Fig. 8

Expression of key ferroptosis markers in SPMS. A Double immunofluorescence labeling of ferritin (FN) in macrophages (CD68/FN) (Ai); and oligodendrocytes (TPPP/FN) (Aii). All immunofluorescence sections are stained with nuclear DAPI stain; L = lesion. Note the clear expression of ferritin in macrophages lined up along the lesion rim (arrows) (Ai). In contrast, fewer oligodendrocytes showed double labeling of TPPP and FN (Aii). FN labeling surrounds some TPPP⁺ cell bodies and may represent FN in cell processes (arrows). Inset shows double labeling of TPPP and FN (arrow) in another section. Aiii Shows double labeling of ferritin and NCOA4 (FN/NCOA4). Note that cells showing strong FN labeling express little or no NCOA4 (1), and cells that show no ferritin strongly express NCOA4 (2), while cells that express some NCOA4 express lower levels of FN (3). B Panels showing double labeling of CD68 and NCOA4 (i-iii) and TPPP and NCOA4 (iv-vi) in Controls (i and iv), NAWM (ii and v) and MAIAPD (iii and vi). All panels stained with DAPI. Bi–Biii In CD68+ and NCOA4 staining, note the absence of CD68⁺ macrophages and lack of NCOA4 in Control tissue. Bii In NAWM only about 10% of the CD68⁺ macrophages express NCOA4 (arrow). Biii In contrast, CD68⁺ macrophages lining the rim of MAIAPD lesions were double labeled with NCOA4. The hatched lines demarcate the lesion rim. Biv–Bvi In TPPP⁺ and NCOA4 staining, in control samples, a small number TPPP⁺ oligodendrocytes express NCOA4 (arrows). Bv The numbers and staining intensity of NCOA4 increased in NAWM (arrows). Bvi In contrast, NCOA4 expression is markedly increased in TPPP⁺ oligodendrocytes along the lesion rim located between the hatched lines. C Panels showing CD68⁺ macrophages (Ci) and TPPP⁺ oligodendrocytes (Cii) double labeled with 4-HNE (arrows). Note that the 4-HNE staining is stronger in macrophages as compared to the sparce labeling in oligodendrocytes. Scale bar in A and B: 30 µm; C and Inset: 10 µm