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TLR4 signaling augments B lymphocyte migration and overcomes the restriction that limits access to germinal center dark zones.

Hwang IY, Park C, Harrison K, Kehrl JH - J. Exp. Med. (2009)

Bottom Line: Intravital microscopy and in vivo tracking studies of B cells transferred to recipient mice revealed that TLR4-activated, but not nonstimulated, B cells accumulated within the dark zones of preexisting germinal centers even when transferred with antigen-specific B cells.The TLR4-activated cells persist much better than nonstimulated cells, expanding both within the memory and plasma cell compartments.TLR-mediated activation of B cells may help to feed and stabilize the spontaneous and ectopic germinal centers that are so commonly found in autoimmune individuals and that accompany chronic inflammation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Immunoregulation, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
B lymphocyte-intrinsic Toll-like receptor (TLR) signals amplify humoral immunity and can exacerbate autoimmune diseases. We identify a new mechanism by which TLR signals may contribute to autoimmunity and chronic inflammation. We show that TLR4 signaling enhances B lymphocyte trafficking into lymph nodes (LNs), induces B lymphocyte clustering and interactions within LN follicles, leads to sustained in vivo B cell proliferation, overcomes the restriction that limits the access of nonantigen-activated B cells to germinal center dark zones, and enhances the generation of memory and plasma cells. Intravital microscopy and in vivo tracking studies of B cells transferred to recipient mice revealed that TLR4-activated, but not nonstimulated, B cells accumulated within the dark zones of preexisting germinal centers even when transferred with antigen-specific B cells. The TLR4-activated cells persist much better than nonstimulated cells, expanding both within the memory and plasma cell compartments. TLR-mediated activation of B cells may help to feed and stabilize the spontaneous and ectopic germinal centers that are so commonly found in autoimmune individuals and that accompany chronic inflammation.

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LPS-activated transferred B cells accumulate in the recipient spleens. (A and B) Transferred LPS-activated B cells are found in the B cell follicle and the marginal zone of the spleen. Three groups of five mice (6 wk old, C57BL/6 CD45.2) were injected subcutaneously on day 0 with 100 µg HEL in CFA. 15 million cells of LPS-activated or nonactivated B cells from C57BL/6 CD45.1 mice were injected intravenously to recipient mice on day 7. Mice were boosted on day 28 with 100 µg HEL antigen. On day 38, the mice were sacrificed and the spleens analyzed by immunohistochemistry to detect CD35 and CD45.1 (10× and 20×; A) and to detect MAdCAM-1 versus CD45.1 (20×; B). Bars, 100 µm. The experiment was performed with five mice with similar results.
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fig7: LPS-activated transferred B cells accumulate in the recipient spleens. (A and B) Transferred LPS-activated B cells are found in the B cell follicle and the marginal zone of the spleen. Three groups of five mice (6 wk old, C57BL/6 CD45.2) were injected subcutaneously on day 0 with 100 µg HEL in CFA. 15 million cells of LPS-activated or nonactivated B cells from C57BL/6 CD45.1 mice were injected intravenously to recipient mice on day 7. Mice were boosted on day 28 with 100 µg HEL antigen. On day 38, the mice were sacrificed and the spleens analyzed by immunohistochemistry to detect CD35 and CD45.1 (10× and 20×; A) and to detect MAdCAM-1 versus CD45.1 (20×; B). Bars, 100 µm. The experiment was performed with five mice with similar results.

Mentions: To examine whether TLR4 ligand–exposed cells can augment an ongoing immune response, we immunized CD45.2 mice with HEL and, 1 wk later, transferred either TLR4 ligand–stimulated CD45.1 B cells or nonstimulated CD45.1 B cells. We measured specific Ig response in serum samples collected 3 wk after the cell transfer and again 10 d after boosting with HEL. We found that the addition of the TLR4 ligand–stimulated cells increased the HEL-specific IgM response 50% and the HEL-specific IgG1 response 15% compared with nonstimulated cells. The transfer of TLR4-activated B cells increased the HEL-specific IgG2a and IgG2b responses at day 28 (∼25%) but did not enhance the levels achieved after reexposure to antigen (Fig. S4). To determine the location of the transferred cells after antigen boosting (day 38 after transfer), we performed immunohistochemistry using either CD35 and CD45.1 or MAdCAM-1 and CD45.1. CD35 allows distinction of the B and T cell zones, whereas MAdCAM-1 delineates the marginal sinus. We found numerous transferred CD45.1 cells in the spleens from mice that had received TLR4-stimulated B cells, which contrasted with the relative paucity in the spleens of mice that had received the non–TLR4-stimulated CD45.1 cells (Fig. 7). Many of the previously stimulated CD45.1 cells resided in the B cell follicles and in the marginal zones of the CD45.2 mice (Fig. 7). In addition, we found CD45.1 cells in ongoing germinal centers in the spleens of mice that had received an antigen boost 10 d previously. Approximately 30% of the splenic germinal centers contained CD45.2 cells. In some instances, the majority of the cells within a germinal center were of CD45.2 origin, whereas in others we detected only a few or no CD45.2 cells (Fig. S5).


TLR4 signaling augments B lymphocyte migration and overcomes the restriction that limits access to germinal center dark zones.

