Figure 2

IL-6 and IL-17F/A levels increase and Stat3 is activated in hIL-1α cTg mice. PolyI-PolyC was injected into eight-week-old Ctl or cTg mice. Three weeks later, peripheral blood was collected and ankle joints removed. Levels of cytokines indicated on the y-axis in peripheral blood sera were assessed by ELISA (a). Data represent mean levels of indicated cytokines ± SD (n = 3 for Ctl; n = 4 for cTg mice; ^**P < 0.01, NS not significant). Ankle joint specimens from Ctl or cTg mice were subjected to hematoxylin and eosin (HE) and immunofluorescence staining with an antibody specific for phosphorylated Stat3 (pStat3) Arrowheads indicate pStat3-psotive cells. (b) Nuclei were visualized by DAPI. Bar, 100 µm. (c) The absolute number of splenocytes in Ctl or cTg mice was determined three weeks after PolyI-PolyC injection. Data represents mean splenocyte number ± SD (n = 6 for Ctl; n = 4 for cTg mice; ^*P < 0.05). (d–f) Flow cytometric analysis was undertaken to detect CD11b, Gr1, CD3, B220, CD4, TCRβ, TCRγδ and IL-17 in cells isolated from spleen (d), lymph nodes (LN) (e) and joints (f) from Ctl (upper panels) or cTg (lower panels) mice. TCRβ⁺TCRγδ⁻CD4⁺IL-17⁺ and TCRβ⁻TCRγδ⁺IL-17⁺ cells are shown as IL-17⁺CD4⁺ and IL-17⁺TCRγδ⁺ cells, respectively. Data represent mean frequency (%) of each fraction ± SD (n = 5 for Ctl; n = 4 for cTg mice; ^*P < 0.05, ^**P < 0.01, ^***P < 0.001, NS not significant). (g) CD11b− (Mac), Gr1− (Neu), CD3− (T cell) or B220-positive (B cell), or CD45-negative cells (CD45−) were sorted from Ctl or cTg splenocytes, and human IL-1α (hIL-1α) expression was determined by realtime PCR. Data represent mean indicated gene expression ± SD (n = 3 for Ctl; n = 3 for cTg mice; ^***P < 0.001).