Background/Purpose:  We have generated a mouse line with Flip conditionally deleted in CD11c-cre expressing cells (CD11c-Flip-KO, named HUPO), which spontaneously develops erosive arthritis resembling rheumatoid arthritis (RA). HUPO mice exhibit significantly reduced conventional dendritic cells (cDC), especially its CD8α⁺ subset, as well as a reduction in the number and percent of CD4⁺ T regulatory cells (Tregs). The reduced Tregs contribute to disease pathogenesis in HUPO mice, permitting the expansion of autoreactive CD4⁺T cells, plasmablasts, and joint specific autoantibodies.  This study is focused on discovering the cause for the reduced Tregs in the HUPO mice.  

Methods: Immune cell phenotyping and apoptosis/necrosis were assessed by antibody-flow cytometric analysis. HUPO mice were crossed with Rag^-/- generated HUPO-Rag^-/- double mutant line. Cells were adoptively transferred intravenously.  Cytokines were determined by quantitative ELISA.  Tregs were identified as CD4⁺CD25⁺Foxp3⁺.

Results:  HUPO-Rag-/- mice developed a very mild arthritis, which was not erosive, mediated by macrophages.   Whole spleen T and B cells were isolated from 3-5 month old control or HUPO mice and adoptively transferred into Rag^-/- or HUPO-Rag^-/- mice.  After 3 weeks, there was no difference in the percent Tregs when control lymphocytes were transferred to Rag^-/- or HUPO-Rag^-/- mice.  Further, no difference in Tregs was observed when HUPO lymphocytes were transferred to Rag^-/- mice.  In contrast, when HUPO lymphocytes were transferred into HUPO-Rag^-/- mice, after 3 weeks Tregs were significantly reduced compared to all the other groups.  Next, isolated control CD45.1⁺CD4⁺CD25⁺ T cells transferred into CD45.2 HUPO mice resulted in reduced CD45.1⁺Foxp3⁺Tregs after 2-3 weeks compared to transfer into control mice.  These data suggest that both HUPO Tregs and the in vivo environment of the HUPO mouse contribute to the reduction of Tregs.  To define the potential mechanisms, in vitro culture of the total splenocytes for 7 days resulted in significantly reduced IL-2 in the HUPO, compared with the littermate control, culture supernatants.  Further, significantly reduced Tregs were present in the HUPO, compared with the control, cultures.  Interestingly, supplement of the culture medium with IL-2 prevented the loss of Tregs in the cultures from the HUPO, but not the control mice.  The reduction of IL-2 was not due to cDCs, however, the production of immuno-reactive TGFβ,  important for Treg differentiation, was significantly reduced with BM differentiated DCs from HUPO, compared with control, mice

Conclusion:  HUPO mice represent a novel arthritis model that may provide insights into the mechanisms that contribute to the initiation of RA.  This study suggests that the mechanisms for the reduction of Tregs in HUPO mice are multifactorial and include a reduction of cDC TGFβ and a reduction of IL-2 which is essential for maintenance of Tregs. Many earlier studied documented the reduction of IL-2 in patients with RA. Our observations suggest that IL-2 may be an effective treatment strategy in the HUPO mouse model of RA.   

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/impaired-regulatory-t-cell-survival-in-the-pathogenesis-of-autoreactive-arthritis-mediated-by-cd11c-deletion-of-flip/