Limits...
Multiple sclerosis: Fas signaling in oligodendrocyte cell death.

D'Souza SD, Bonetti B, Balasingam V, Cashman NR, Barker PA, Troutt AB, Raine CS, Antel JP - J. Exp. Med. (1996)

Bottom Line: In contrast to the activity of fas in other cellular systems, dying OLs did not exhibit evidence of apoptosis, assessed morphologically and by terminal transferase-mediated d-uridine triphosphate-biotin nick-end-labeling staining for DNA fragmentation.Other stimuli such as C2-ceramide were capable of inducing rapid apoptosis in OLs.Antibodies directed at other surface molecules expressed on OLs or the M33 non-activating anti-fas monoclonal antibody did not induce cytolysis of OLs.

View Article: PubMed Central - PubMed

Affiliation: Neuroimmunology Unit, McGill University, Montreal Neurological Institute, Quebec, Canada.

ABSTRACT
Fas is a cell surface receptor that transduces cell death signals when cross-linked by agonist antibodies or by fas ligand. In this study, we examined the potential of fas to contribute to oligodendrocyte (OL) injury and demyelination as they occur in the human demyelinating disease multiple sclerosis (MS). Immunohistochemical study of central nervous system (CNS) tissue from MS subjects demonstrated elevated fas expression on OLs in chronic active and chronic silent MS lesions compared with OLs in control tissue from subjects with or without other neurologic diseases. In such lesions, microglia and infiltrating lymphocytes displayed intense immunoreactivity to fas ligand. In dissociated glial cell cultures prepared from human adult CNS tissue, fas expression was restricted to OLs. Fas ligation with the anti-fas monoclonal antibody M3 or with the fas-ligand induced rapid OL cell membrane lysis, assessed by LDH release and trypan blue uptake and subsequent cell death. In contrast to the activity of fas in other cellular systems, dying OLs did not exhibit evidence of apoptosis, assessed morphologically and by terminal transferase-mediated d-uridine triphosphate-biotin nick-end-labeling staining for DNA fragmentation. Other stimuli such as C2-ceramide were capable of inducing rapid apoptosis in OLs. Antibodies directed at other surface molecules expressed on OLs or the M33 non-activating anti-fas monoclonal antibody did not induce cytolysis of OLs. Our results suggest that fas-mediated signaling might contribute in a novel cytolytic manner to immune-mediated OL injury in MS.

Show MeSH

Related in: MedlinePlus

Expression of fas and related molecules in normal and MS CNS tissue. (a) Normal human white matter reacted with M3 anti-fas mAb. Note  the faintly positive cells (brown) that have the morphology of oligodendrocytes. Frozen section: DAB-reacted and counterstained with hematoxylin. (b)  Normal human white matter immunoreactive for fasL. Note the faint positivity on the fine processes of the ramified microglial cells. Frozen section:  DAB and hematoxylin. (c) White matter adjacent to a chronic active MS lesion shows numerous fas-positive interfascicular oligodendrocytes (brown).  DAB and hematoxylin. (d ) Detail of fas-positive oligodendrocytes from the edge of a chronic silent MS lesion. Note the typical bipolar outline of the  cells. DAB and hematoxylin. (e) Interfascicular oligodendrocytes in white matter adjacent to a chronic silent MS lesion show positive immunoreactivity  for Leu-7. Similar, but less intense staining was obtained with anti-CNPase antiserum. DAB and hematoxylin. (  f  ) Double-staining of interfascicular oligodendrocytes from white matter adjacent to a chronic silent lesion with M3 and Leu-7 (Brown and blue, respectively). DAB and NBT/BCIP. No counterstain. (  g) The edge of a chronic active MS lesion shows fas-positive infiltrating cells around a blood vessel. DAB-immunoreacted, no counterstain. (h)  Microglial cells at the periphery of a chronic silent MS lesion display intense immunoreactivity for fasL. DAB and hematoxylin. Original magnifications:  (A, D, F, G) ×750; (B, C, E, H) ×300.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2196365&req=5

Figure 1: Expression of fas and related molecules in normal and MS CNS tissue. (a) Normal human white matter reacted with M3 anti-fas mAb. Note the faintly positive cells (brown) that have the morphology of oligodendrocytes. Frozen section: DAB-reacted and counterstained with hematoxylin. (b) Normal human white matter immunoreactive for fasL. Note the faint positivity on the fine processes of the ramified microglial cells. Frozen section: DAB and hematoxylin. (c) White matter adjacent to a chronic active MS lesion shows numerous fas-positive interfascicular oligodendrocytes (brown). DAB and hematoxylin. (d ) Detail of fas-positive oligodendrocytes from the edge of a chronic silent MS lesion. Note the typical bipolar outline of the cells. DAB and hematoxylin. (e) Interfascicular oligodendrocytes in white matter adjacent to a chronic silent MS lesion show positive immunoreactivity for Leu-7. Similar, but less intense staining was obtained with anti-CNPase antiserum. DAB and hematoxylin. (  f  ) Double-staining of interfascicular oligodendrocytes from white matter adjacent to a chronic silent lesion with M3 and Leu-7 (Brown and blue, respectively). DAB and NBT/BCIP. No counterstain. (  g) The edge of a chronic active MS lesion shows fas-positive infiltrating cells around a blood vessel. DAB-immunoreacted, no counterstain. (h) Microglial cells at the periphery of a chronic silent MS lesion display intense immunoreactivity for fasL. DAB and hematoxylin. Original magnifications: (A, D, F, G) ×750; (B, C, E, H) ×300.

