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Pasteurella multocida toxin-stimulated osteoclast differentiation is B cell dependent.

Infection and immunity (2010-10-20)
Dagmar Hildebrand, Klaus Heeg, Katharina F Kubatzky
ABSTRACT

Pasteurella multocida is a Gram-negative bacillus that infects a number of wild and domestic animals, causing respiratory diseases. Toxigenic Pasteurella multocida strains produce a protein toxin (PMT) that leads to atrophic rhinitis in swine due to enhanced osteoclastogenesis and the inhibition of osteoblast function. We show that PMT-induced osteoclastogenesis is promoted by an as-yet-uncharacterized B-cell population. The toxin, however, is not acting at the level of hematopoietic stem cells, since purified CD117(+) cells from murine hematopoietic progenitor cells cultivated with PMT did not mature into osteoclasts. The early macrophages contained within this cell population (CD117(+)/CD11b(+)) did not further differentiate into osteoclasts but survived and were able to phagocytose. Within the CD117(-) population, however, we detected PMT-induced generation of a B220(+)/CD19(+) and B220(+)/IgM(+) B-cell population that was able to take up fluorescently labeled PMT. Using purified B-cell and macrophage populations, we show that these B cells are needed to efficiently generate osteoclasts from macrophages. Cells of the immune system are thought to affect osteoclast formation and function by secreting cytokines and growth factors. We show here that PMT-stimulated B cells produce elevated levels of the osteoclastogenic factors interleukin-1β (IL-1β), IL-6, tumor necrosis factor alpha, and receptor activator of nuclear factor receptor ligand (RANKL) compared to B cells generated through incubation with IL-7. These results suggest that the osteoclastic properties characteristic for PMT may result from a cross talk between bone cells and lymphoid cells and that B cells might be an important target of Pasteurella multocida.

MATERIALS
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Product Description

Sigma-Aldrich
Latex beads, carboxylate-modified polystyrene, fluorescent yellow-green, aqueous suspension, 2.0 μm mean particle size
Sigma-Aldrich
Atto 647, BioReagent, suitable for fluorescence, ≥90% (HPCE)
Sigma-Aldrich
Atto 647 NHS ester, BioReagent, suitable for fluorescence, ≥80% (coupling to amines)
Sigma-Aldrich
Atto 647 maleimide, BioReagent, suitable for fluorescence, ≥90% (coupling to thiols)

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