miR-134-5p expression is decreased during osteoclast differentiation.A, BMMs were cultured in the presence of 30 ng/ml M-CSF and 50 mg/ml RANKL for 7 days and were identified under a light microscope. Scale bars: 20×: 100 μm; 40×: 50 μm. B and C, TRAP staining was performed to analyze osteoclast activity. The number of TRAP+ multinuclear osteoclasts was counted. The scale bars represent 20×: 100 μm; 40×: 50 μm. D and E, BMMs were induced to differentiate into osteoclasts and stained for actin ring formation using phalloidin. The number of F-actin rings per field was counted. The scale bars represent 100 μm. F, the mRNA expression levels of TRAP, CTSK, and NFATc1 were analyzed by quantitative RT-PCR at 1, 3, and 7 days. β-Actin was used as an internal reference gene. G, the expression level of miR-134-5p was analyzed by quantitative RT-PCR after induction for 1, 3, and 7 days. U6 was used as an internal reference gene. All the data were expressed as the means ± SDs. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001. BMMs, bone marrow macrophages.