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Functional anergy in a subpopulation of naive B cells from healthy humans that express autoreactive immunoglobulin receptors.

Duty JA, Szodoray P, Zheng NY, Koelsch KA, Zhang Q, Swiatkowski M, Mathias M, Garman L, Helms C, Nakken B, Smith K, Farris AD, Wilson PC - J. Exp. Med. (2008)

Bottom Line: Analysis of 95 recombinant antibodies expressed from the variable genes of single B(ND) cells demonstrated that they are predominantly autoreactive, binding to HEp-2 cell antigens and DNA.Upon B cell receptor cross-linkage, B(ND) cells have a reduced capacity to mobilize intracellular calcium or phosphorylate tyrosines, demonstrating that they are anergic.This is the first identification of a distinct mature human B cell subset that is naturally autoreactive and controlled by the tolerizing mechanism of functional anergy.

View Article: PubMed Central - PubMed

Affiliation: Immunobiology and Cancer, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA.

ABSTRACT
Self-reactive B cells not controlled by receptor editing or clonal deletion may become anergic. We report that fully mature human B cells negative for surface IgM and retaining only IgD are autoreactive and functionally attenuated (referred to as naive IgD(+)IgM(-) B cells [B(ND)]). These B(ND) cells typically make up 2.5% of B cells in the peripheral blood, have antibody variable region genes in germline (unmutated) configuration, and, by all current measures, are fully mature. Analysis of 95 recombinant antibodies expressed from the variable genes of single B(ND) cells demonstrated that they are predominantly autoreactive, binding to HEp-2 cell antigens and DNA. Upon B cell receptor cross-linkage, B(ND) cells have a reduced capacity to mobilize intracellular calcium or phosphorylate tyrosines, demonstrating that they are anergic. However, intense stimulation causes B(ND) cells to fully respond, suggesting that these cells could be the precursors of autoantibody secreting plasma cells in autoimmune diseases such as systemic lupus erythematosus or rheumatoid arthritis. This is the first identification of a distinct mature human B cell subset that is naturally autoreactive and controlled by the tolerizing mechanism of functional anergy.

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The BND phenotype is naive mature B cells. Peripheral blood B cells were isolated and stained with anti-CD19, CD27, IgM, and IgD, as well as one of each of the antibodies shown in the figure, to delineate phenotype differences between BND and naive populations for immaturity (A), maturity (B), or activation markers (C). Isotype controls were included and, where appropriate, CD19+ human BM, marginal zone–like peripheral blood B cells (CD19+, CD27+, IgM+, and IgD+), or fractions representing IgG memory B cells (CD27+, IgM−, and IgD−) were included for an internal positive and negative control. Histograms are representative of three to eight donors in at least three independent experiments. RT-PCR analysis was performed on sorted BND and naive cells for the presence of VpreB (CD179a) transcripts. Actin RT-PCR was included as a template control.
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fig3: The BND phenotype is naive mature B cells. Peripheral blood B cells were isolated and stained with anti-CD19, CD27, IgM, and IgD, as well as one of each of the antibodies shown in the figure, to delineate phenotype differences between BND and naive populations for immaturity (A), maturity (B), or activation markers (C). Isotype controls were included and, where appropriate, CD19+ human BM, marginal zone–like peripheral blood B cells (CD19+, CD27+, IgM+, and IgD+), or fractions representing IgG memory B cells (CD27+, IgM−, and IgD−) were included for an internal positive and negative control. Histograms are representative of three to eight donors in at least three independent experiments. RT-PCR analysis was performed on sorted BND and naive cells for the presence of VpreB (CD179a) transcripts. Actin RT-PCR was included as a template control.

Mentions: As immature B cells before completing primary selection may be autoreactive, we hypothesize that BND cells might be immature. However, BND cells do not express markers found on immature B cells such as the B cell progenitor marker CD10 and, like mature cells, express only low levels of the development markers CD38 and CD24, which are commonly used to distinguish recent BM immigrant immature/transitional B cells that are CD38hiCD24hi (Fig. 3 A) (32). Furthermore, the BND and naive populations are both positive for the BCR-associated regulator CD22 that is expressed on mature B cells (33). In addition, recent reports have demonstrated that transitional B cells, as well as CD27+ IgG+ memory cells, in circulation can be distinguished because they do not express the ATP binding cassette ABCB1 transporter and, subsequently, they retain mitochondrial dyes such as Rhodamine 123 or MitoTracker Green (MTG) (34). Like naive cells, BND cells actively extrude MTG, further demonstrating that they are neither transitional B cells nor are they memory cells (Fig. 3 A, middle). Finally, BND cells do not produce CD179a (surrogate light chain; VpreB) transcripts, which distinguishes them from an autoreactive population of naive B cells that are in the process of receptor editing (35, 36) (Fig. 3 A, bottom).


Functional anergy in a subpopulation of naive B cells from healthy humans that express autoreactive immunoglobulin receptors.

