Background/Purpose:  We found that systemic lupus erythematosus (SLE) was induced experimentally by repeatedly immunizing the mice normally not prone to autoimmune diseases by any exogenous antigen so far examined (Tsumiyama K. et al. PLoS ONE 4(12):e8382, 2009). We have then proposed a novel ‘self-organized criticality theory’ that takes place when host’s immune system is overstimulated by repeated exposure to antigen to levels that surpass the immune system’s stability limit, i.e., self-organized criticality. The autoreactive lymphocyte clones, which we name autoantibody-inducing CD4 T cells (aiCD4 T cells), are newly generated via de novo T cell receptor (TCR) revision from thymus-passed non-autoreactive clones at peripheral lymphoid organs. They not only stimulated B cells to generate varieties of autoantibodies but also helped final differentiation of CD8 T cell into cytotoxic T lymphocyte (CTL) via antigen cross-presentation to induce tissue injuries identical to SLE. We further showed that the transfer of CD45RB^lo122^lo CD4 T cell of repeatedly-immunized mice could did induce autoantibodies in the naïve recipients, indicating that aiCD4 T cell seemed to belong to CD45RB^lo122^lo CD4 T subpopulation. To identify the phenotype of aiCD4 T cell in more detail, we here performed microarray analysis and cell transfer assay to show that aiCD4 T cell belongs to PD-1⁺CD45RB^lo122^loCD4 subpopulation.

Methods:  BALB/c mice were repeatedly immunized with ovalbumin (OVA). To investigate gene expression profiles of aiCD4 T cell, we performed microarray analysis (Whole Mouse Genome Microarray; Agilent Technologies) of CD45RB^lo122^lo CD4 T cell after immunization 12x with OVA. We also studied protein expression on CD45RB^lo122^lo CD4 T cell by flow cytometric analysis. CD45RB^lo122^loCD4 T cells were isolated additionally referring to PD-1 expression, and these fractionated cells were adoptively transferred into naïve recipients. Autoantibodies in sera of recipient mice were measured 2 weeks after cell transfer.

Results: Upon repeated immunization 12x with OVA, varieties of autoantibodies including RF, anti-Sm and anti-dsDNA antibodies were generated. Under microarray analysis, we compared the gene expression profile between CD45RB^lo122^lo CD4 T cell and the rest of CD4 T cell subsets after immunization 12x with OVA. We found that gene expression of programmed cell death 1 (PD-1) was increased x2 in the CD45RB^lo122^lo CD4 subset. Simultaneously, surface expression of PD-1 protein was also significantly increased in this subset: The PD-1⁺ cell was concentrated up to 25.2 ± 6.3% in CD45RB^lo122^lo CD4 T subset versus 7.5 ± 2.4% in CD45RB^hi and/or CD122^hi CD4 T subset (P < 0.001). Adoptive cell transfer from 12x immunized mice showed that RF and anti-dsDNA antibody in the transferred recipients with PD-1⁺CD45RB^lo122^lo CD4 T cell were 36.3 ± 336.0 U/ml and 0.5 ± 0.007 AU, whereas those with PD-1^–CD45RB^lo122^lo CD4 T cell were 12.6 ± 2.9 U/ml and 0.4 ± 0.002 AU, respectively (P < 0.05 and P < 0.05). These results indicated that PD-1⁺CD45RB^lo122^loCD4 T cells were capable of inducing autoantibodies in the naïve recipients.

Conclusion: The aiCD4 T cell that induces SLE belongs to a PD-1⁺CD45RB^lo122^lo CD4 subpopulation.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/pd-1cd45rblo122lo-autoantibody-inducing-cd4-t-cells-aicd4-t-cells-as-a-key-in-the-pathogenesis-of-systemic-lupus-erythematosus-sle/