Figure 2.

Vanishing white matter disease (VWM) astrocytes proliferate actively but do not reach full maturity. (A) There are significantly higher numbers of Ki-67-positive astrocytes in the white matter of VWM patients than in controls (n = 5 patients, 3 controls; p < 0.0001; bars = SD). (B) Double stain for glial fibrillary acidic protein (GFAP) and Ki-67 shows cells with incomplete separation of the nucleus and abnormal morphology with blunt processes (patient VWM3). (C) Scratch-wound assays of control 5 (left panel) and patient VWM44 (right panel) show increased proliferation of primary cultured astrocytes in the scratch 48 hours after administration of a mechanical stress. Nuclei of the proliferating cells inside the scratch area are blue. Nuclei of the unstressed cells outside this area are red; their net number is not changed vs. controls. (D) The slope of proliferation in the scratch area; the individual net values are indicated as dots (n = 4 patients, 3 controls; p = 0.06; the lack of significance is due to the large variance of cell growth among the different VWM patients vs. controls). (E, F) GFAP-positive astrocytes from VWM patients express the intermediate filament nestin (patient VWM99) (E), and are S100β-negative (F) patient VWM3), suggesting that they are immature. (G) Astrocytes from controls show normal morphology with fine arborizations and both nuclear and cytoplasmic S100β-immunoreactivity (control 2; single S100β staining in inset). Nuclei are stained with DAPI (blue) in all panels. Original magnifications: (B, F) bars indicate 2 µm and 10 µm, respectively; original magnifications: E, G, 400×.