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Outer periarteriolar lymphoid sheath arrest and subsequent differentiation of both naive and tolerant immunoglobulin transgenic B cells is determined by B cell receptor occupancy.

Cook MC, Basten A, Fazekas de St Groth B - J. Exp. Med. (1997)

Bottom Line: In addition, tolerant B cells were found to relocate from the follicles to the outer PALS of HEL/anti-HEL double Tg mice in which the concentration of soluble self-antigen had been increased by zinc feeding.Thus, subtle variations in antigen concentration resulted in dramatic changes in positioning of B cells within the spleen.A series of mixed bone marrow chimeras in which the effective antigen concentration was inversely related to the number of self-reactive B cells due to absorption of antigen by transgene-encoded membrane and secreted Ig, was used to confirm that alteration in B cell position previously attributed to changes in follicular composition could be explained on the basis of available antigen concentration, rather than the diversity of the repertoire.

View Article: PubMed Central - PubMed

Affiliation: Centenary Institute for Cancer Medicine and Cell Biology, Newtown, Sydney, New South Wales, Australia 2042.

ABSTRACT
T-dependent B cell responses in the spleen are initiated in the outer periarteriolar lymphoid sheath (PALS) and culminate in the generation of proliferative foci and germinal center reactions. By pulsing anti-hen egg lysozyme (HEL) immunoglobulin transgenic (IgTg) B cells with various concentrations of HEL in vitro before adoptive transfer into normal recipients, it was shown that a critical number of B cell receptors (BCRs) must be ligated for B cells to undergo arrest in the outer PALS. T cell help was manipulated independently of the BCR stimulus by incubating B cells expressing the appropriate major histocompatibility complex class II antigen with a peptide recognized by CD4(+) TCR Tg T cells. B cells which either failed to arrest in the outer PALS due to a subthreshold BCR stimulus, or arrested only transiently due to the brevity of the BCR stimulus, underwent an abortive response within the follicles when provided with T cell help. In contrast, naive B cells stimulated by a sustained, suprathreshold concentration of either foreign or self-antigen and given T cell help, proliferated in the outer PALS and then differentiated. Outer PALS arrest was not influenced by the nature of the B cells occupying the follicle, but appeared to be determined solely by the magnitude of BCR stimulation. Thus antigen-pulsed B cells arrested in the outer PALS in an identical manner irrespective of whether the follicles comprised a population of normal B cells with multiple specificities, a monoclonal naive population, or a monoclonal population of tolerant B cells. In addition, tolerant B cells were found to relocate from the follicles to the outer PALS of HEL/anti-HEL double Tg mice in which the concentration of soluble self-antigen had been increased by zinc feeding. Similarly, when anti-HEL Tg mice were crossed with a second HEL Tg strain expressing a higher concentration of soluble HEL, the tolerant anti-HEL Tg B cells were located constitutively in the outer PALS. Thus, subtle variations in antigen concentration resulted in dramatic changes in positioning of B cells within the spleen. A series of mixed bone marrow chimeras in which the effective antigen concentration was inversely related to the number of self-reactive B cells due to absorption of antigen by transgene-encoded membrane and secreted Ig, was used to confirm that alteration in B cell position previously attributed to changes in follicular composition could be explained on the basis of available antigen concentration, rather than the diversity of the repertoire.

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Location of self-reactive B cells is determined by  the prevailing concentration of  self-antigen. Fluorescent micrographs of frozen sections of  spleen. HEL-binding B cells are  stained green (fluorescein) and  B220+ cells are stained red  (Texas red). (A and B) Sections  of spleen from MD4 × ML5  double Tg mice treated with oral  zinc sulfate (25 mM). The section obtained 1 d after starting  zinc but before a change in serum HEL concentration (A)  shows HEL-binding B cells predominantly within the follicles  (F), whereas the section obtained  4 d after starting zinc, 1 d after  the increase in serum HEL concentration, shows relocation of  HEL-binding B cells to the outer  PALS (B). (C and D) Sections of  spleen from unmanipulated  MD4 × AL3 double Tg mice.  HEL-binding B cells are located  in the follicles in male mice (C),  but in the outer PALS in females  (D). (E and F) Sections of spleen  from mixed bone marrow chimeras. The section from a HEL  Tg (ML5) mouse reconstituted  with IgTg/non-Tg bone marrow in a ratio of 95:5 shows  HEL-binding B cells distributed  throughout the follicles and  PALS (E). A section from a HEL  Tg (ML5) mouse reconstituted  with IgTg/non-Tg marrow in a  ratio of 80:20 shows HEL-binding B cells predominantly in the  outer PALS (F).
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Figure 4: Location of self-reactive B cells is determined by the prevailing concentration of self-antigen. Fluorescent micrographs of frozen sections of spleen. HEL-binding B cells are stained green (fluorescein) and B220+ cells are stained red (Texas red). (A and B) Sections of spleen from MD4 × ML5 double Tg mice treated with oral zinc sulfate (25 mM). The section obtained 1 d after starting zinc but before a change in serum HEL concentration (A) shows HEL-binding B cells predominantly within the follicles (F), whereas the section obtained 4 d after starting zinc, 1 d after the increase in serum HEL concentration, shows relocation of HEL-binding B cells to the outer PALS (B). (C and D) Sections of spleen from unmanipulated MD4 × AL3 double Tg mice. HEL-binding B cells are located in the follicles in male mice (C), but in the outer PALS in females (D). (E and F) Sections of spleen from mixed bone marrow chimeras. The section from a HEL Tg (ML5) mouse reconstituted with IgTg/non-Tg bone marrow in a ratio of 95:5 shows HEL-binding B cells distributed throughout the follicles and PALS (E). A section from a HEL Tg (ML5) mouse reconstituted with IgTg/non-Tg marrow in a ratio of 80:20 shows HEL-binding B cells predominantly in the outer PALS (F).

