Figure 7: Critical-sized calvarial bone-defect repair in overiectomy-induced osteoporotic (OVX) mice.

(a) Cell-free PLLA scaffolds with physically coated HP/mi-R26a or NC polyplexes (miR-26a-bolus or NC-bolus), (b) cell-free PLLA scaffolds with immobilized PLGA 6.5-K microspheres loaded with HP/miR-26a or NC polyplexes (miR-26a-HP-short or NC-HP-short) and (c) cell-free PLLA scaffolds with immobilized PLGA 64-K microspheres loaded with HP/miR-26a or NC polyplexes (miR-26a-HP-long or NC-HP-long) were implanted in 5-mm critical-sized calvarial bone defects in OVX mice. MicroCT images (left) and H&E staining (middle and right) of calvarial defects were recorded after implantation for 2 months. Scale bars, 5 mm in microCT images, 2.0 mm in H&E images in the middle, 200 μm in the high-mag H&E images on the right. (d) Quantitative analysis showing the new bone volumes and new bone mineral densities in the above-described groups. (e) Osteoblast surface/bone surface ratios and osteoblast number/bone perimeter ratios in OVX mice under the above-described groups. (f) Immunofluorescence staining for OCN (green) and DAPI staining for nuclei (blue) in histological sections of the healing calvarial bone defects in OVX mice in the above-described groups, where the scale bars, 500 μm (left), 200 μm (right). (g) New bone-formation front in the calvarial bone defect was sequentially labelled with fluorescent calcein and xylenol in OVX mice in the six groups. Representative fluorescent micrographs show the xylenol (red) and calcein (green) bands, where the scale bars, 2 mm. (h) The Oc.S/BS and Oc.N/B.Pm were determined using tartrate-resistant acid phosphatase (TRAP) staining. *P<0.05; **P<0.01; #P<0.05; ##P<0.01. All experiments were performed in triplicate. n=5 per group. Data are mean±s.d.