Fig. 7

ITPR3-mediated NF-ĸB/CD44 signaling in BCa cells. a Immunofluorescence indicating the nuclear translocation of NF-κB (P65) induced by TNFα (10 ng/mL) (column 1 vs. column 2), which can be inhibited by ITPR3 knockdown (column 2 vs. column 3). Bar: 50 μm, arrows: NF-κB nuclear translocated cells. b GSEA results showed that “HALLMARK_TNFA_SIGNALING_VIA_NFKB” was upregulated and responded to high ITPR3 expression. c The expression level of NF-κB in nuclear and cytosolic fractions was analyzed by western blotting. Histone H3 and β-actin were used as the loading controls for the nucleus and cytosol, respectively. d We examined the NF-κB pathway-related proteins P-NF-κB, NF-κB, P-IKBa, and IKBα by western blotting with specific antibodies in 5637 and 253 J cells transfected with ITPR3 shRNA or shCon. β-actin served as an internal control. e CD44 expression was analyzed by immunofluorescence in 5637 and 253 J cells transfected with ITPR3 shRNA or shCon. Scale bars, 100 μm. f western blot analysis of ITPR3, CD44, NF-κB, and P- NF-κB in 5637 and 253 J cells treated with TNFα for 24 h after transfection with ITPR3 shRNA or shCon. β-actin was used as a loading control