Fig 1: RANK promoted the metastasis of CRC cells in vitro and in vivo.a Protein expressions of RANK were detected by western blot analysis in seven CRC cell lines. b, c RANK stable overexpression or knockdown efficiency was confirmed by western blotting in CRC cells. d, e RANK overexpression promoted the migration and invasion of SW480 and Caco2 cells. And the addition of 100 ng/ml RANKL moderately increased the migration and invasion of RANK-overexpressing SW480 and Caco2 cells. Scales bars = 100 μm. f Knockdown of RANK inhibited the migration and invasion of HT29 cells. Scales bars = 100 μm. g Bioluminescent images of representative mice at week 15 after implantation. h The total tumor burden is quantified as total photons measured by bioluminescent technology. i Livers were excised for ex vivo bioluminescent imaging. j Representative results for staining of metastatic nodules (black arrows) in the livers. Scales bars = 500 μm (40×), 200 μm (100×). k The statistical analyses of metastatic nodules. Data are mean ± SD (n = 3; n = 6 mice/group for in vivo experiments). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.
Fig 2: Hematoxylin-eosin and RANKL stains on Ewing’ sarcoma.(7a) Uniform small round cells were observed. (7b) No RANKL-positive cells were detected.
Fig 3: Evaluation of RANKL expression in articular cartilage by immunoelectron microscopy.The localization of 10 nm colloidal gold particles labeled RANKL antibody are observed as electron–dense punctate. (A and C) Lower magnification micrograph of deep layer chondrocyte and surrounding matrix. (B) Insets of A showing localization of immunogold at pericellular matrix (a, b), cytoplasm and cell membrane (c, d, e), and territorial matrix surrounding chondron (f, g, h, i). Scale bars = 500 nm. (D) Inset of C showing extracellular vesicles scattered randomly with the territorial matrix without labeling by colloidal gold, indicating these vesicles do not contain RANKL. Ch, chondrocyte; N, nucleus; EVs, extracellular vesicles.
Fig 4: Cytokines induced gene expression in chondrocytes.(A) RANKL expression and the RANKL/OPG ratio were up–regulated after 24 hours of stimulation with LT–α or TNF–α, but not after 12 hours of stimulation. (B) RANKL expression and angiogenic factors were up–regulated in a concentration–dependent manner after 24 hours of cytokine stimulation (n = 6 per experimental group). * / § p < 0.05, ** / §§ p < 0.01, *** / §§§ p < 0.001.
Fig 5: Proposed model for the potential function of TRPS1 on the selective mechanism of MPA in EC and BC cells.TRPS1 functioned as a cosuppressor recruited by PR combining with HDAC2 to enhance the sensitivity of cells to MPA via deacetylating RANKL in EC cells, while in different cellular contexts of breast cancer, TRPS1 served as a coactivator and the complex of PR/TRPS1/HDAC2 was disassociated, resulting in the acetylation of RANKL and facilitating the cancer-promoting effect of MPA in BC. Denosumab, as a monoclonal antibody directed to RANKL, could effectively enhance the tumor suppressor effect of MPA in EC, while inhibiting the growth of breast cancer.
Supplier Page from Abcam for Anti-RANKL antibody