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. 2000 Dec;157(6):1991-2002.
doi: 10.1016/S0002-9440(10)64838-9.

Intracerebral Recruitment and Maturation of Dendritic Cells in the Onset and Progression of Experimental Autoimmune Encephalomyelitis

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Free PMC article

Intracerebral Recruitment and Maturation of Dendritic Cells in the Onset and Progression of Experimental Autoimmune Encephalomyelitis

B Serafini et al. Am J Pathol. .
Free PMC article

Abstract

Dendritic cells (DCs) are thought to be key elements in the initiation and maintenance of autoimmune diseases. In this study, we sought evidence that DCs recruited to the central nervous system (CNS), a site that is primarily devoid of resident DCs, play a role in the effector phase and propagation of the immune response in experimental autoimmune encephalomyelitis (EAE). After immunization of SJL mice with proteolipid protein 139-151 peptide, process-bearing cells expressing the DC markers DEC-205 and CD11c appeared early in the spinal cord. During acute, chronic, and relapsing EAE, DEC-205(+) DCs expressing a lymphostimulatory phenotype (including the mature DC marker MIDC-8, major histocompatibility complex class II, CD40, and CD86 molecules) accumulated within the CNS inflammatory cell infiltrates. More prominent infiltration of the spinal cord parenchyma by mature DCs was observed in mice with relapsing disease. Macrophage inflammatory protein 3alpha, a chemokine active on DCs and lymphocytes, and its receptor CCR6 were up-regulated in the CNS during EAE. These findings suggest that intracerebral recruitment and maturation of DCs may be crucial in the local stimulation and maintenance of autoreactive immune responses, and that therapeutic strategies aimed at manipulating DC migration could be useful in the treatment of CNS autoimmune disorders.

