Migration of dendritic cells into the brain in a mouse model of prion disease

Migration of dendritic cells into the brain in a mouse model of prion disease

Journal of Neuroimmunology 165 (2005) 114 – 120 www.elsevier.com/locate/jneuroim Migration of dendritic cells into the brain in a mouse model of prio...

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Journal of Neuroimmunology 165 (2005) 114 – 120 www.elsevier.com/locate/jneuroim

Migration of dendritic cells into the brain in a mouse model of prion disease Barbara Rosicarelli, Barbara Serafini, Marco Sbriccoli, Mei Lu, Franco Cardone, Maurizio Pocchiari, Francesca Aloisi* Department of Cell Biology and Neurosciences, Istituto Superiore di Sanita`, Viale Regina Elena 299 00161, Rome, Italy Received 17 March 2005; accepted 29 April 2005

Abstract The immune system plays a key role in the dissemination of prion infections from the periphery to the central nervous system (CNS). While follicular dendritic cells are critical for prion replication in lymphoid tissue and subsequent neuroinvasion, myeloid dendritic cells (DCs) have been implicated in both the clearance and propagation of pathological prion protein. Since nothing is known on the ability of DCs to migrate to the CNS during prion diseases, we investigated the immunohistochemical localization of CD205+ DCs in the brain of C57BL/6 mice intraperitoneally infected with the mouse-adapted KFu strain of Gerstmann – Stra¨ussler – Scheinker syndrome, a human genetic prion disorder. In normal brain, CD205+ cells were present in the meninges and choroid plexus, whereas in the majority of mice sacrificed between 120 and 300 days post infection, CD205+ DCs were also detected in the cerebral cortex, subcortical white matter, thalamus and medulla oblongata. These findings demonstrate that DCs can enter the CNS of prion-infected mice, suggesting a possible role for these cells in the pathogenesis of prion disorders. D 2005 Elsevier B.V. All rights reserved. Keywords: Prion disease; Dendritic cells; Immune system; Mouse brain

1. Introduction Prion diseases, also known as transmissible spongiform encephalopathies (TSEs), are inevitably fatal neurodegenerative disorders which affect humans and a variety of animal species. In humans, TSEs may have an undetermined (sporadic Creutzfeldt – Jakob disease [sCJD]), infectious (variant CJD, iatrogenic CJD, kuru), or inherited (familial CJD, Gerstmann – Stra¨ussler – Scheinker Syndrome, Fatal Familial Insomnia) etiology. All TSEs are characterized by the accumulation in the CNS of a protein called PrPTSE, which is an abnormally folded form of a self-encoded protein (cellular prion protein or PrPc) and is associated with TSE infectivity (Bolton et al., 1982). In spite of the lack of any signs of a typical immune response, it is now well established that the immune * Corresponding author. Tel.: +39 0649902087; fax: +39 064957821. E-mail address: [email protected] (F. Aloisi). 0165-5728/$ - see front matter D 2005 Elsevier B.V. All rights reserved. doi:10.1016/j.jneuroim.2005.04.017

system has a critical role in TSE pathogenesis (Aucouturier and Carnaud, 2002). After infection through peripheral routes (oral, parenteral), replication of most TSE agents primarily occurs in lymphoid tissues, such as spleen and lymph nodes, before neuroinvasion and detection in the CNS (Aguzzi, 2003). Experimental studies indicate that follicular dendritic cells (FDCs) residing in the B-cell follicles of secondary lymphoid organs are the primary sites of PrPTSE accumulation and participation in the process of neuroinvasion (Montrasio et al., 2000; Mabbott et al., 2000). However, since FDCs are not migratory cells and are unlikely to be responsible for the transport of the infectious agent into the CNS, the innervation of secondary lymphoid organs, and more specifically the topographical relationships between sympathetic nerves and FDCs, have been proposed to contribute to neuroinvasion (Aguzzi et al., 2004). Recent experimental evidence suggests that DCs might also play a role in TSE pathogenesis and affect the balance