Mentions: The phenotype of tolerant B cells from the soluble HEL double Tg mice resembles that of a subset of primary follicular and follicular mantle zone B cells from normal mice (IgMloIgDhi) (29). Histological examination of the spleens of double Tg mice has revealed that they are deficient in marginal zones in which memory cells are thought to reside (30), whereas the organization of PALS and follicles appears to be relatively normal (31). An alternative method of investigating the importance of follicular composition on B cell location was to determine the effect of the concentration of serum HEL on the in situ distribution of tolerant B cells in intact double Tg mice. This was achieved by administering zinc supplements to MD4 × ML5 double Tg mice in which the HEL transgene is under the transcriptional regulation of the zinc-inducible mouse metallothionein 1 promoter (15; summarized in Table 2). Before induction, the HEL-binding B cells were distributed throughout the primary splenic follicles (Fig. 4 A). However, on day 4 after zinc induction, i.e., 24 h after the serum HEL concentration had peaked, the HEL-binding B cells had redistributed to the outer PALS (Fig. 4 B), despite the fact that the percentage of these cells had remained unchanged during this period.


Outer periarteriolar lymphoid sheath arrest and subsequent differentiation of both naive and tolerant immunoglobulin transgenic B cells is determined by B cell receptor occupancy.

Cook MC, Basten A, Fazekas de St Groth B - J. Exp. Med. (1997)

Location of self-reactive B cells is determined by  the prevailing concentration of  self-antigen. Fluorescent micrographs of frozen sections of  spleen. HEL-binding B cells are  stained green (fluorescein) and  B220+ cells are stained red  (Texas red). (A and B) Sections  of spleen from MD4 × ML5  double Tg mice treated with oral  zinc sulfate (25 mM). The section obtained 1 d after starting  zinc but before a change in serum HEL concentration (A)  shows HEL-binding B cells predominantly within the follicles  (F), whereas the section obtained  4 d after starting zinc, 1 d after  the increase in serum HEL concentration, shows relocation of  HEL-binding B cells to the outer  PALS (B). (C and D) Sections of  spleen from unmanipulated  MD4 × AL3 double Tg mice.  HEL-binding B cells are located  in the follicles in male mice (C),  but in the outer PALS in females  (D). (E and F) Sections of spleen  from mixed bone marrow chimeras. The section from a HEL  Tg (ML5) mouse reconstituted  with IgTg/non-Tg bone marrow in a ratio of 95:5 shows  HEL-binding B cells distributed  throughout the follicles and  PALS (E). A section from a HEL  Tg (ML5) mouse reconstituted  with IgTg/non-Tg marrow in a  ratio of 80:20 shows HEL-binding B cells predominantly in the  outer PALS (F).
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Figure 4: Location of self-reactive B cells is determined by the prevailing concentration of self-antigen. Fluorescent micrographs of frozen sections of spleen. HEL-binding B cells are stained green (fluorescein) and B220+ cells are stained red (Texas red). (A and B) Sections of spleen from MD4 × ML5 double Tg mice treated with oral zinc sulfate (25 mM). The section obtained 1 d after starting zinc but before a change in serum HEL concentration (A) shows HEL-binding B cells predominantly within the follicles (F), whereas the section obtained 4 d after starting zinc, 1 d after the increase in serum HEL concentration, shows relocation of HEL-binding B cells to the outer PALS (B). (C and D) Sections of spleen from unmanipulated MD4 × AL3 double Tg mice. HEL-binding B cells are located in the follicles in male mice (C), but in the outer PALS in females (D). (E and F) Sections of spleen from mixed bone marrow chimeras. The section from a HEL Tg (ML5) mouse reconstituted with IgTg/non-Tg bone marrow in a ratio of 95:5 shows HEL-binding B cells distributed throughout the follicles and PALS (E). A section from a HEL Tg (ML5) mouse reconstituted with IgTg/non-Tg marrow in a ratio of 80:20 shows HEL-binding B cells predominantly in the outer PALS (F).
Mentions: The phenotype of tolerant B cells from the soluble HEL double Tg mice resembles that of a subset of primary follicular and follicular mantle zone B cells from normal mice (IgMloIgDhi) (29). Histological examination of the spleens of double Tg mice has revealed that they are deficient in marginal zones in which memory cells are thought to reside (30), whereas the organization of PALS and follicles appears to be relatively normal (31). An alternative method of investigating the importance of follicular composition on B cell location was to determine the effect of the concentration of serum HEL on the in situ distribution of tolerant B cells in intact double Tg mice. This was achieved by administering zinc supplements to MD4 × ML5 double Tg mice in which the HEL transgene is under the transcriptional regulation of the zinc-inducible mouse metallothionein 1 promoter (15; summarized in Table 2). Before induction, the HEL-binding B cells were distributed throughout the primary splenic follicles (Fig. 4 A). However, on day 4 after zinc induction, i.e., 24 h after the serum HEL concentration had peaked, the HEL-binding B cells had redistributed to the outer PALS (Fig. 4 B), despite the fact that the percentage of these cells had remained unchanged during this period.

