Figure 4.

Tyk2^P and IL−12R^−/− mice are not protected in murine models of lupus. (A) Experimental schematic for BM12 CD4⁺ T cell adoptive transfer model. 5.0 × 10⁶ BM12 CD4⁺ T cells were adoptively transferred into Tyk2^NP/NP, Tyk2^NP/P, or Tyk2^P/P recipient mice and autoantibodies and splenic cell populations (not shown) were assessed at indicated times. Serum ELISA analysis for: (B) anti-dsDNA IgG and (C) anti-dsDNA IgG2c autoantibodies. (D) Schematic for establishment of B cell-specific bone marrow (BM) chimeras using an 80%:20% mixture of bone marrow from μMT^−/− mice and WT or Wiskott-Aldrich knock-out (WAS^−/−) donor cells with the indicated Tyk2 or IL-12R alleles (WAS^−/−Tyk2^NP/NP, WAS^−/−Tyk2^NP/P, WAS^−/−Tyk2^P/P, WAS^−/−IL-12R^−/−), respectively. Yellow lightning bolt represents irradiation of recipient mice. See methods for additional details of experimental design. (E,F) ELISA analysis of serum at 16 week post-transplantation for: (E) anti-dsDNA IgG2c and (F) anti-smRNP IgG2c autoantibodies in indicated recipient mice. (G,H) Splenocytes were isolated at 16 week and analyzed by flow cytometry for frequency of: (G) GC and (H) Tfh cells as described in Figure 2. Small horizontal lines indicate the mean (± s.e.m.). ELISA results are displayed as absorbance at 450 nm normalized to results of a blank well and presented in arbitrary units (AU). (B,C,E,F) Statistical analysis was performed using one-way ANOVA with Tukey's multiple comparisons test. Data are representative of two independent experiments (B,C) or data combined from two independent experiments (E–H). Each symbol represents an individual biological replicate;Tyk2^NP/NP n = 3, Tyk2^NP/P n = 4,Tyk2^P/P n = 3 or PBS n = 2 (A-C); WT n = 4,Tyk2^NP/NP n = 9, Tyk2^NP/P n = 12,Tyk2^P/P n = 7 or IL-12R^−/− n = 8 (E,F); WT n = 8,Tyk2^NP/NP n = 9, Tyk2^NP/P n = 12,Tyk2^P/P n = 7 or IL-12R^−/− n = 8 (G,H).