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between propagation and clearance of PrPTSE in the organism (reviewed by Huang and Mac Pherson, 2004). Data by Huang et al. (2002) indicate that CD11c+ myeloid DCs can transport PrPTSE from the intestinal lumen to mesenteric lymph nodes. Aucouturier et al. (2001) have shown that infected CD11c+ splenic DCs alone, when injected systemically in RAG knockout mice, are sufficient for infectivity propagation and transport to the CNS. In vitro studies suggest that the PrP fragment 106 – 126 can act as a chemoattractant for monocyte-derived DCs (Le et al., 2001; Kaneider et al., 2003), and that DCs may protect against the infection by causing an efficient degradation of PrPTSE (Luhr et al., 2002). In normal conditions, DCs are excluded from the CNS parenchyma and can be found only in brain-associated tissues, such as the meninges and the choroid plexus (McMenamin, 1999). However, in a variety of neuropathological conditions, including ischemia (Kostulas et al., 2002; Reichmann et al., 2002); parasitic infection (Fischer et al., 2000); neurodegenerative disorders such as amyotrophic lateral sclerosis (Henkel et al., 2004), multiple sclerosis and its animal model experimental autoimmune encephalomyelitis (Serafini et al., 2000; Suter et al., 2000; Kivisa¨ kk et al., 2004; Serafini et al., submitted for publication), DCs are actively recruited to the CNS where they accumulate predominantly in the perivascular space of cerebral blood vessels, where they are thought to play a role in regulating local immune reactivity (Greter et al., 2005; McMahon et al., 2005). Since nothing is known on the ability of DCs to migrate to the CNS during TSE infection, in this study we used immunohistochemical techniques to investigate the presence of CD205+ DCs in the brain of C57BL/6 mice that were infected intraperitoneally with the mouse-adapted KFu strain of Gerstmann – Stra¨ussler – Scheinker syndrome, a human genetic prion disorder (Tateishi et al., 1979).

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Groups of three animals were sacrificed by CO2 asphyxia and their brains were removed at 120, 180, 240 days post infection (d.p.i.) and at the terminal stage of the disease (300 T 13.2 d.p.i. [mean T SD]). Animals of the control group were injected with normal mouse brain homogenate and sacrificed at the same time points. Animals were observed daily and scored for the presence of clinical signs.

2. Materials and methods 2.1. Animals Six week-old female C57BL/6 mice (purchased from Charles River, Calco, Como, Italy) were housed 8– 10 per cage in a controlled environment in accordance with the guidelines of the European Community Council of the Welfare of Experimental Animals (86/609/EEC). All experimental procedures were approved by the Italian Ministry of Health. 2.2. Mouse inoculation Mice were injected intraperitoneally with the mouseadapted KFu strain of Gerstmann –Stra¨ussler– Scheinker syndrome (Tateishi et al., 1979) obtained from a 10% (w/v) PBS suspension of pooled brains from clinically ill animals.

Fig. 1. Histopathological features of brains from C57BL/6 mice infected with the mouse-adapted KFu strain of Gerstmann – Stra¨ussler – Scheinker syndrome. (A) First signs of spongiform changes in the white matter of the medulla oblongata in a KFu-infected mouse at 180 days post-infection. (B) Severe spongiform changes in the cerebellar white matter of KFu-infected mice at the terminal stage of the disease. (C) Scattered fine deposits of PrPTSE in the thalamus (arrow). Severe spongiform changes are also evident. Original magnifications: A and C = 500, B = 250.

