T cells are not the cause of the AA-like phenotype in ARE-del mice. (A) Total RNA was extracted from tissues. Equal amounts of RNA were reverse transcribed into mRNA. IFN-γ mRNA levels were detected by using real-time polymerase chain reaction. Data were normalized against glyceraldehyde-3-phosphate dehydrogenase mRNA levels and calculated relative to IFN-γ expression levels in WT tissue set to 1 (n = 3). (B) Serum from animals was analyzed by using a cytometric bead array to determine IFN-γ levels (n = 7). (C) Single-cell suspensions of BM cells were prepared and counted, and total cell numbers are shown (n = 7). (D) Sternums from 6 WT and 6 ARE-del mice were fixed and stained with hematoxylin and eosin. Depicted here are representative photographs from 1 set of animals. (E) Seven-color flow cytometry analyses were performed on total BM cells. T cells were gated on the live NKp46^–CD11b^–B220^–Gr1^–CD3⁺ population. The bar graphs show both the total cell number and percentage of live cells (n = 4). (F) T cells were selected from BM. The function of T cells was determined by the intensity of cytokine responses against TCR agonists (anti-CD3/anti-CD28 antibodies). Cytokine levels in the medium were measured 6 hours after stimulation by using a cytometric bead array. The bar graphs displays the concentration of each cytokine measured (n = 4). All experiments were performed at least 3 times. Results are expressed as mean ± standard deviation (SD). *P < .05; **P < .01; ***P < .001; ****P < .0001. SPL, spleen.