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Pleural innate response activator B cells protect against pneumonia via a GM-CSF-IgM axis.

Weber GF, Chousterman BG, Hilgendorf I, Robbins CS, Theurl I, Gerhardt LM, Iwamoto Y, Quach TD, Ali M, Chen JW, Rothstein TL, Nahrendorf M, Weissleder R, Swirski FK - J. Exp. Med. (2014)

Bottom Line: We show that in response to lung infection, B1a B cells migrate from the pleural space to the lung parenchyma to secrete polyreactive emergency immunoglobulin M (IgM).The strategic location of these cells, coupled with the capacity to produce GM-CSF-dependent IgM, ensures effective early frontline defense against bacteria invading the lungs.The study describes a previously unrecognized GM-CSF-IgM axis and positions IRA B cells as orchestrators of protective IgM immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Systems Biology, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114 Department of Visceral, Thoracic and Vascular Surgery, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, 01307 Dresden, Germany fswirski@mgh.harvard.edu georg.weber@uniklinikum-dresden.de.

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Generation of mixed chimeras. (A) Leukocyte reconstitution in WT/µMT, GM/WT, and GM/µMT mice in the pleural space, blood, and lung 10 wk after irradiation and bone marrow transfer. Data show the relative percentages of myeloid (CD11b+) and nonmyeloid (CD11b−) cells in the three compartments (n = 4, mean ± SD). (B) B cells in the pleural space of a WT mouse. Gating strategy for B1a (CD19+IgMhighCD43+CD5+), B1b (CD19+IgMhighCD43+CD5−), B2 (CD19+IgMlowCD43−CD5−), and B reg (CD19+IgM+CD43−CD5+) B cells is shown. (C) Relative proportions of total B cells and B1a B cells in the pleural space of WT/µMT, GM/WT, and GM/µMT chimeric mice and non-irradiated WT mice in the steady state. The chimeric mice were analyzed 8 wk after bone marrow transfer (n = 3–5, mean ± SD). (D) Enumeration of the various subsets in the three chimeras (n = 3–5, mean ± SD). (E) CD45.2 expression on blood leukocytes in CD45.1+ mice that had been lethally irradiated and reconstituted with CD45.2+ bone marrow cells 10 wk earlier. The plot is representative from that of the three chimeras. (F) Lung H&E in the three sets of chimeric mice 10 wk after bone marrow reconstitution (bars, 200 µm). (G) Turbidity analysis of BAL at 600 nm from WT/µMT, GM/µMT, WTGM (lethal irradiation of WT mouse and 100% reconstitution with Csf2−/− BM), and GMWT (lethal irradiation of Csf2−/− mouse and 100% reconstitution with WT BM) chimera (n = 3; mean ± SD). (H) BAL of WT/µMT, GM/WT, GM/µMT, WTGM, and GMWT chimeric mice.
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fig4: Generation of mixed chimeras. (A) Leukocyte reconstitution in WT/µMT, GM/WT, and GM/µMT mice in the pleural space, blood, and lung 10 wk after irradiation and bone marrow transfer. Data show the relative percentages of myeloid (CD11b+) and nonmyeloid (CD11b−) cells in the three compartments (n = 4, mean ± SD). (B) B cells in the pleural space of a WT mouse. Gating strategy for B1a (CD19+IgMhighCD43+CD5+), B1b (CD19+IgMhighCD43+CD5−), B2 (CD19+IgMlowCD43−CD5−), and B reg (CD19+IgM+CD43−CD5+) B cells is shown. (C) Relative proportions of total B cells and B1a B cells in the pleural space of WT/µMT, GM/WT, and GM/µMT chimeric mice and non-irradiated WT mice in the steady state. The chimeric mice were analyzed 8 wk after bone marrow transfer (n = 3–5, mean ± SD). (D) Enumeration of the various subsets in the three chimeras (n = 3–5, mean ± SD). (E) CD45.2 expression on blood leukocytes in CD45.1+ mice that had been lethally irradiated and reconstituted with CD45.2+ bone marrow cells 10 wk earlier. The plot is representative from that of the three chimeras. (F) Lung H&E in the three sets of chimeric mice 10 wk after bone marrow reconstitution (bars, 200 µm). (G) Turbidity analysis of BAL at 600 nm from WT/µMT, GM/µMT, WTGM (lethal irradiation of WT mouse and 100% reconstitution with Csf2−/− BM), and GMWT (lethal irradiation of Csf2−/− mouse and 100% reconstitution with WT BM) chimera (n = 3; mean ± SD). (H) BAL of WT/µMT, GM/WT, GM/µMT, WTGM, and GMWT chimeric mice.

