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. 2014 Mar 13;23(18):2156–2169. doi: 10.1089/scd.2013.0344

FIG. 5.

FIG. 5.

Transmission electron microscopy images of bone nodules formed in osteogenically induced MSCs (A–D) and iPSCs (E–H) cultures. Cells are indicated as “C” in the panels. (A) Cross-section through the MSCs on day 10. Calcium phosphate-containing vesicles (black spot areas) were visible around the cells and in the extracellular spaces, which contained fibrous ECM (asterisks). Scale bar: 5 μm. (B) Magnified image of the square in (A). Micrograph showing mineral aggregates (asterisk) immediately outside the cell. Scale bar: 100 nm. Inset: the selected area electron diffraction (SAED) pattern of the mineral aggregate was similar to that of hydroxyapatite. (C) Micrograph of a cross-section through the MSCs on day 30. Electron-dense vesicles (dense black spots) were evident immediately outside cells (arrowheads), within membrane invaginations (black arrow), and within cells (white arrow). Fibrous ECM was visible (asterisks). Scale bar: 5 μm. (D) Magnified image of the dotted square in (C). Micrograph showing dense mineral (asterisk) immediately outside the cell membrane. Scale bar: 500 nm. Inset: SAED analysis of the dense areas was indicative of hydroxyapatite. (E) Cross-section through the iPSCs on day 10. An electron-sparse structure (asterisks) surrounded the cells. Scale bar: 2 μm. (F) Magnified image of the square in (E). Electron-dense needle-like minerals are scattered (asterisk). Scale bar: 50 nm. Inset: SAED analysis of the scattered mineral structure indicated a lack of a textured crystalline diffraction pattern, which suggests that the structure is relatively amorphous and that mineralization has been initiated. (G) Cross-section through the bone nodule portion of iPSCs on day 30. Heavily electron-dense vesicles (asterisks) were evident in the ECM spaces. A vesicle, which appeared to be dispersing its mineral contents (arrowheads), was also visible. Fibrous structures were not evident in the extracellular spaces. Scale bar: 5 μm. (H) Magnification of the dotted square in (G). The SAED pattern of an electron-dense vesicle (asterisk) was identical to that of typical hydroxyapatite (inset). Scale bar: 500 nm.