14800-DvRappard

95 Donor macrophages and remyelination in metachromatic leukodystrophy 6 Figure 2. Donor cells reach the brain of transplanted MLD patients. (A, patient 5) Stain with Klüver (blue dye for myelin) and periodic acid Schiff (PAS, pink, stain for sulfatides in macrophages) of the cerebral periventricular white matter shows loss of myelin and abundance of cells loaded with PAS-positive granular material. (B, patient 1) Hematoxylin&Eosin stain of the frontal subcortical white matter shows presence of macrophages with intense eosinophilic cytoplasm (arrows) next to macrophages loaded with clearer granular material. (C, patient 1) A Klüver-PAS stain of the same region confirms the presence of a double population of macrophages, more (open arrows) and less (closed arrows) intensely PAS-positive. (D, patient 2) Toluidine stain of the parietal white matter reveals that only a subset of macrophages is metachromatic (purple, i.e. loaded with sulfatides), the remaining being orthochromatic (brown). (E, patient 1) Metabolic competence of a subset of macrophages is confirmed by their ability to digest sulfatides to cholesterol, as shown in this Oil Red-O stain for neutral fats. (F, patient 1) FISH against the X and Y chromosomes confirms cells of both sexes in the brain of this transplanted child. Looking at oligodendrocytes, we found that numbers of both oligodendrocyte precursors and mature myelin-forming oligodendrocytes was significantly higher in transplanted than untreated patients (figure 4). The amount of total myelin as assessed by PLP immunopositivity was unchanged, indicating that in the white matter of treated patients, oligodendrocytes and their precursors may survive despite a similar degree of myelin loss.