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Table 1 Neuropathological characteristics and quantification of CD205+ DCs in the CNS of KFu-infected mice Mouse

D.p.i.a

Spongiosis

PrPTSE

366 – 3 366 – 4 366 – 5 369 – 3 369 – 4 369 – 5 370 – 2 370 – 3 370 – 4 370 – 8 370 – 9 370 – 10

120 µ µ 180 µ µ 240 µ µ 300 T 13.2 µ µ

No No No No No Yes Yes Yes No Yes Yes Yes

No No No No No No Yes Yes No Yes Yes Yes

Frequency of CD205+ cells at each CNS level examinedb A

B c

C

D

+ +

++

+ +++ +

+ +

+ + +

+ +

+ ++

++ + + + +

a

D.p.i.=days post infection. Mice were inoculated intraperitoneally with KFu strain and at the indicated days were sacrificed and processed for spongiosis evaluation and immunohistochemistry for PrPTSE and CD205. b For each mouse, four brain levels were analysed for the presence of CD205+ DCs: A=anterior cortex and septal nuclei; B=thalamus, hypothalamus, hippocampus and posterior cortex; C=superior colliculus; D=cerebellum and medulla oblongata. c The frequency of CD205+ DCs was evaluated using the following score: = no cells/section; + = 1 – 5 cells/section; ++ = 6 – 10 cells/section; +++ = > 10 cells/section.

Prior to processing and embedding, brains were trimmed coronally into four 2-mm thick slices, one including anterior cortex and the septal nuclei (level A), one including thalamus, hypothalamus, hippocampus and posterior cortex (level B), one including the superior colliculus (level C), and one including the cerebellum and the medulla oblongata (level D). The slices were immersed in formic acid (98%) for 1 h to reduce infectivity, dehydrated in graded alcohols and then embedded in paraffin wax. Five-Am-thick sections were cut on microtome and stained with hematoxylin/eosin for the evaluation of neuronal damage and for the construction of the lesion profile according to the scoring system introduced by Fraser and Dickinson (1968) on a scale of evaluation graded from 1 to 3. Brain sections were immunostained for the presence of the pathological prion protein (PrPTSE) using the mouse monoclonal antibody (mAb) SAF84 (SPI-BIO, Massy, France). DCs were identified using anti-CD205 rat mAb (NLDC 145, Serotec, Oxford, UK), which binds an integral membrane glycoprotein highly expressed on DCs (Jiang et al., 1995). CD205 antigen is not expressed in macrophages and T cells from bone marrow and peripheral blood (Inaba et al., 1995) and has not been detected in the normal mouse brain (Jiang et al., 1995). PrPTSE immunostaining was carried out according to the ABC method. Briefly, sections were deparaffinized, rehydrated, autoclaved at 121 -C for 30 min in distilled water (hydrated autoclaving), treated with 98% formic acid for 1 min at room temperature to enhance staining, rinsed and then incubated overnight at 4 -C with mouse mAb SAF84 (1.5 Ag/ml). Subsequent antibody detection involved incubation with biotinylated goat antimouse secondary antibody for 1 h (1 : 200 dilution, Vector Laboratories) at room temperature, followed by incubation

with the avidin – biotin – peroxidase complex (Vectastain ABC-Elite kit, Vector Laboratories). The samples were stained with 3,3V diaminobenzidine (DAB, Sigma) to visualize the reaction product and then counterstained with hematoxylin. Immunostaining with anti-CD205 mAb was performed following the Tyramide Signal Amplification technique using the TSA Biotin System Kit-NEN (Perkin Elmer Life Science, Inc. Boston, MA) according to the manufacturer’s instructions. Briefly, deparaffinized brain and cerebellum sections were submitted to the high temperature antigen unmasking procedure, performed in Dako target retrieval solution (Dako, Carpinteria, CA) for 30 min at 95 -C, incubated with blocking solution for 1 h, and then with antiCD205 rat mAb, at the dilution of 1 : 20, overnight at 4 -C. After extensive washes with PBS, sections were treated for 1 h at room temperature with the biotinylated secondary 3,5 3

Vacuolation score

2.3. Histology and immunohistochemistry

2,5 2 1,5 1 0,5 0 1

2

3

4

5

6

7

8

9

Scoring area

Fig. 2. Lesion profile in infected mice at the terminal stage of disease. 1: medulla oblongata; 2: cerebellar cortex; 3: superior colliculus; 4: hypothalamus; 5: thalamus; 6: hippocampus; 7: septum; 8 and 9: posterior and anterior cerebral cortex, respectively. Plotted data are mean T SD from three animals.