Mentions: To determine whether IRA B cells are important in the host response to airway infection, we generated mixed chimeric mice with a B cell–restricted GM-CSF deficiency (Fig. 4). The procedure involved lethal irradiation of WT mice and reconstitution with a mixture of bone marrow cells from µMT and GM-CSF–deficient mice. Accordingly, the µMT bone marrow gave rise to GM-CSF–sufficient leukocytes, but not B cells, whereas the GM-CSF–deficient bone marrow gave rise to GM-CSF–deficient leukocytes, including B cells. After 10 wk of reconstitution, all B cells (which necessarily derived from GM-CSF–deficient mice) lacked the capacity to produce GM-CSF in the mixed chimeras (GM/µMT). The remaining leukocytes were a mixture of WT and GM-CSF–deficient cells, whereas radiation-resistant and tissue-resident nonhematopoietic cells were GM-CSF–sufficient. We also generated two types of controls: WT mice reconstituted with a mixture of WT and µMT bone marrow cells (WT/µMT), which controlled for the µMT contribution, and WT mice reconstituted with a mixture of GM-CSF–deficient and WT bone marrow cells (GM/WT), which controlled for the contribution of any GM-CSF–deficient non–B cells in the GM/µMT group. The leukocyte profiles in the pleural space, blood, and lung in the steady-state were most similar between the GM/µMT and WT/µMT chimeras (Fig. 4 A), but all three chimeras developed various B cell subsets (Baumgarth, 2011), including B1a B cells, as expected (Düber et al., 2009; Esplin et al., 2009; Holodick et al., 2009). Even though reconstitution was sub-optimal compared with non-irradiated WT mice (Fig. 4, B–D), the chimeras’ leukocytes were of donor origin (Fig. 4 E), and the mice had normal lung histology (Fig. 4 F) without evidence of alveolar proteinosis (Fig. 4, G and H). Complete GM-CSF deficiency leads to the development of spontaneous alveolar proteinosis (Dranoff et al., 1994; Stanley et al., 1994). Previous data have shown (Huffman et al., 1996), and our data confirm, that pulmonary epithelial (i.e., nonhematopoietic) GM-CSF–producing cells prevent proteinosis by stimulating alveolar macrophages to clear surfactant; B cells are dispensable in this context.


Pleural innate response activator B cells protect against pneumonia via a GM-CSF-IgM axis.

Weber GF, Chousterman BG, Hilgendorf I, Robbins CS, Theurl I, Gerhardt LM, Iwamoto Y, Quach TD, Ali M, Chen JW, Rothstein TL, Nahrendorf M, Weissleder R, Swirski FK - J. Exp. Med. (2014)

