Figure 4.

IL-9 blockade abrogates bone loss via modulating Th9 and Th17 cells: (A) Upon sacrifice, BMCs were harvested from all the groups and cultured in the media supplemented with M-CSF (30 ng/mL) and RANKL (60 ng/mL) and after 4 d of incubation TRAP staining was performed. (B) Number of TRAP positive cells with >3nuclei. (C-E) Bar graph representing relative expression of cathepsin K, RANKL, OPG genes in representative groups. (F) Bar graph representing RANKL/OPG ratio. (G-H) ELISA data representing the level of bone biochemical biomarkers viz. CTX-1 and PINP in the sera of respective groups. (I) Contour plots representing CD4⁺IL-9⁺IL-17⁺ Th cells, CD4⁺IL-9⁺IL-17^- Th9 cells, CD4⁺IL-9^-IL-17⁺ Th17 cells in BM. (J-L) Individual plot representing CD4⁺IL-9⁺IL-17^- Th9 cells, CD4⁺IL-9^-IL-17⁺ Th17 cells, and CD4⁺IL-9⁺IL-17⁺ Th17 cells in BM. (M-Q) Individual plots representing IL-17a, Rorγt, Foxo-1 gene expression, IRF4, and Ccl 20 gene expression in BM of representative groups. Data are expressed as mean ± SEM. Data were analyzed by unpaired Student t test and analyzed by one-way ANOVA. ^*p ≤ 0.05, ^**p ≤ 0.01, ^***p ≤ 0.001, ^****p ≤ 0.0001) compared with the indicated group. Abbreviation: IRF4, interferon regulatory factor 4.