Mentions: In normal CNS white matter, faint fas immunoreactivity was detected on scattered glial cells by immunohistochemistry using the M3 mAb. This occurred on cells with small, round nuclei, a thin rim of cytoplasm, and one or two tenous processes (Fig. 1 a). Such elements were identified as OLs by their morphologic phenotype and positive staining for Leu-7 and CNPase in serial sections. Fas was also constitutively expressed on endothelial cells of small blood vessels. Other glial cells and neurons were invariably fas-negative. Immunostaining for fasL on normal CNS tissue revealed low-level constitutive reactivity on microglial cells (Fig. 1 b). The specificity of the fasL reactivity was confirmed by peptide preabsorption, the latter resulting in a lack of fasL immunoreactivity. In tissue from all cases of MS, fas reactivity was prominent on OLs along the margin of lesions and in adjacent white matter (Fig. 1, c and d). These same cells also stained positively for CNPase and Leu-7 in serial sections (Fig. 1 e), although the pattern of staining was different with Leu-7 and CNPase staining the cell body and M3, the cell membrane, and its fine processes. Definitive identification of these cells as faspositive oligodendrocytes was confirmed by double-staining with M3 and Leu-7 antibodies (Fig. 1 f   ). Fas IR on OLs in MS lesions was confirmed using another anti-fas mAb, UB2 (Immunotech Inc., Westbrook, ME), and similar results were obtained (data not shown). Apart from endothelial cells and infiltrating lymphocytes (Fig. 1 g), no other cell type showed fas reactivity. Staining for fas ligand in MS lesions (Fig. 1 h) revealed intense positivity on microglia and scattered infiltrating lymphocytes but not on OLs or astrocytes. In sections from OND cases, immunostaining for fas and fasL was comparable to the normal controls.


Multiple sclerosis: Fas signaling in oligodendrocyte cell death.

D'Souza SD, Bonetti B, Balasingam V, Cashman NR, Barker PA, Troutt AB, Raine CS, Antel JP - J. Exp. Med. (1996)

Expression of fas and related molecules in normal and MS CNS tissue. (a) Normal human white matter reacted with M3 anti-fas mAb. Note  the faintly positive cells (brown) that have the morphology of oligodendrocytes. Frozen section: DAB-reacted and counterstained with hematoxylin. (b)  Normal human white matter immunoreactive for fasL. Note the faint positivity on the fine processes of the ramified microglial cells. Frozen section:  DAB and hematoxylin. (c) White matter adjacent to a chronic active MS lesion shows numerous fas-positive interfascicular oligodendrocytes (brown).  DAB and hematoxylin. (d ) Detail of fas-positive oligodendrocytes from the edge of a chronic silent MS lesion. Note the typical bipolar outline of the  cells. DAB and hematoxylin. (e) Interfascicular oligodendrocytes in white matter adjacent to a chronic silent MS lesion show positive immunoreactivity  for Leu-7. Similar, but less intense staining was obtained with anti-CNPase antiserum. DAB and hematoxylin. (  f  ) Double-staining of interfascicular oligodendrocytes from white matter adjacent to a chronic silent lesion with M3 and Leu-7 (Brown and blue, respectively). DAB and NBT/BCIP. No counterstain. (  g) The edge of a chronic active MS lesion shows fas-positive infiltrating cells around a blood vessel. DAB-immunoreacted, no counterstain. (h)  Microglial cells at the periphery of a chronic silent MS lesion display intense immunoreactivity for fasL. DAB and hematoxylin. Original magnifications:  (A, D, F, G) ×750; (B, C, E, H) ×300.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2196365&req=5