Figures

Figure 1.
Figure 1.
Localization of DC markers in the CNS of normal SJL mice (A–C) and of mice immunized with PLP 139-151 in CFA before the onset of EAE clinical signs (day 10 after immunization) (D–H). Representative cerebellar (A) and thoracic spinal cord (B–H) sections are shown. CNS sections were stained for the indicated markers and visualized with DAB, as described in Materials and Methods. Control CNS: DEC-205+ process-bearing cells in the choroid plexus of the fourth ventricle (A), DEC-205+ (B) and CD11c+ (C) process-bearing cells in the spinal cord meninges. Preclinical EAE: DEC-205+ (D and E), CD11c+ (F), and MHC class II+ (H) cells with thin cytoplasmic processes are present in the subpial white matter of the ventral and lateral spinal cord. Note the ramified morphology of a DEC-205+ cell immediately beneath the pial surface (inset in E). Double staining with anti-CD11b (brown color, DAB) and anti-CD11c (red color, fuchsin) mAbs reveals that CD11c mAb recognizes intraparenchymal cells that are distinct from CD11b+ microglia (G). Original magnifications: ×500 (A, D, F, and H), ×1000 (B, C, E, and G), and ×1575 (inset).
Figure 2.
Figure 2.
Cellular composition of the immune infiltrates and localization of DC markers in the CNS of mice with acute EAE. Representative lumbar (B, D, and I) and thoracic (A, C, F, G, and H) spinal cord and cerebellar (E) sections from a SJL mouse developing acute EAE (grade 4) are shown. CNS sections were stained for the indicated markers and visualized with DAB, as described in Materials and Methods. Infiltration of CD11b+ macrophages (A) and CD4+ T cells (B) into the spinal cord white matter is shown. Numerous DEC-205+ (C and D) and MIDC-8+ (F and G) cells are present within the perivascular and submeningeal inflammatory cell infiltrates in the spinal cord. Note the typical DC morphology of DEC-205+ cells accumulating within the cerebellar meningeal spaces (E). Staining of spinal cord sections for CD86 using the Adams’ amplification procedure (see Materials and Methods) evidenced the presence of CD86 immunoreactivity on most infiltrating cells, some activated microglia (H) and rare cells with irregularly shaped cell body and thin cytoplasmic processes typical of DCs (inset). Omission of the amplification step allowed to visualize only scattered CD86+ process-bearing cells, likely DCs, in the center of a submeningeal inflammatory cell infiltrate (I). Original magnifications: ×500 (A, B, C, E, and H) and ×1000 (D, F, G, I, and inset).
Figure 3.
Figure 3.
Persistence of mature DCs in the CNS of mice with chronic EAE. Representative thoracic (A, B, F, G, and H) and cervical (C) spinal cord and cerebellar (D and E) sections from a SJL mouse with chronic EAE (grade 3) are shown. CNS sections were stained for the indicated markers and visualized with DAB, as described in Materials and Methods. The immune infiltrates remain primarily confined to the meninges and around blood vessels and are composed of CD11b+ macrophages (A), CD4+ T cells (B), CD8+ T cells (C), as well as B220+ B cells (D). Cells positive for DEC-205 (E) and for the mature DC marker MIDC-8 (F) accumulate perivascularly and in the meninges. Anti-DEC-205 mAb stains process-bearing cells within the dorsal spinal cord white matter (G). Some of the MHC class II+ cells detected in the spinal cord meninges and white matter (H) have a DC-like morphology (arrowheads). Original magnifications: ×500 (AC and E–H), and ×1000 (D).
Figure 4.
Figure 4.
Diffuse infiltration of DCs with mature phenotype into the spinal cord white matter in EAE relapses. Representative thoracic (A and C–M) spinal cord and cerebellar (B) sections from a SJL mouse undergoing an EAE relapse (grade 2) are shown. CNS sections were stained for the indicated markers and visualized with DAB, as described in Materials and Methods. A–C: Shown are the distributions of CD4+ T cells, CD8+ T cells, and B220+ B cells recruited to the CNS during the EAE relapse phase. Numerous DEC-205+ (D) and MIDC-8+ (E) cells are present in the perivascular location as well as within the spinal cord parenchyma. DEC-205+ cells (D) co-localize with the CD4+ T cell infiltrates (A; adjacent spinal cord sections are shown in A and D). Note the different distribution and morphology of CD11b+-activated microglia (F), intraparenchymal DEC-205+ (G), and MIDC-8+ (H) DCs (dorsal part of adjacent spinal cord sections). CD11b+ microglia have elongated cell bodies and thick ramified processes (inset in F) whereas DEC-205+ DCs have a more rounded, irregular cell body with thin and long cytoplasmic processes (inset in G). I–M: Shown are the distributions of MHC class II, CD40, and CD86 molecules on intraparenchymal cells most of which exhibit a DC-like morphology. Original magnifications: ×500 (A–D, F, G, I, L, and M) and ×1000 (E, H, and insets).
Figure 5.
Figure 5.
MIP-3α and CCR6 transcripts are up-regulated in the CNS of SJL mice with PLP 139-151-induced EAE. RNA was extracted from different CNS areas and from control tissues (spleen and small intestine), reverse-transcribed, and subjected to PCR amplification using MIP-3α- and CCR6-specific primers, as described in Materials and Methods. A RT-PCR using glyceraldehyde-3-phosphate dehydrogenase-specific primers is also shown as an internal control. RNA was extracted from control mice (PBS/CFA-injected) and from mice at the indicated EAE stages. The data shown were obtained in mice with grade 3 acute EAE and grade 4 relapsing EAE. The PCR products were visualized by ethidium bromide staining. Lane 1, cerebrum; lane 2, cerebellum; lane 3, spinal cord; lane 4, spleen; and lane 5, small intestine.
Figure 6.
Figure 6.
Localization of MIP-3α immunoreactivity in the spinal cord of SJL mice with acute (A) and relapsing (B and C) EAE. Representative lumbar spinal cord sections are shown. Sections were stained for MIP-3α and visualized with DAB, as described in Materials and Methods. During acute EAE, MIP-3α immunoreactivity is localized in most cells within meningeal, submeningeal, and perivascular inflammatory infiltrates and in a few cells scattered in the spinal cord white matter (A). After an EAE relapse, numerous MIP-3α+ cells are also found within the spinal cord (B). At higher magnification, most intraparenchymal MIP-3α+ cells show the typical astrocytic morphology (C). Original magnifications: ×500 (A and B) and ×1000 (C).

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