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septal nuclei (level A), posterior cortex, hippocampus, thalamus and hypothalamus (level B), superior colliculus (level C), cerebellum and medulla oblongata (level D). Counting of DCs in brain sections from the different animal groups was performed by two independent observers in a blinded fashion. The score used was: = no cells/section; + = 1– 5 cells/section; ++ = 6– 10 cells/section; +++ = > 10 cells/section.

3. Results Histological analysis of the brains of KFu-infected mice revealed that the earliest vacuolar changes occurred in the internal capsule and medulla oblongata, and were detected in some mice already at 180 d.p.i. (Fig. 1A and Table 1). By 240 d.p.i., spongiform changes also developed in the corpus callosum, thalamus and cerebellar white matter. In mice sacrificed at the terminal stage of the disease, severe spongiform changes were evident in all brain areas examined, including the cerebellum (Fig. 1B and Table 1). The lesion profile observed at the terminal stage of the disease was similar to that previously described after intracerebral infection (Kordek et al., 1999) (Fig. 2).

Fig. 3. CD205 immunostaining in normal and control mouse brains. In control mice, both non-injected (A and B) and injected intraperitoneally with non-infected brain homogenate (C), anti-CD205 mAb stains a few ramified cells in the meninges (arrows in A and C) and in the choroid plexus (arrow in B), whereas no CD205+ cells are present in the brain parenchyma (C). Original magnifications: A and B = 1575; C = 250.

antibody (rabbit anti-rat IgG, 10 Ag/ml, Vector Laboratories, Burlingame, CA) and then subjected to the signal amplification procedure. Staining reaction was visualized using DAB as substrate. Lymph nodes and spleen from healthy mice were used as positive controls; negative controls were performed by omission of the primary antibody from the incubation medium. All sections were counterstained with hematoxylin and viewed with a Zeiss Axiophot photomicroscope. For quantification of CD205+ DCs, positive cells were counted at the level of the anterior cortex and

Fig. 4. Detection of CD205+ dendritic cells in the medulla oblongata of KFu-infected mice at 180 d.p.i. Numerous CD205+ cells displaying irregular shapes and short cytoplasmic processes (arrows) are present in the white matter of the medulla oblongata adjacent to the spinal trigeminal tract (stt). Inset in A is a high power magnification of a portion of the field shown in panel A. Original magnifications: A = 1000; B and inset = 1575.

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Immunostaining for PrPTSE, the patholological prion protein, consistently revealed the presence of small deposits in the ventromedial and ventral posterolateral thalamic nuclei at 240 d.p.i. and more prominent accumulation at the terminal stage of the disease (Fig. 1C). At this stage, small deposits of PrPTSE were also found in the medulla oblongata (data not shown). Immunohistochemical evaluation of CD205+ DCs was performed in brain tissue sections obtained from mice sacrificed at 120 (n = 3), 180 (n = 3) and 240 (n = 3) d.p.i., and at the terminal stage of the disease (around 300 d.p.i.) (n = 3). For each mouse, four different brain levels were analysed for the presence of CD205 immunoreactivity (Table 1). In agreement with previous studies (Serafini et al., 2000), in both healthy and control mice CD205 mAb stained only a few ramified cells in the meninges and choroid plexus, but none inside the cerebral parenchyma (Fig. 3A –C). Variable numbers of CD205+ cells were detected in the brain of all mice injected intraperitoneally with the KFu strain. Numerous, sparse CD205+ cells were observed in the CNS of one out of three mice sacrificed at 120 d.p.i., and of two out of three mice sacrificed at 180 d.p.i. (Fig. 4 and Table 1). In these mice, which showed no evident intracerebral PrPTSE accumulation, CD205+ DCs with irregular cell bodies and often short cytoplasmic processes were