Bottom Line: In addition, tolerant B cells were found to relocate from the follicles to the outer PALS of HEL/anti-HEL double Tg mice in which the concentration of soluble self-antigen had been increased by zinc feeding.Thus, subtle variations in antigen concentration resulted in dramatic changes in positioning of B cells within the spleen.A series of mixed bone marrow chimeras in which the effective antigen concentration was inversely related to the number of self-reactive B cells due to absorption of antigen by transgene-encoded membrane and secreted Ig, was used to confirm that alteration in B cell position previously attributed to changes in follicular composition could be explained on the basis of available antigen concentration, rather than the diversity of the repertoire.

View Article: PubMed Central - PubMed

Affiliation: Centenary Institute for Cancer Medicine and Cell Biology, Newtown, Sydney, New South Wales, Australia 2042.

ABSTRACT
T-dependent B cell responses in the spleen are initiated in the outer periarteriolar lymphoid sheath (PALS) and culminate in the generation of proliferative foci and germinal center reactions. By pulsing anti-hen egg lysozyme (HEL) immunoglobulin transgenic (IgTg) B cells with various concentrations of HEL in vitro before adoptive transfer into normal recipients, it was shown that a critical number of B cell receptors (BCRs) must be ligated for B cells to undergo arrest in the outer PALS. T cell help was manipulated independently of the BCR stimulus by incubating B cells expressing the appropriate major histocompatibility complex class II antigen with a peptide recognized by CD4(+) TCR Tg T cells. B cells which either failed to arrest in the outer PALS due to a subthreshold BCR stimulus, or arrested only transiently due to the brevity of the BCR stimulus, underwent an abortive response within the follicles when provided with T cell help. In contrast, naive B cells stimulated by a sustained, suprathreshold concentration of either foreign or self-antigen and given T cell help, proliferated in the outer PALS and then differentiated. Outer PALS arrest was not influenced by the nature of the B cells occupying the follicle, but appeared to be determined solely by the magnitude of BCR stimulation. Thus antigen-pulsed B cells arrested in the outer PALS in an identical manner irrespective of whether the follicles comprised a population of normal B cells with multiple specificities, a monoclonal naive population, or a monoclonal population of tolerant B cells. In addition, tolerant B cells were found to relocate from the follicles to the outer PALS of HEL/anti-HEL double Tg mice in which the concentration of soluble self-antigen had been increased by zinc feeding. Similarly, when anti-HEL Tg mice were crossed with a second HEL Tg strain expressing a higher concentration of soluble HEL, the tolerant anti-HEL Tg B cells were located constitutively in the outer PALS. Thus, subtle variations in antigen concentration resulted in dramatic changes in positioning of B cells within the spleen. A series of mixed bone marrow chimeras in which the effective antigen concentration was inversely related to the number of self-reactive B cells due to absorption of antigen by transgene-encoded membrane and secreted Ig, was used to confirm that alteration in B cell position previously attributed to changes in follicular composition could be explained on the basis of available antigen concentration, rather than the diversity of the repertoire.

Show MeSH
Related in: MedlinePlus