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. 2016 Nov;101(11):e440–e444. doi: 10.3324/haematol.2016.149278

Figure 3.

Figure 3.

IL-6Rα signaling in T cells directly regulates T-follicular helper cell (TFH) differentiation in response to stored HOD red blood cell (RBC) transfusions. To determine whether direct IL-6Rα signaling in T cells regulates their differentiation into antigen specific TFH cells in response to transfusions with stored HOD RBCs, TFH differentiation in IL-6RαKO OT-II and WT OT-II cells was directly compared in wild-type (WT) recipients transfused with HOD RBCs. (A) Representative flow cytometry contour plots depicting the net expansion of WT and IL-6RαKO (KO) OT-II cells in response to fresh or stored HOD RBC transfusions. Panels show total gated CD4+ population in splenocytes, with frequencies indicated for WT OT-II (CD45.1+CD45.2- gate) and IL-6RαKO OT-II (CD45.1-CD45.2+ gate) populations within the splenic CD4+ T cells. (B) Number of WT or IL-6RαKO (KO) OT-II cells per 1×106 splenocytes in mice that were untransfused or transfused with either fresh (Fresh) or stored HOD RBCs (Stored). (C) Number of WT and IL-6RαKO (KO) OT-II CXCR5hiPD-1hi TFH cells per 1×106 splenocytes in mice transfused with either fresh (Fresh) or stored HOD RBCs (Stored) or left untransfused. (D) WT and IL-6RαKO (KO) OT-II CXCR5hiBcl-6-1hi TFH cell numbers per 1×106 splenocytes in recipients that were untransfused or transfused with fresh (Fresh) or stored HOD RBCs (Stored). (E) Ratios of WT to IL-6RαKO OT-II cell numbers in each recipient animal. Pre-Transfer indicates the initial ratio of total WT OT-II to IL-6RαKO OT-II cells in the OT-II cell mixed population adoptively transferred into recipient animals. Untransfused, Fresh HOD RBCs, and Stored HOD RBCs indicate the ratio of total splenic WT OT-II to IL-6RαKO OT-II cell numbers in mice that were untransfused, transfused with fresh HOD RBCs, or transfused with stored HOD RBCs, respectively. CXCR5hi + PD-1hi and CXCR5hi + Bcl-6hi indicate ratios of the number of WT to IL-6RαKO CXCR5hiPD-1hi TFH OT-II and WT to IL-6RαKO CXCR5hiBcl-6hi TFH OT-II cell number respectively within each recipient transfused with stored HOD RBCs. Bars on scatter plots show median values. *P<0.05; ***P<0.001; ****P<0.0001; ns=P>0.05.