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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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Pleural B1a B cells accumulating in the lung produce opsonizing IgM that is sufficient to confer survival. (A) Intracellular IgM (IgM (ic)) reservoirs in steady-state pleural GFP+ B1a cells (left dot plot) and in GFP+ B1a cells that had infiltrated LPS-challenged lungs after intrapleural transfer (right dot plot). The dotted line represents the upper 99% limit of intracellular IgM staining in steady-state cells. Representative analysis from n = 4 is shown. (B) Mean fluorescence intensity (MFI) of IgM (ic) from A (n = 4). (C) IgM ELISPOT analysis of cells as in A. Representative analysis from n = 2 experiments is shown. (D) Kaplan-Meier survival curves in response to E. coli infection in WT mice; µMT mice; and µMT mice that received WT pleural B cells in the pleural space, Csf2−/− pleural cells in the pleural space, and WT blood cells into the blood at the time of infection (n = 10 mice). (E) Opsonization of bacteria with IgM. Data show Pkh26-labeled E. coli retrieved from the BAL of either µMT mice or µMT mice spiked with pleural WT B cells in the pleural space. Bacteria (i.t.) and cell transfer (i.pls.) were conducted 6 h before BAL. An antibody against IgM shows opsonization of labeled bacteria. A representative analysis of n = 3 is shown. (F) IgM is polyclonal. WT mice received PBS or LPS. 6 h later, BAL was collected, and capacity of IgM to bind to S. aureus and P. aeruginosa was measured. Data show binding relative to a commercially available polyclonal IgM (n = 3). (G) IgM ELISA of pleural fluid and BAL after E. coli infection. µMT mice received WT or Csf2−/− pleural B1a cells into the pleural space at the time of infection (n = 3). Relevant data are presented as mean ± SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001.
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fig8: Pleural B1a B cells accumulating in the lung produce opsonizing IgM that is sufficient to confer survival. (A) Intracellular IgM (IgM (ic)) reservoirs in steady-state pleural GFP+ B1a cells (left dot plot) and in GFP+ B1a cells that had infiltrated LPS-challenged lungs after intrapleural transfer (right dot plot). The dotted line represents the upper 99% limit of intracellular IgM staining in steady-state cells. Representative analysis from n = 4 is shown. (B) Mean fluorescence intensity (MFI) of IgM (ic) from A (n = 4). (C) IgM ELISPOT analysis of cells as in A. Representative analysis from n = 2 experiments is shown. (D) Kaplan-Meier survival curves in response to E. coli infection in WT mice; µMT mice; and µMT mice that received WT pleural B cells in the pleural space, Csf2−/− pleural cells in the pleural space, and WT blood cells into the blood at the time of infection (n = 10 mice). (E) Opsonization of bacteria with IgM. Data show Pkh26-labeled E. coli retrieved from the BAL of either µMT mice or µMT mice spiked with pleural WT B cells in the pleural space. Bacteria (i.t.) and cell transfer (i.pls.) were conducted 6 h before BAL. An antibody against IgM shows opsonization of labeled bacteria. A representative analysis of n = 3 is shown. (F) IgM is polyclonal. WT mice received PBS or LPS. 6 h later, BAL was collected, and capacity of IgM to bind to S. aureus and P. aeruginosa was measured. Data show binding relative to a commercially available polyclonal IgM (n = 3). (G) IgM ELISA of pleural fluid and BAL after E. coli infection. µMT mice received WT or Csf2−/− pleural B1a cells into the pleural space at the time of infection (n = 3). Relevant data are presented as mean ± SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Mentions: The leukocyte profile and the differences between the two delivery routes suggested that the pleural space was the preferential source of lung-accumulating serosal B cells. B1a B cells retrieved from the lung contained large reservoirs of IgM (Fig. 8, A and B), which they secreted locally, as measured by ELISPOT assays performed on GFP+ cells that had relocated from the pleural space to lung tissue (Fig. 8 C). To determine the importance of pleural B cells to the host response, we profiled mortality in WT and µMT (i.e., B cell–deficient) mice infected with a high dose of E. coli. After 48 h, ∼40% of WT mice died but ∼60% completely recovered. In contrast, ∼80% of B cell–deficient µMT mice died within 12 h (Fig. 8 D). Additionally, we used ICAPS to transfer pleural B cells from WT mice to the pleural spaces of µMT mice. The µMT pleural B cell recipients, which now contained B cells but only in the pleural space, were then infected with E. coli. Remarkably, this B cell supplementation completely reversed the severe mortality otherwise observed in µMT mice (Fig. 8 D). For controls, we transferred pleural B cells from Csf2−/− mice to the pleural spaces of µMT mice and B cells sorted from the blood of WT mice to the blood (by i.v.) of µMT mice. Neither approach rescued the µMT mice, indicating that GM-CSF produced by pleural B cells was required for protection. The data show that pleural B cells defend against pneumonia.


