Fig. 8

Ultrastructural identification of intracytoplasmic OPN in reactive astrocytes. a–c Immunoelectron microscopic (EM) images in the lesioned striatum at 28 days post-lesion, showing that degenerated neurites delineated by DAB grains are in close proximity to, or internalized by, reactive astrocytes (As, cyan) showing extensive cell body hypertrophy and cytoplasmic processes. The boxed areas in a are enlarged in b and c. Note that intracytoplasmic OPN-positive profiles are similar in shape to those outside the astrocytes, and occasionally contain small and highly electron-dense mitochondria (m in b). The EM image shows representative data for immunoelectron staining of three sections from three rats. As astrocytes, cyan; f glial filaments; M microglia/macrophage, magenta; N nucleus. d–g Triple labeling with OPN, GFAP, and LAMP1 in control striatum (d) and in the lesioned striatum at 28 days post-lesion (e–g). e–g Confocal z-stack of images (e), an image including only intracytoplasmic OPN and LAMP1 within reactive astrocytes from e (f) and an image displaying the LAMP1 signals within reactive astrocytes (g) showing prominent LAMP1 staining with a typical punctate pattern, is localized within reactive astrocytes that contained OPN puncta. h The three-dimensional rendering of reactive astrocytes that contain numerous lysosomes and OPN puncta. Cell nuclei appear blue after DAPI staining. Scale bars = 2 μm for a; 0.5 μm for b, c; 10 μm for d–h