Fig. 2: Staining of osteoblasts with or without mechanical stimulation.
From: Bone mechano-response is driven by locomotion transitions during vertebrate evolution
Osteoblasts were cultured in osteoinductive media with or without mechanical stimulation. The cells under the static condition started the osteoblast differentiation after 1 week and differentiated into mature osteoblasts after 4 weeks. The cells exposed to mechanical stimulation differentiated into osteoblasts after 2 weeks, and mineral deposition occurred after 4 weeks. ALP staining revealed that the mechanical stimulation enhanced osteoblast differentiation. Scale bar for ALP images = 500 μm. Immunofluorescent staining shows that the osteoblasts expressed the target proteins. Merged images of actin (rhodamine phalloidin, red), nuclei (Hoechst, blue) and the targeted proteins (green) are shown. The red, green, and blue color-separated images are available in Fig. S1–5. Four proteins (RSPO1, annexin A1, osteonectin, and TGF-β3) were expressed from early to late stages of osteoblast differentiation. Two proteins (RSPO1 and annexin A1) were expressed under both the static and mechanical stimulation conditions. Five proteins (BSP, MATN3, osteopontin, fetuin A, and fetuin B) were observed as dot-like structures in the early stage of osteoblast differentiation and bundle-like structures or aggregated structures in the late stage of osteoblast differentiation. Scale bar for immunofluorescent images = 100 μm.
