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. 2017 Mar 7;18(3):581. doi: 10.3390/ijms18030581

Figure 3.

Figure 3

Methyl gallate attenuates F-actin ring formation and bone-resorbing activity of mature osteoclasts. (A) BMMs were cultured with M-CSF (30 ng/mL) and RANKL (100 ng/mL) in the presence of the indicated concentrations of methyl gallate. The cells were fixed, permeabilized, and stained with phalloidin and 4′,6-diamidino-2-phenylindole (DAPI). The cells were examined under a confocal laser scanning microscope at the indicated magnification (10×); (B) Mature osteoclasts from the coculture system were seeded in 48-well plates with incubation for 6 h, in hydroxyapatite-coated plates with incubation for 24 h, or on dentin slices with incubation for 48 h with or without methyl gallate (10 µM). After that, cells attached to 48-well plates were stained with a TRAP solution, and the cells on hydroxyapatite-coated plates or dentin slices were removed and photographed under a light microscope at the indicated magnification (10×); (C) Pit areas on hydroxyapatite-coated plates or dentin slices were quantified using the Image Pro-PLUS (ver. 4.5) software, and the number of TRAP-positive NMCs (nuclei > 5) was determined. *** p < 0.001 vs. control group; (D) The mRNA expression levels of DC-, OC-STAMP, Atp6v0d2, and Cathepsin K were analyzed by quantitative real-time RT-PCR. ** p < 0.01; *** p < 0.001 vs. control group at the corresponding time point.