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)

Pleural B1a B cells accumulating in the lung produce opsonizing IgM that is sufficient to confer survival. (A) Intracellular IgM (IgM (ic)) reservoirs in steady-state pleural GFP+ B1a cells (left dot plot) and in GFP+ B1a cells that had infiltrated LPS-challenged lungs after intrapleural transfer (right dot plot). The dotted line represents the upper 99% limit of intracellular IgM staining in steady-state cells. Representative analysis from n = 4 is shown. (B) Mean fluorescence intensity (MFI) of IgM (ic) from A (n = 4). (C) IgM ELISPOT analysis of cells as in A. Representative analysis from n = 2 experiments is shown. (D) Kaplan-Meier survival curves in response to E. coli infection in WT mice; µMT mice; and µMT mice that received WT pleural B cells in the pleural space, Csf2−/− pleural cells in the pleural space, and WT blood cells into the blood at the time of infection (n = 10 mice). (E) Opsonization of bacteria with IgM. Data show Pkh26-labeled E. coli retrieved from the BAL of either µMT mice or µMT mice spiked with pleural WT B cells in the pleural space. Bacteria (i.t.) and cell transfer (i.pls.) were conducted 6 h before BAL. An antibody against IgM shows opsonization of labeled bacteria. A representative analysis of n = 3 is shown. (F) IgM is polyclonal. WT mice received PBS or LPS. 6 h later, BAL was collected, and capacity of IgM to bind to S. aureus and P. aeruginosa was measured. Data show binding relative to a commercially available polyclonal IgM (n = 3). (G) IgM ELISA of pleural fluid and BAL after E. coli infection. µMT mice received WT or Csf2−/− pleural B1a cells into the pleural space at the time of infection (n = 3). Relevant data are presented as mean ± SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001.
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fig8: Pleural B1a B cells accumulating in the lung produce opsonizing IgM that is sufficient to confer survival. (A) Intracellular IgM (IgM (ic)) reservoirs in steady-state pleural GFP+ B1a cells (left dot plot) and in GFP+ B1a cells that had infiltrated LPS-challenged lungs after intrapleural transfer (right dot plot). The dotted line represents the upper 99% limit of intracellular IgM staining in steady-state cells. Representative analysis from n = 4 is shown. (B) Mean fluorescence intensity (MFI) of IgM (ic) from A (n = 4). (C) IgM ELISPOT analysis of cells as in A. Representative analysis from n = 2 experiments is shown. (D) Kaplan-Meier survival curves in response to E. coli infection in WT mice; µMT mice; and µMT mice that received WT pleural B cells in the pleural space, Csf2−/− pleural cells in the pleural space, and WT blood cells into the blood at the time of infection (n = 10 mice). (E) Opsonization of bacteria with IgM. Data show Pkh26-labeled E. coli retrieved from the BAL of either µMT mice or µMT mice spiked with pleural WT B cells in the pleural space. Bacteria (i.t.) and cell transfer (i.pls.) were conducted 6 h before BAL. An antibody against IgM shows opsonization of labeled bacteria. A representative analysis of n = 3 is shown. (F) IgM is polyclonal. WT mice received PBS or LPS. 6 h later, BAL was collected, and capacity of IgM to bind to S. aureus and P. aeruginosa was measured. Data show binding relative to a commercially available polyclonal IgM (n = 3). (G) IgM ELISA of pleural fluid and BAL after E. coli infection. µMT mice received WT or Csf2−/− pleural B1a cells into the pleural space at the time of infection (n = 3). Relevant data are presented as mean ± SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001.
Mentions: The leukocyte profile and the differences between the two delivery routes suggested that the pleural space was the preferential source of lung-accumulating serosal B cells. B1a B cells retrieved from the lung contained large reservoirs of IgM (Fig. 8, A and B), which they secreted locally, as measured by ELISPOT assays performed on GFP+ cells that had relocated from the pleural space to lung tissue (Fig. 8 C). To determine the importance of pleural B cells to the host response, we profiled mortality in WT and µMT (i.e., B cell–deficient) mice infected with a high dose of E. coli. After 48 h, ∼40% of WT mice died but ∼60% completely recovered. In contrast, ∼80% of B cell–deficient µMT mice died within 12 h (Fig. 8 D). Additionally, we used ICAPS to transfer pleural B cells from WT mice to the pleural spaces of µMT mice. The µMT pleural B cell recipients, which now contained B cells but only in the pleural space, were then infected with E. coli. Remarkably, this B cell supplementation completely reversed the severe mortality otherwise observed in µMT mice (Fig. 8 D). For controls, we transferred pleural B cells from Csf2−/− mice to the pleural spaces of µMT mice and B cells sorted from the blood of WT mice to the blood (by i.v.) of µMT mice. Neither approach rescued the µMT mice, indicating that GM-CSF produced by pleural B cells was required for protection. The data show that pleural B cells defend against pneumonia.

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