Background/Purpose: Alterations in B cell immune tolerance are important in the development of autoimmune rheumatoid arthritis (RA). Recent studies in healthy human subjects have identified a subset of potentially autoreactive CD19+CD27-IgD+IgMlow/- late transitional cells that constitute ~2.5% of peripheral blood B cells and have anergic signaling properties. These cells are hyporesponsive to antigen receptor stimulation in vitroand have the capacity to produce autoreactive antibodies (Abs). The function and signaling properties of these cells have not been characterized in RA (or any other human autoimmune disorder to date). Early alterations in signaling profile of these cells are of significant value for the prediction of the onset of autoimmunity as pathogenic Abs and other clinical manifestations of RA are detectable only after a prolonged period of autoantibody expansion. 

Methods: Phosphorylation of intracellular signaling proteins and intracellular Ca2+ levels in CD19+CD27-IgD+IgMlow/- B cells at baseline and in response to in vitroBCR stimulation with polyclonal anti-human Ig were measured by multicolor flow cytometry in the peripheral blood cells from 32 normal healthy control donors and 20 patients with RA. Signaling properties of anergic B cells were compared to other peripheral blood B cell populations both in normal controls and RA patients.

Results: B cell signaling profiles at baseline and in response to anti-BCR stimulation were examined in 32 healthy donors (mean age 35.7; male/female ratio 0.68) and 20 RA patients (mean age 55.9; male/female ratio 0.54) from UC Denver Hospital Rheumatology Clinic. RA patients were recently diagnosed; most were serum RF and/or anti-CCP positive and have not undergone B cell targeted treatments. No significant phenotypic or quantitative differences were found between IgM+ and CD19+CD27-IgD+IgMlow/- B cell subsets obtained from RA patients and normal controls. CD19+CD27-IgD+IgMlow/- B cells from RA patients demonstrated increased baseline total phospho tyrosine (pTyr) protein phosphorylation as compared to normal controls. In contrast to the control group, in RA patients, CD19+CD27-IgD+IgMlow/- B cells total pTyr and Ca2+ responses to anti-BCR stimulation resembled those of normal non-anergic IgM+ B cells. Comparison of baseline phosphorylation levels of individual signaling molecules between unstimulated RA and control CD19+CD27-IgD+IgMlow/- B cells  revealed pronounced increases in Blnk, SHP2 and Jnk in the RA group. Comparison of signaling properties CD19+CD27-IgD+IgMlow/- B cells in RA patients and healthy controls revealed a reversal of pTyr and Ca2+ anergic signaling features in patients, accompanied by phosphorylation decreases of Blnk, Syk, SHP2, CD19. 

Conclusion: CD19+CD27-IgD+IgMlow/- B cells under normal conditions showed characteristic signaling inhibition, which was reversed in this cell population in RA patients. A distinct phosphoryation pattern for major signal transduction proteins in CD19+CD27-IgD+IgMlow/- B cells in RA patients as compared to healthy controls at baseline and in response to BCR stimulation was demonstrated.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/rheumatoid-arthritis-is-associated-with-signaling-alterations-in-naturally-occurring-autoreactive-b-lymphocytes/