Generation of mixed chimeras. (A) Leukocyte reconstitution in WT/µMT, GM/WT, and GM/µMT mice in the pleural space, blood, and lung 10 wk after irradiation and bone marrow transfer. Data show the relative percentages of myeloid (CD11b+) and nonmyeloid (CD11b−) cells in the three compartments (n = 4, mean ± SD). (B) B cells in the pleural space of a WT mouse. Gating strategy for B1a (CD19+IgMhighCD43+CD5+), B1b (CD19+IgMhighCD43+CD5−), B2 (CD19+IgMlowCD43−CD5−), and B reg (CD19+IgM+CD43−CD5+) B cells is shown. (C) Relative proportions of total B cells and B1a B cells in the pleural space of WT/µMT, GM/WT, and GM/µMT chimeric mice and non-irradiated WT mice in the steady state. The chimeric mice were analyzed 8 wk after bone marrow transfer (n = 3–5, mean ± SD). (D) Enumeration of the various subsets in the three chimeras (n = 3–5, mean ± SD). (E) CD45.2 expression on blood leukocytes in CD45.1+ mice that had been lethally irradiated and reconstituted with CD45.2+ bone marrow cells 10 wk earlier. The plot is representative from that of the three chimeras. (F) Lung H&E in the three sets of chimeric mice 10 wk after bone marrow reconstitution (bars, 200 µm). (G) Turbidity analysis of BAL at 600 nm from WT/µMT, GM/µMT, WTGM (lethal irradiation of WT mouse and 100% reconstitution with Csf2−/− BM), and GMWT (lethal irradiation of Csf2−/− mouse and 100% reconstitution with WT BM) chimera (n = 3; mean ± SD). (H) BAL of WT/µMT, GM/WT, GM/µMT, WTGM, and GMWT chimeric mice.
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fig4: Generation of mixed chimeras. (A) Leukocyte reconstitution in WT/µMT, GM/WT, and GM/µMT mice in the pleural space, blood, and lung 10 wk after irradiation and bone marrow transfer. Data show the relative percentages of myeloid (CD11b+) and nonmyeloid (CD11b−) cells in the three compartments (n = 4, mean ± SD). (B) B cells in the pleural space of a WT mouse. Gating strategy for B1a (CD19+IgMhighCD43+CD5+), B1b (CD19+IgMhighCD43+CD5−), B2 (CD19+IgMlowCD43−CD5−), and B reg (CD19+IgM+CD43−CD5+) B cells is shown. (C) Relative proportions of total B cells and B1a B cells in the pleural space of WT/µMT, GM/WT, and GM/µMT chimeric mice and non-irradiated WT mice in the steady state. The chimeric mice were analyzed 8 wk after bone marrow transfer (n = 3–5, mean ± SD). (D) Enumeration of the various subsets in the three chimeras (n = 3–5, mean ± SD). (E) CD45.2 expression on blood leukocytes in CD45.1+ mice that had been lethally irradiated and reconstituted with CD45.2+ bone marrow cells 10 wk earlier. The plot is representative from that of the three chimeras. (F) Lung H&E in the three sets of chimeric mice 10 wk after bone marrow reconstitution (bars, 200 µm). (G) Turbidity analysis of BAL at 600 nm from WT/µMT, GM/µMT, WTGM (lethal irradiation of WT mouse and 100% reconstitution with Csf2−/− BM), and GMWT (lethal irradiation of Csf2−/− mouse and 100% reconstitution with WT BM) chimera (n = 3; mean ± SD). (H) BAL of WT/µMT, GM/WT, GM/µMT, WTGM, and GMWT chimeric mice.
Mentions: To determine whether IRA B cells are important in the host response to airway infection, we generated mixed chimeric mice with a B cell–restricted GM-CSF deficiency (Fig. 4). The procedure involved lethal irradiation of WT mice and reconstitution with a mixture of bone marrow cells from µMT and GM-CSF–deficient mice. Accordingly, the µMT bone marrow gave rise to GM-CSF–sufficient leukocytes, but not B cells, whereas the GM-CSF–deficient bone marrow gave rise to GM-CSF–deficient leukocytes, including B cells. After 10 wk of reconstitution, all B cells (which necessarily derived from GM-CSF–deficient mice) lacked the capacity to produce GM-CSF in the mixed chimeras (GM/µMT). The remaining leukocytes were a mixture of WT and GM-CSF–deficient cells, whereas radiation-resistant and tissue-resident nonhematopoietic cells were GM-CSF–sufficient. We also generated two types of controls: WT mice reconstituted with a mixture of WT and µMT bone marrow cells (WT/µMT), which controlled for the µMT contribution, and WT mice reconstituted with a mixture of GM-CSF–deficient and WT bone marrow cells (GM/WT), which controlled for the contribution of any GM-CSF–deficient non–B cells in the GM/µMT group. The leukocyte profiles in the pleural space, blood, and lung in the steady-state were most similar between the GM/µMT and WT/µMT chimeras (Fig. 4 A), but all three chimeras developed various B cell subsets (Baumgarth, 2011), including B1a B cells, as expected (Düber et al., 2009; Esplin et al., 2009; Holodick et al., 2009). Even though reconstitution was sub-optimal compared with non-irradiated WT mice (Fig. 4, B–D), the chimeras’ leukocytes were of donor origin (Fig. 4 E), and the mice had normal lung histology (Fig. 4 F) without evidence of alveolar proteinosis (Fig. 4, G and H). Complete GM-CSF deficiency leads to the development of spontaneous alveolar proteinosis (Dranoff et al., 1994; Stanley et al., 1994). Previous data have shown (Huffman et al., 1996), and our data confirm, that pulmonary epithelial (i.e., nonhematopoietic) GM-CSF–producing cells prevent proteinosis by stimulating alveolar macrophages to clear surfactant; B cells are dispensable in this context.

Bottom Line: We show that in response to lung infection, B1a B cells migrate from the pleural space to the lung parenchyma to secrete polyreactive emergency immunoglobulin M (IgM).The strategic location of these cells, coupled with the capacity to produce GM-CSF-dependent IgM, ensures effective early frontline defense against bacteria invading the lungs.The study describes a previously unrecognized GM-CSF-IgM axis and positions IRA B cells as orchestrators of protective IgM immunity.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Systems Biology, Massachusetts General Hospital, Harvard Medical School, Boston, MA 02114 Department of Visceral, Thoracic and Vascular Surgery, Medizinische Fakultät Carl Gustav Carus, Technische Universität Dresden, 01307 Dresden, Germany fswirski@mgh.harvard.edu georg.weber@uniklinikum-dresden.de.

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Related in: MedlinePlus