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Regulation of anti-double-stranded DNA B cells in nonautoimmune mice: localization to the T-B interface of the splenic follicle.

Mandik-Nayak L, Bui A, Noorchashm H, Eaton A, Erikson J - J. Exp. Med. (1997)

Bottom Line: Because the VH3H9 H chain can pair with endogenous L chains to generate anti-single-stranded DNA, anti-dsDNA, and non-DNA B cells, this allowed us to study the regulation of anti-dsDNA B cells in the context of a diverse B cell repertoire.We have identified anti-dsDNA B cells that are located at the T-B interface in the splenic follicle where they have an increased in vivo turnover rate.These anti-dsDNA B cells exhibit a unique surface phenotype suggesting developmental arrest due to antigen exposure.

View Article: PubMed Central - PubMed

Affiliation: The Wistar Institute, Philadelphia, Pennsylvania 19104, USA.

ABSTRACT
Systemic lupus erythematosus (SLE) and the MRL-lpr/lpr murine model for SLE are characterized by the presence of serum anti-double-stranded (ds)DNA antibodies (Abs), whereas nonautoimmune individuals have negligible levels of these Abs. To increase the frequency of anti-DNA B cells and identify the mechanisms involved in their regulation in nonautoimmune mice, we have used Ig transgenes (tgs). In the present study, we used the VH3H9 heavy (H) chain tg which expresses an H chain that was repeatedly isolated from anti-dsDNA Abs from MRL-lpr/lpr mice. Because the VH3H9 H chain can pair with endogenous L chains to generate anti-single-stranded DNA, anti-dsDNA, and non-DNA B cells, this allowed us to study the regulation of anti-dsDNA B cells in the context of a diverse B cell repertoire. We have identified anti-dsDNA B cells that are located at the T-B interface in the splenic follicle where they have an increased in vivo turnover rate. These anti-dsDNA B cells exhibit a unique surface phenotype suggesting developmental arrest due to antigen exposure.

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VH3H9/λ anti-dsDNA B cells are present with a reduced Ig  density. Bone marrow (left), spleen (middle), and lymph node (right) cells  from Tg(−) (top) and VH3H9 tg (bottom) mice were stained with anti-B220-biotin/streptavidin-Red670 and anti-λ-FITC. MFI is given for the  λ+ cells in the boxed region. These are representative plots of n = 19  mice of each genotype.
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Figure 1: VH3H9/λ anti-dsDNA B cells are present with a reduced Ig density. Bone marrow (left), spleen (middle), and lymph node (right) cells from Tg(−) (top) and VH3H9 tg (bottom) mice were stained with anti-B220-biotin/streptavidin-Red670 and anti-λ-FITC. MFI is given for the λ+ cells in the boxed region. These are representative plots of n = 19 mice of each genotype.

Mentions: In this study, we use VH3H9 H chain only tg mice to increase the frequency of anti-dsDNA B cells while maintaining a polyclonal repertoire. Transfection and hybridoma analysis have identified germline Vλ1 as an L chain that pairs with the VH3H9 H chain to generate an anti-dsDNA Ab (see J558LT and MRL1-45 in Table 1; references 9, 10). Because the VH3H9 H chain tg has been shown to be a good excluder of endogenous H chain rearrangement on the BALB/c background (Table 1; references 23, 30), we can follow the fate of anti-dsDNA B cells in VH3H9 tg mice using anti-λ specific reagents. Several different reagents were used to track λ+ and λ1+ B cells (LS136, R11-153, JC5, and R26-46). Using these reagents and flow cytometry, we have shown that the majority of λ+ B cells in VH3H9 tg mice are λ1 (79 ± 19%). Therefore, for the remainder of our studies, we have used anti-pan λ reagents to detect anti-dsDNA B cells. As is shown in Fig. 1, λ+ B cells are present in the bone marrow, spleen, and lymph node. It is also apparent that the levels of λ Ig on these cells are lower in the periphery compared to λ+ B cells from tg(−) mice (spleen: mean fluorescence intensity [MFI] 52 versus 181; LN: MFI 52 versus 216). Interestingly, Ig levels are also decreased on the λ+ B cells from the bone marrow (MFI 47 versus 197). There is precedent for antigen encounter leading to a decrease in Ig density in other tolerance model systems; for example, anti–hen egg lysozyme (HEL) B cells have a reduced level of Ig when in the presence of HEL (3), but when these B cells are removed from Ag, either by in vivo parking or in vitro cultures, their surface Ig levels increase (31, 32). The decreased Ig density on the anti-dsDNA B cells suggests that these cells are encountering their Ag and that this encounter initially occurred in the bone marrow.