Hwang IY, Park C, Harrison K, Kehrl JH - J. Exp. Med. (2009)

LPS-activated transferred B cells accumulate in the recipient spleens. (A and B) Transferred LPS-activated B cells are found in the B cell follicle and the marginal zone of the spleen. Three groups of five mice (6 wk old, C57BL/6 CD45.2) were injected subcutaneously on day 0 with 100 µg HEL in CFA. 15 million cells of LPS-activated or nonactivated B cells from C57BL/6 CD45.1 mice were injected intravenously to recipient mice on day 7. Mice were boosted on day 28 with 100 µg HEL antigen. On day 38, the mice were sacrificed and the spleens analyzed by immunohistochemistry to detect CD35 and CD45.1 (10× and 20×; A) and to detect MAdCAM-1 versus CD45.1 (20×; B). Bars, 100 µm. The experiment was performed with five mice with similar results.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
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getmorefigures.php?uid=PMC2806604&req=5

fig7: LPS-activated transferred B cells accumulate in the recipient spleens. (A and B) Transferred LPS-activated B cells are found in the B cell follicle and the marginal zone of the spleen. Three groups of five mice (6 wk old, C57BL/6 CD45.2) were injected subcutaneously on day 0 with 100 µg HEL in CFA. 15 million cells of LPS-activated or nonactivated B cells from C57BL/6 CD45.1 mice were injected intravenously to recipient mice on day 7. Mice were boosted on day 28 with 100 µg HEL antigen. On day 38, the mice were sacrificed and the spleens analyzed by immunohistochemistry to detect CD35 and CD45.1 (10× and 20×; A) and to detect MAdCAM-1 versus CD45.1 (20×; B). Bars, 100 µm. The experiment was performed with five mice with similar results.
Mentions: To examine whether TLR4 ligand–exposed cells can augment an ongoing immune response, we immunized CD45.2 mice with HEL and, 1 wk later, transferred either TLR4 ligand–stimulated CD45.1 B cells or nonstimulated CD45.1 B cells. We measured specific Ig response in serum samples collected 3 wk after the cell transfer and again 10 d after boosting with HEL. We found that the addition of the TLR4 ligand–stimulated cells increased the HEL-specific IgM response 50% and the HEL-specific IgG1 response 15% compared with nonstimulated cells. The transfer of TLR4-activated B cells increased the HEL-specific IgG2a and IgG2b responses at day 28 (∼25%) but did not enhance the levels achieved after reexposure to antigen (Fig. S4). To determine the location of the transferred cells after antigen boosting (day 38 after transfer), we performed immunohistochemistry using either CD35 and CD45.1 or MAdCAM-1 and CD45.1. CD35 allows distinction of the B and T cell zones, whereas MAdCAM-1 delineates the marginal sinus. We found numerous transferred CD45.1 cells in the spleens from mice that had received TLR4-stimulated B cells, which contrasted with the relative paucity in the spleens of mice that had received the non–TLR4-stimulated CD45.1 cells (Fig. 7). Many of the previously stimulated CD45.1 cells resided in the B cell follicles and in the marginal zones of the CD45.2 mice (Fig. 7). In addition, we found CD45.1 cells in ongoing germinal centers in the spleens of mice that had received an antigen boost 10 d previously. Approximately 30% of the splenic germinal centers contained CD45.2 cells. In some instances, the majority of the cells within a germinal center were of CD45.2 origin, whereas in others we detected only a few or no CD45.2 cells (Fig. S5).

Bottom Line: Intravital microscopy and in vivo tracking studies of B cells transferred to recipient mice revealed that TLR4-activated, but not nonstimulated, B cells accumulated within the dark zones of preexisting germinal centers even when transferred with antigen-specific B cells.The TLR4-activated cells persist much better than nonstimulated cells, expanding both within the memory and plasma cell compartments.TLR-mediated activation of B cells may help to feed and stabilize the spontaneous and ectopic germinal centers that are so commonly found in autoimmune individuals and that accompany chronic inflammation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Immunoregulation, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

ABSTRACT
B lymphocyte-intrinsic Toll-like receptor (TLR) signals amplify humoral immunity and can exacerbate autoimmune diseases. We identify a new mechanism by which TLR signals may contribute to autoimmunity and chronic inflammation. We show that TLR4 signaling enhances B lymphocyte trafficking into lymph nodes (LNs), induces B lymphocyte clustering and interactions within LN follicles, leads to sustained in vivo B cell proliferation, overcomes the restriction that limits the access of nonantigen-activated B cells to germinal center dark zones, and enhances the generation of memory and plasma cells. Intravital microscopy and in vivo tracking studies of B cells transferred to recipient mice revealed that TLR4-activated, but not nonstimulated, B cells accumulated within the dark zones of preexisting germinal centers even when transferred with antigen-specific B cells. The TLR4-activated cells persist much better than nonstimulated cells, expanding both within the memory and plasma cell compartments. TLR-mediated activation of B cells may help to feed and stabilize the spontaneous and ectopic germinal centers that are so commonly found in autoimmune individuals and that accompany chronic inflammation.

Show MeSH
Related in: MedlinePlus