Figure 4
Paraspeckles appear predominantly in spinal motor neurons in ALS. A. RNA-FISH using DIG-labeled NEAT1_2 probe indicates that NEAT1_2 lncRNA often appears in the nuclei of human motor neurons in sporadic ALS cases. Right panel shows the profile image of fluorescence intensity of NEAT1_2 lncRNA and TDP-43 along the arrow in the nucleus (ZEN software, Carl Zeiss). Most NEAT1_2 signals overlapped with parts of aggregated TDP-43 in the nucleus (arrowheads). Dotted lines represent the outline of the nucleus. B. No NEAT1_2 expression is detected in most motor neurons in control cases. Dotted lines represent the outline of the nucleus. C. A 40-fold magnified view of a spinal motor neuron in an ALS case is shown with DAB-hematoxylin counterstain using anti-PSP1 antibody. The arrowhead indicates the nucleolus. Arrows indicate PSP1 aggregates in the nucleus. D, E. RNA-FISH using DIG- or FITC-labeled NEAT1_2 probe in the nuclei of ALS motor neurons. PSF ((D), ALS-Pt C), PSP1 (upper in (E), ALS-Pt D) and p54nrb (lower in (E), ALS-Pt B) overlap with NEAT1_2 foci. Observation at 647 nm wavelength was performed to rule out autofluorescent signals. Right panels display images on orthogonal sections (ZEN software). Asterisks indicate lipofuscin in the cytoplasm. Dotted line: outline of the nucleus. Scale bars, 10 μm.