Figure 1: Expression of fas and related molecules in normal and MS CNS tissue. (a) Normal human white matter reacted with M3 anti-fas mAb. Note the faintly positive cells (brown) that have the morphology of oligodendrocytes. Frozen section: DAB-reacted and counterstained with hematoxylin. (b) Normal human white matter immunoreactive for fasL. Note the faint positivity on the fine processes of the ramified microglial cells. Frozen section: DAB and hematoxylin. (c) White matter adjacent to a chronic active MS lesion shows numerous fas-positive interfascicular oligodendrocytes (brown). DAB and hematoxylin. (d ) Detail of fas-positive oligodendrocytes from the edge of a chronic silent MS lesion. Note the typical bipolar outline of the cells. DAB and hematoxylin. (e) Interfascicular oligodendrocytes in white matter adjacent to a chronic silent MS lesion show positive immunoreactivity for Leu-7. Similar, but less intense staining was obtained with anti-CNPase antiserum. DAB and hematoxylin. (  f  ) Double-staining of interfascicular oligodendrocytes from white matter adjacent to a chronic silent lesion with M3 and Leu-7 (Brown and blue, respectively). DAB and NBT/BCIP. No counterstain. (  g) The edge of a chronic active MS lesion shows fas-positive infiltrating cells around a blood vessel. DAB-immunoreacted, no counterstain. (h) Microglial cells at the periphery of a chronic silent MS lesion display intense immunoreactivity for fasL. DAB and hematoxylin. Original magnifications: (A, D, F, G) ×750; (B, C, E, H) ×300.
Mentions: In normal CNS white matter, faint fas immunoreactivity was detected on scattered glial cells by immunohistochemistry using the M3 mAb. This occurred on cells with small, round nuclei, a thin rim of cytoplasm, and one or two tenous processes (Fig. 1 a). Such elements were identified as OLs by their morphologic phenotype and positive staining for Leu-7 and CNPase in serial sections. Fas was also constitutively expressed on endothelial cells of small blood vessels. Other glial cells and neurons were invariably fas-negative. Immunostaining for fasL on normal CNS tissue revealed low-level constitutive reactivity on microglial cells (Fig. 1 b). The specificity of the fasL reactivity was confirmed by peptide preabsorption, the latter resulting in a lack of fasL immunoreactivity. In tissue from all cases of MS, fas reactivity was prominent on OLs along the margin of lesions and in adjacent white matter (Fig. 1, c and d). These same cells also stained positively for CNPase and Leu-7 in serial sections (Fig. 1 e), although the pattern of staining was different with Leu-7 and CNPase staining the cell body and M3, the cell membrane, and its fine processes. Definitive identification of these cells as faspositive oligodendrocytes was confirmed by double-staining with M3 and Leu-7 antibodies (Fig. 1 f   ). Fas IR on OLs in MS lesions was confirmed using another anti-fas mAb, UB2 (Immunotech Inc., Westbrook, ME), and similar results were obtained (data not shown). Apart from endothelial cells and infiltrating lymphocytes (Fig. 1 g), no other cell type showed fas reactivity. Staining for fas ligand in MS lesions (Fig. 1 h) revealed intense positivity on microglia and scattered infiltrating lymphocytes but not on OLs or astrocytes. In sections from OND cases, immunostaining for fas and fasL was comparable to the normal controls.

Bottom Line: In contrast to the activity of fas in other cellular systems, dying OLs did not exhibit evidence of apoptosis, assessed morphologically and by terminal transferase-mediated d-uridine triphosphate-biotin nick-end-labeling staining for DNA fragmentation.Other stimuli such as C2-ceramide were capable of inducing rapid apoptosis in OLs.Antibodies directed at other surface molecules expressed on OLs or the M33 non-activating anti-fas monoclonal antibody did not induce cytolysis of OLs.

View Article: PubMed Central - PubMed

Affiliation: Neuroimmunology Unit, McGill University, Montreal Neurological Institute, Quebec, Canada.

ABSTRACT
Fas is a cell surface receptor that transduces cell death signals when cross-linked by agonist antibodies or by fas ligand. In this study, we examined the potential of fas to contribute to oligodendrocyte (OL) injury and demyelination as they occur in the human demyelinating disease multiple sclerosis (MS). Immunohistochemical study of central nervous system (CNS) tissue from MS subjects demonstrated elevated fas expression on OLs in chronic active and chronic silent MS lesions compared with OLs in control tissue from subjects with or without other neurologic diseases. In such lesions, microglia and infiltrating lymphocytes displayed intense immunoreactivity to fas ligand. In dissociated glial cell cultures prepared from human adult CNS tissue, fas expression was restricted to OLs. Fas ligation with the anti-fas monoclonal antibody M3 or with the fas-ligand induced rapid OL cell membrane lysis, assessed by LDH release and trypan blue uptake and subsequent cell death. In contrast to the activity of fas in other cellular systems, dying OLs did not exhibit evidence of apoptosis, assessed morphologically and by terminal transferase-mediated d-uridine triphosphate-biotin nick-end-labeling staining for DNA fragmentation. Other stimuli such as C2-ceramide were capable of inducing rapid apoptosis in OLs. Antibodies directed at other surface molecules expressed on OLs or the M33 non-activating anti-fas monoclonal antibody did not induce cytolysis of OLs. Our results suggest that fas-mediated signaling might contribute in a novel cytolytic manner to immune-mediated OL injury in MS.

Show MeSH
Related in: MedlinePlus