in +/+ and −/− mice, but the total (kappa) anti-TNP response decayed more rapidly in Msh2 −/− mice. This more rapid decay appeared to be accounted for by a severe deficiency in the IgG3 response, particularly at late times after immunization (Fig. 3 D). Interestingly, total serum IgG levels were found not to differ significantly in Msh2 +/+, +/−, and −/− mice (data not shown), suggesting that long-term homeostatic regulation of these levels is not perturbed by an Msh2 deficiency.


Severe attenuation of the B cell immune response in Msh2-deficient mice.

Vora KA, Tumas-Brundage KM, Lentz VM, Cranston A, Fishel R, Manser T - J. Exp. Med. (1999)

Serum antibody responses of Msh2-deficient mice. Immunization was performed with either NP-CGG (A, B, and C) or TNP-Ficoll (D), as described in Materials and Methods, and mice were bled at various times thereafter. Levels of antigen-specific antibody of a particular isotype were determined by  ELISA. In many of these assays, anti-NP mAbs with similar affinities for NP were used as standards, allowing the results to be presented as micrograms per milliliter of these standards. In other assays, relative levels of antigen-binding antibodies in different sera were determined using serial dilutions. The points at which  the resulting OD curves were 50% maximal were then used to calculate the relative dilution factor giving an equivalent OD for each serum sample. The data  illustrated in A and C were obtaining using sera pooled from at least three different animals of the each genotype. The data illustrated in B and D were obtained  from at least three individual sera per time point, and error bars are shown (except for the +/+ anti–TNP-Ficoll data, which were obtained from one animal).
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Related In: Results  -  Collection

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Figure 3: Serum antibody responses of Msh2-deficient mice. Immunization was performed with either NP-CGG (A, B, and C) or TNP-Ficoll (D), as described in Materials and Methods, and mice were bled at various times thereafter. Levels of antigen-specific antibody of a particular isotype were determined by ELISA. In many of these assays, anti-NP mAbs with similar affinities for NP were used as standards, allowing the results to be presented as micrograms per milliliter of these standards. In other assays, relative levels of antigen-binding antibodies in different sera were determined using serial dilutions. The points at which the resulting OD curves were 50% maximal were then used to calculate the relative dilution factor giving an equivalent OD for each serum sample. The data illustrated in A and C were obtaining using sera pooled from at least three different animals of the each genotype. The data illustrated in B and D were obtained from at least three individual sera per time point, and error bars are shown (except for the +/+ anti–TNP-Ficoll data, which were obtained from one animal).
Mentions: These observations led us to conduct detailed analysis of the anti–NP-CGG serum antibody responses of Msh2-deficient mice. As shown in Fig. 3 A, at early stages of this response, antigen-specific IgM levels were only slightly lower in Msh2 +/− and −/− mice compared with +/+ mice. However, dramatic differences in the levels of NP-specific κ, λ, and IgG isotypes were apparent between Msh2 +/+ and −/− mice at all times in the primary response, with Msh2 +/− mice displaying an intermediate phenotype in most cases (Fig. 3, A and B). Msh2 +/− mice also showed a delayed serum IgG1 response (peaking at day 21 instead of 14), and expressed increased levels of IgG2a, although this isotype was a minor component of the total response in all mice. Also readily apparent was the severely diminished or reduced secondary serum antibody response in Msh2 −/− and +/− mice, respectively. In −/− mice, this secondary response was of a magnitude not noticeably different from peak levels in the primary response in most assays. This result was not idiosyncratic to the anti-NP response, as the secondary response to the carrier, CGG, was also severely blunted in −/− mice and reduced in +/− mice (Fig. 3 C). Analysis of the serum antibody response to TNP-Ficoll, a T cell–independent antigen, revealed analogous IgM responses in +/+ and −/− mice, but the total (kappa) anti-TNP response decayed more rapidly in Msh2 −/− mice. This more rapid decay appeared to be accounted for by a severe deficiency in the IgG3 response, particularly at late times after immunization (Fig. 3 D). Interestingly, total serum IgG levels were found not to differ significantly in Msh2 +/+, +/−, and −/− mice (data not shown), suggesting that long-term homeostatic regulation of these levels is not perturbed by an Msh2 deficiency.

Bottom Line: Recently, results obtained from mice with targeted inactivations of postreplication DNA mismatch repair (MMR) genes have been interpreted to demonstrate a direct role for MMR in antibody variable (V) gene hypermutation.These include lack of progression of the germinal center (GC) reaction associated with increased intra-GC apoptosis, severely diminished antigen-specific immunoglobulin G responses, and near absence of anamnestic responses.Interpretation of the impact of an MMR deficiency on the mechanism of V gene somatic hypermutation could be easily confounded by these perturbations.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology and Immunology and The Kimmel Cancer Institute, Jefferson Medical College, Philadelphia, Pennsylvania 19107, USA.

ABSTRACT
Recently, results obtained from mice with targeted inactivations of postreplication DNA mismatch repair (MMR) genes have been interpreted to demonstrate a direct role for MMR in antibody variable (V) gene hypermutation. Here we show that mice that do not express the MMR factor Msh2 have wide-ranging defects in antigen-driven B cell responses. These include lack of progression of the germinal center (GC) reaction associated with increased intra-GC apoptosis, severely diminished antigen-specific immunoglobulin G responses, and near absence of anamnestic responses. Mice heterozygous for the Msh2 deficiency display an "intermediate" phenotype in these regards, suggesting that normal levels of Msh2 expression are critical for the B cell response. Interpretation of the impact of an MMR deficiency on the mechanism of V gene somatic hypermutation could be easily confounded by these perturbations.

Show MeSH
Related in: MedlinePlus