Figure 1. RA-induced tongue malformation at E14.5 and E15.5, and morphology of the genioglossus at E18.5.

(a): HE staining of coronal sections of the tongue from E14.5 mouse fetus. The tongue (T) was at higher positions and bilateral palatal shelves (PS) were vertically positioned that formed a cleft in the +RA fetus (ii) versus control fetus (i). The bilateral palatal shelves (PS) in control fetus (i) already elevated above the tongue to the horizontal position and merged each other. Bar = 100 μm. (b): Myosin immunocytochemical staining of tongue muscles. Myosin expression in the tongue body (iii) and the genioglossus muscle (iv) were decreased in the +RA fetus versus control fetus. A higher magnification of the tongue intrinsic muscle from control fetuses showed multinucleated myotubes expressing high levels of myosin ((i), arrow). In +RA fetuses, myosin expression in myotubes ((iii) arrow) became weaker. In a transverse section of the genioglossus muscle, myosin was expressed at high levels in the control myotubes ((ii), arrowhead); in +RA fetuses, myosin staining was very weak. Bar = 40 μm. Upper right inserts in (i and iii) show images at lower magnifications, Bar = 100 μm. (c): Transmission electron microscopy examination of sagittal sections of the genioglossus. In control (i), myofibrils (white arrow) and sarcomeres were arranged longitudinally with integral I and A bands, and clear Z- and M-lines. In +RA (ii), the classic structures of sarcomeres of myofibrils were not found, myofilament bundles were arranged longitudinally (white arrow) and transversely (black arrow) in a single cell. Bar = 500 nm. Cont.: control mouse fetus; +RA: RA-exposed mouse fetus.PS: palatal shelves; T: tongue.