Background We previously obtained a FcγRIIB-deficient C57BL/6 (B6) congenic strain of mice, which spontaneously developed severe rheumatoid arthritis (RA). The introduction of the Yaa (Y-linked autoimmune acceleration) mutation, which is a consequence of a translocation from the telomeric end of the X chromosome containing the Tlr7 gene onto the Y chromosome, to the FcγRIIB-deficient B6 (B6.FcγRIIB-/-.Yaa) mice developed lupus like nephritis but not RA. This phenotype conversion did not depend on autoantibody specificity, since the mice showed a marked increase in serum levels of both lupus-related and RA related autoantibodies.
Objectives To examine the mechanism of the disease phenotype conversion from RA (B6.FcγRIIB-/- mice) to lupus (B6.FcγRIIB-/-.Yaa mice), we compared subpopulations of peripheral blood mononuclear cells and spleen cells, also cytokine expression of stimulated spleen cells.
Methods Peripheral blood mononuclear cells were analyzed by flow cytometry and two distinct subpopulation of Gr-1 positive and negative monocytes were analyzed among B6, B6.Yaa, B6.FcγRIIB-/-, and B6.FcγRIIB-/-.Yaa mice at 2 months of age. Comparisons of cell surface phenotypes of splenic lymphocytes were analyzed using flow cytometry. Spleen cells were triple-stained and the frequency of marginal zone (MZ) B cells, activation/maturation status of B cells, peanut agglutinin (PNA)+ germinal center (GC) B cells, and CD138+ plasma cells were analyzed. Also activation/maturation status of T cells, PD1+ICOS+CD4+ and PD1+CXCR5+CD4+ follicular helper T cells were analyzed. Quantitative real-time PCR analysis was performed to compare mRNA expression levels of notable cytokines in spleen between B6.FcγRIIB-/- and B6.FcγRIIB-/-.Yaa mice at 4 months of age. PMA/ionomycin-stimulated spleen cells from these mice were also analyzed by flow cytometry.
Results Flow cytometric analysis of peripheral blood monocytes from 2-month-old mice revealed monocytosis in B6.FcγRIIB-/-.Yaa mice but not in B6.FcγRIIB-/- mice. The frequency of Gr-1- “resident” monocytes was increased in B6.FcγRIIB-/-.Yaa mice. Flow cytometric analysis of spleen cells from 4-month-old mice revealed that, while there were no differences in frequencies of B220+ B cells per total spleen cells among four strains, there was a significant decrease in frequencies of CD21+CD23- marginal zone B cells in B6.Yaa and B6.FcγRIIB-/-.Yaa mice. As for the activation/maturation status of B cells, frequencies of CD69+ activated B cells, PNA+ GC B cells, and CD138+ plasma cells were significantly higher in B6.FcγRIIB-/-.Yaa mice than those found in other strains of mice. As for T cells, the frequency of CD69+ activated T cells was markedly increased in B6.FcγRIIB-/-.Yaa mice. As for about comparisons of cytokine synthesis, the expression of IL-6, IL-10 and IL-21 were up regulated in B6.FcγRIIB-/-.Yaa mice. Both IL-10 and IL-21 were secreted from CD4+ T cells and the frequencies of IL-10 and IL-21-secreting cells per total CD4+ T cells were significantly higher in B6.FcγRIIB-/-.Yaa mice than those in B6.FcγRIIB-/- mice.
Conclusions The present studies suggest that the mechanism of the disease phenotype conversion from RA to lupus was likely to be due to the activation of monocytes, B cells and T cells leading to the change in cytokine milieus from the activated follicular helper T cells.
Disclosure of Interest None declared