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We thank Poubelle et al for their interest in our publication "Cytokine mRNA profiling identifies B cells as a major source of RANKL in rheumatoid arthritis "
We share an enthusiasm with Professor Poubelle for the role of neutrophils in the pathophysiology of rheumatoid arthritis and agree that neutrophils are often the dominant cell population in rheumatoid synovial fluid.
However, we disagr...
However, we disagree with a number of comments made in their letter. They state that we did not mention the possibility that other cell types beside B cells produce RANKL in the joint; however, this is not the case as we stated in the discussion that "previous studies have reported the expression of RANKL by RA synovial fibroblasts, T cells and chondrocytes".
We regret that due to space restrictions we did not cite their interesting work. In their studies [2, 3] which are largely based on in vitro differentiation of a myeloid cell line towards neutrophil-like cells and on in vitro culture of peripheral blood neutrophils, they have shown
increased mRNA expression and surface expression of RANKL upon stimulation. However, in their experiments on ex vivo isolated synovial fluid neutrophils, protein expression of RANKL was not significantly increased, and there was no induction of RANKL mRNA expression above that
found in peripheral blood neutrophils. Our data are consistent with this - that there is no mRNA over-expression of RANKL in neutrophils isolated from synovial fluid of RA patients ex vivo when compared to peripheral
blood cells. For us, it was a key objective to get as close as possible to the in vivo situation in the RA patient, without allowing the effects of cell culture to confound results. Hence we restricted our studies to
freshly obtained, unstimulated, ex vivo samples.
Furthermore, Poubelle et al. suggested that we should use more than one anti-RANKL antibody. Indeed, this is what we did. As mentioned in our Methods section  we used two different antibodies, a mouse monoclonal
anti-RANKL as well as a polyclonal rabbit anti-RANKL (Figure 5 Yeo et al.).
A further suggestion was that we should use more than one method to confirm our data, and indeed we three methods: used real-time PCR and flow cytometry to analyze synovial fluid cells, and immunofluorescence staining
for synovial tissue (Figure 5 Yeo et al).
Our data are clear that, among the synovial fluid cell populations isolated ex vivo, B cells are the cell type that expressed the highest level of RANKL mRNA.
 Yeo L, Toellner KM, Salmon M et al. Cytokine mRNA profiling identifies B cells as a major source of RANKL in rheumatoid arthritis. Ann Rheum Dis Jul 8. epub ahead of print.
 Poubelle PE, Chakravarti A, Fernandes MJ et al. Differential expression of RANK, RANK-L, and osteoprotegerin by synovial fluid neutrophils from patients with rheumatoid arthritis and by healthy human
blood neutrophils. Arthritis Res Ther 2007;9:R25.
 Chakravarti A, Raquil MA, Tessier P et al. Surface RANKL of Toll-like receptor 4-stimulated human neutrophils activates osteoclastic bone resorption. Blood 2009;114:1633-44.