Duty JA, Szodoray P, Zheng NY, Koelsch KA, Zhang Q, Swiatkowski M, Mathias M, Garman L, Helms C, Nakken B, Smith K, Farris AD, Wilson PC - J. Exp. Med. (2008)

The BND phenotype is naive mature B cells. Peripheral blood B cells were isolated and stained with anti-CD19, CD27, IgM, and IgD, as well as one of each of the antibodies shown in the figure, to delineate phenotype differences between BND and naive populations for immaturity (A), maturity (B), or activation markers (C). Isotype controls were included and, where appropriate, CD19+ human BM, marginal zone–like peripheral blood B cells (CD19+, CD27+, IgM+, and IgD+), or fractions representing IgG memory B cells (CD27+, IgM−, and IgD−) were included for an internal positive and negative control. Histograms are representative of three to eight donors in at least three independent experiments. RT-PCR analysis was performed on sorted BND and naive cells for the presence of VpreB (CD179a) transcripts. Actin RT-PCR was included as a template control.
© Copyright Policy
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC2626668&req=5

fig3: The BND phenotype is naive mature B cells. Peripheral blood B cells were isolated and stained with anti-CD19, CD27, IgM, and IgD, as well as one of each of the antibodies shown in the figure, to delineate phenotype differences between BND and naive populations for immaturity (A), maturity (B), or activation markers (C). Isotype controls were included and, where appropriate, CD19+ human BM, marginal zone–like peripheral blood B cells (CD19+, CD27+, IgM+, and IgD+), or fractions representing IgG memory B cells (CD27+, IgM−, and IgD−) were included for an internal positive and negative control. Histograms are representative of three to eight donors in at least three independent experiments. RT-PCR analysis was performed on sorted BND and naive cells for the presence of VpreB (CD179a) transcripts. Actin RT-PCR was included as a template control.
Mentions: As immature B cells before completing primary selection may be autoreactive, we hypothesize that BND cells might be immature. However, BND cells do not express markers found on immature B cells such as the B cell progenitor marker CD10 and, like mature cells, express only low levels of the development markers CD38 and CD24, which are commonly used to distinguish recent BM immigrant immature/transitional B cells that are CD38hiCD24hi (Fig. 3 A) (32). Furthermore, the BND and naive populations are both positive for the BCR-associated regulator CD22 that is expressed on mature B cells (33). In addition, recent reports have demonstrated that transitional B cells, as well as CD27+ IgG+ memory cells, in circulation can be distinguished because they do not express the ATP binding cassette ABCB1 transporter and, subsequently, they retain mitochondrial dyes such as Rhodamine 123 or MitoTracker Green (MTG) (34). Like naive cells, BND cells actively extrude MTG, further demonstrating that they are neither transitional B cells nor are they memory cells (Fig. 3 A, middle). Finally, BND cells do not produce CD179a (surrogate light chain; VpreB) transcripts, which distinguishes them from an autoreactive population of naive B cells that are in the process of receptor editing (35, 36) (Fig. 3 A, bottom).

Bottom Line: Analysis of 95 recombinant antibodies expressed from the variable genes of single B(ND) cells demonstrated that they are predominantly autoreactive, binding to HEp-2 cell antigens and DNA.Upon B cell receptor cross-linkage, B(ND) cells have a reduced capacity to mobilize intracellular calcium or phosphorylate tyrosines, demonstrating that they are anergic.This is the first identification of a distinct mature human B cell subset that is naturally autoreactive and controlled by the tolerizing mechanism of functional anergy.

View Article: PubMed Central - PubMed

Affiliation: Immunobiology and Cancer, Oklahoma Medical Research Foundation, Oklahoma City, OK 73104, USA.

ABSTRACT
Self-reactive B cells not controlled by receptor editing or clonal deletion may become anergic. We report that fully mature human B cells negative for surface IgM and retaining only IgD are autoreactive and functionally attenuated (referred to as naive IgD(+)IgM(-) B cells [B(ND)]). These B(ND) cells typically make up 2.5% of B cells in the peripheral blood, have antibody variable region genes in germline (unmutated) configuration, and, by all current measures, are fully mature. Analysis of 95 recombinant antibodies expressed from the variable genes of single B(ND) cells demonstrated that they are predominantly autoreactive, binding to HEp-2 cell antigens and DNA. Upon B cell receptor cross-linkage, B(ND) cells have a reduced capacity to mobilize intracellular calcium or phosphorylate tyrosines, demonstrating that they are anergic. However, intense stimulation causes B(ND) cells to fully respond, suggesting that these cells could be the precursors of autoantibody secreting plasma cells in autoimmune diseases such as systemic lupus erythematosus or rheumatoid arthritis. This is the first identification of a distinct mature human B cell subset that is naturally autoreactive and controlled by the tolerizing mechanism of functional anergy.

Show MeSH
Related in: MedlinePlus