Background Abnormal selection of self-reactive B cells has been shown to occur at the transitional B cell stage, the tolerance checkpoint II, in systemic lupus erythematosus (SLE). This study investigated novel mechanisms of IFN-beta (IFNb)-dependent tolerance loss of transitional B cells.
Materials and methods Using a La-peptide specific tetramer, La13-27 autoreactive B cells from the spleens of B6 and autoimmune BXD2 mice were analysed for the development of CD93+ transitional B cell subsets. Mice were treated with IFNa, IFNb, or anti-IFNAR to induce or block type I IFNs. qRT-PCR was used to determine expression of IFN and genes involved in type I IFN induction and responses. IFNB1 expression in human SLE patients and mouse B cells was determined by intracellular flow cytometry analysis.
Results Enhanced IFNAR provided a needed signal to promote transitional (CD93+) autoreactive (La13-27+) B cell maturation and survival in BXD2 mice. IFNb, compared to IFNa, exhibited a more potent effect to stimulate BXD2 transitional B cells. Surprisingly, there was abnormal elevation of IFNb in transitional T1 B cells of BXD2 mice. Autocrine production and stimulation by type I IFN was necessary for optimal anti-IgM-induced transitional B cell activation in purified B cells from BXD2, and the effect was abrogated by IFNAR blockade. Despite the higher expression of IFNb, there was lower expression of genes involved in nucleic acid sensing and TLR pathway (Rig1, Mda5, Pkr, Zbp1, Irf3, and Irf7) in BXD2 T1 B cells, compared to B6 T1 B cells, suggesting non-conventional induction of Ifnb in BXD2 T1 B cells. Interestingly, in vivo immune complex stimulation enhanced Ifnb levels in BXD2 T1 cells. Further, BXD2 but not B6 T1 B cells were susceptible to anti-IgM induction of IFNb. Higher expression of Ifnb1 was also found in La(+) B cells compared to La(-) B cells, suggesting that BCR stimulation may provide a signal to enhance type I IFN expression in BXD2 B cells. Similar to the mouse finding, elevation of IFNb was identified in 9G4+ transitional B cells from SLE patient, compared to B cells from healthy controls.
Conclusions These results suggest that transitional B cells from BXD2 mice exhibit autocrine stimulation by intracellular IFNb. In combination with BCR signalling, this facilitates survival and maturation of autoreactive B cells that otherwise will be deleted. Identification of the molecular mechanism leading to INFb autocrine stimulation in transitional B cells should unveil new pathways for development of autoreactive B cells.
Acknowledgements This worked was supported by grants from the NIH (grants R01-AI-071110 to Dr. Mountz, R01-AI-083705 to Dr. Hsu, P30-AR-048311 and P30-AI-027767 to the UAB Comprehensive Flow Cytometry Core), the Department of Veterans Affairs Merit Review grant 1I01BX000600-01 to Dr. Mountz, P30-AR-048311 P&F Project support to Dr. Jun Li, the Lupus Foundation of America Finzi Summer Fellowship and the UAB Immunology T32 training grant support to Jennie Hamilton, and the Lupus Research Institute Novel Research Award to Dr. Hsu. Dr. Sanz is supported by NIH R37AI049660 and NIH U19 AI110483 Autoimmunity Centre of Excellence.
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