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. 2016 Mar 7;7:10872. doi: 10.1038/ncomms10872

Figure 3. Reduced bone formation in OC-miR-214-3p mice.

Figure 3

(a) A schematic diagram illustrating the experimental design. (b,c) Schematic diagrams of the development strategy of the OC-miR-214-3p mice. The ROSA26-miR-214-3p knock-in mice containing the miR-214-3p knock-in allele were generated, and then crossed with the Ctsk-cre transgenic mice to obtain the OC-miR-214-3p mice. One unit included the mmu-miR-214-3p stem–loop, a 200-bp 5′ flanking sequence and a 200-bp 3′ flanking sequence. (d) Real-time PCR analysis of pri-miR-214-3p (left), pre-miR-214-3p (middle left) and miR-214-3p (middle right) levels in osteoclasts (OCs) versus non-osteoclasts (non-OCs) and serum exosomal miR-214-3p level (right) from WT and OC-miR-214-3p mice. The OCs and non-OCs were the Ctsk+ and Ctsk− cells isolated from the bone marrow cells by magnetic-activated cell sorting, respectively. (e) Real-time PCR analysis of the mRNA levels of bone formation marker genes (Alp, Opn, BSP and Bglap) in bone specimens from WT and OC-miR-214-3p mice. (f) Representative micro-CT images of the distal femur from WT and OC-miR-214-3p mice. (g) The values of micro-CT parameters (BMD, BV/TV, Tb.Th and Tb.N) at the distal femur metaphysis from WT and OC-miR-214-3p mice. (h) Representative images of new bone formation assessed by double labelling with calcein green and xylenol orange at the cortical bone (CB) and trabecular bone (TB) of distal femur from WT and OC-miR-214-3p mice. Scale bars, 10 μm. (i) The values of bone histomorphometry parameters (MAR, BFR/BS, Ob.S/BS, Ob.N/B.Pm, Oc.S/BS and Oc.N/B.Pm) at the distal femur metaphysis from WT and OC-miR-214-3p mice. (j) Representative images of osteoid formation indicated by Masson's trichrome staining at the distal femur metaphysis of WT and OC-miR-214-3p mice. Scale bars, 20 μm. (k) The values of osteoid-related parameters (Osteoid surface/bone surface and Osteoid thickness) at the distal femur metaphysis from WT and OC-miR-214-3p mice. The n value for each group is indicated at the bottom of each histogram. All data are the mean±s.d. *P<0.05 versus WT. **P<0.01 versus WT. Student's t-test was performed.

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