Regulation of anti-double-stranded DNA B cells in nonautoimmune mice: localization to the T-B interface of the splenic follicle.

Mandik-Nayak L, Bui A, Noorchashm H, Eaton A, Erikson J - J. Exp. Med. (1997)

VH3H9/λ anti-dsDNA B cells are present with a reduced Ig  density. Bone marrow (left), spleen (middle), and lymph node (right) cells  from Tg(−) (top) and VH3H9 tg (bottom) mice were stained with anti-B220-biotin/streptavidin-Red670 and anti-λ-FITC. MFI is given for the  λ+ cells in the boxed region. These are representative plots of n = 19  mice of each genotype.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199093&req=5

Figure 1: VH3H9/λ anti-dsDNA B cells are present with a reduced Ig density. Bone marrow (left), spleen (middle), and lymph node (right) cells from Tg(−) (top) and VH3H9 tg (bottom) mice were stained with anti-B220-biotin/streptavidin-Red670 and anti-λ-FITC. MFI is given for the λ+ cells in the boxed region. These are representative plots of n = 19 mice of each genotype.
Mentions: In this study, we use VH3H9 H chain only tg mice to increase the frequency of anti-dsDNA B cells while maintaining a polyclonal repertoire. Transfection and hybridoma analysis have identified germline Vλ1 as an L chain that pairs with the VH3H9 H chain to generate an anti-dsDNA Ab (see J558LT and MRL1-45 in Table 1; references 9, 10). Because the VH3H9 H chain tg has been shown to be a good excluder of endogenous H chain rearrangement on the BALB/c background (Table 1; references 23, 30), we can follow the fate of anti-dsDNA B cells in VH3H9 tg mice using anti-λ specific reagents. Several different reagents were used to track λ+ and λ1+ B cells (LS136, R11-153, JC5, and R26-46). Using these reagents and flow cytometry, we have shown that the majority of λ+ B cells in VH3H9 tg mice are λ1 (79 ± 19%). Therefore, for the remainder of our studies, we have used anti-pan λ reagents to detect anti-dsDNA B cells. As is shown in Fig. 1, λ+ B cells are present in the bone marrow, spleen, and lymph node. It is also apparent that the levels of λ Ig on these cells are lower in the periphery compared to λ+ B cells from tg(−) mice (spleen: mean fluorescence intensity [MFI] 52 versus 181; LN: MFI 52 versus 216). Interestingly, Ig levels are also decreased on the λ+ B cells from the bone marrow (MFI 47 versus 197). There is precedent for antigen encounter leading to a decrease in Ig density in other tolerance model systems; for example, anti–hen egg lysozyme (HEL) B cells have a reduced level of Ig when in the presence of HEL (3), but when these B cells are removed from Ag, either by in vivo parking or in vitro cultures, their surface Ig levels increase (31, 32). The decreased Ig density on the anti-dsDNA B cells suggests that these cells are encountering their Ag and that this encounter initially occurred in the bone marrow.

Bottom Line: Because the VH3H9 H chain can pair with endogenous L chains to generate anti-single-stranded DNA, anti-dsDNA, and non-DNA B cells, this allowed us to study the regulation of anti-dsDNA B cells in the context of a diverse B cell repertoire.We have identified anti-dsDNA B cells that are located at the T-B interface in the splenic follicle where they have an increased in vivo turnover rate.These anti-dsDNA B cells exhibit a unique surface phenotype suggesting developmental arrest due to antigen exposure.

View Article: PubMed Central - PubMed

Affiliation: The Wistar Institute, Philadelphia, Pennsylvania 19104, USA.

ABSTRACT
Systemic lupus erythematosus (SLE) and the MRL-lpr/lpr murine model for SLE are characterized by the presence of serum anti-double-stranded (ds)DNA antibodies (Abs), whereas nonautoimmune individuals have negligible levels of these Abs. To increase the frequency of anti-DNA B cells and identify the mechanisms involved in their regulation in nonautoimmune mice, we have used Ig transgenes (tgs). In the present study, we used the VH3H9 heavy (H) chain tg which expresses an H chain that was repeatedly isolated from anti-dsDNA Abs from MRL-lpr/lpr mice. Because the VH3H9 H chain can pair with endogenous L chains to generate anti-single-stranded DNA, anti-dsDNA, and non-DNA B cells, this allowed us to study the regulation of anti-dsDNA B cells in the context of a diverse B cell repertoire. We have identified anti-dsDNA B cells that are located at the T-B interface in the splenic follicle where they have an increased in vivo turnover rate. These anti-dsDNA B cells exhibit a unique surface phenotype suggesting developmental arrest due to antigen exposure.

Show MeSH
Related in: MedlinePlus