found predominantly in the white matter of the medulla oblongata, close to the spinal trigeminal tract and the cerebellar peduncle (Fig. 4A –B). At both 120 and 180 d.p.i., a few CD205+ DCs were also found in other brain areas, except the cerebellum and superior colliculus (Table 1). These data indicate that the presence of CD205+ cells precedes the appearance of vacuolar changes in the medulla oblongata of the infected mice. Compared to other CNS levels examined, this area had the highest vacuolation score also at the terminal stage of the disease (Fig. 2), suggesting an association between DC recruitment and brain damage. As shown in Table 1, CD205+ cells were present in the CNS of all six mice examined at 240 and 300 d.p.i., five of which showed both vacuolar changes and PrPTSE accumulation. Compared to control mice, increased numbers of CD205+ cells were detected inside the meninges (Fig. 5A) and choroid plexus (data not shown). Isolated CD205+ cells with an irregular or ramified morphology were also observed around some intraparenchymal blood vessels (Fig. 5B) and inside the neural parenchyma (Fig. 5C –F).

4. Conclusions In this study we provide immunohistochemical evidence that CD205+ DCs are present in the CNS parenchyma of

Fig. 5. Presence of CD205+ DCs in the CNS of KFu-infected mice at the terminal stage of the disease. CD205+ cells (arrows) are present inside the cerebral meninges (A), and around intraparenchymal blood vessels located in the area comprised between the hippocampus (CA3 field) and the thalamus (laterodorsal nucleus) (B, arrows). Isolated CD205+ dendritic cells are present in the neural parenchyma in close proximity to the 3rd ventricle (v) (C and D, arrows) and in the white matter of the medulla oblongata (E and F, arrows). Inset in C shows that CD205+ cells display the ramified morphology and irregular shape typical of dendritic cells. Original magnifications: A= 500; B – D and inset = 1000; E and F = 1575.

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C57BL/6 mice following intraperitoneal injection of the mouse-adapted KFu strain of Gerstmann – Stra¨ ussler – Scheinker syndrome. Since DCs are normally excluded from the CNS parenchyma, those present in the CNS of prion-infected mice are likely to derive from DCs or DC progenitors recruited from the blood circulation. This view is consistent with the presence of CD205 + cells in perivascular location in the brain of KFu-infected mice. The finding that intracerebral CD205+ DCs were detected in infected mice starting from day 120 post-infection to the terminal stage of the disease indicates continuous migration of DCs to the degenerating CNS. Notably, the observation that CD205+ DCs were detected in the medulla oblongata already at 120 –180 d.p.i. and that the first signs of vacuolar changes became evident in the same area at 180 d.p.i., suggests an early involvement of DCs in prion-induced brain pathology. DCs could be attracted by chemokines produced in the infected brain or their presence could be consequent to substantial damage of the blood-brain barrier. Due to the lack of substantial mononuclear cell infiltration in the CNS of KFu-infected mice (unpublished observations), we favour the idea of a chemokine-driven mechanism of DC recruitment. In line with this, RANTES, a chemokine which binds to the promiscuous chemokine receptors CCR1, CCR3 and CCR5, and also acts as a DC chemoattractant (Caux et al., 2000) was recently shown to be produced in the brain of scrapieinfected mice (Lee et al., 2005). Whether DCs migrating to the CNS facilitate neuroinvasion by transfer of the infectious agent (Aucouturier et al., 2001), or have a protective role by favoring proteolytic clearance of PrPTSE (Luhr et al., 2002), remains to be established. We would like to propose that, due to their ability to reach the CNS already during the early stages of prion propagation in the CNS, DCs could be considered as useful vectors for transfer of therapeutic molecules to the prion-infected brain.

Acknowledgements This work has been supported by grant ALZ/6 from the Italian Ministry of Health and research project 2ADI from the Istituto Superiore di Sanita`.

References Aucouturier, P., Carnaud, C., 2002. The immune system and prion disease: a relationship of complicity and blindness. J. Leukoc. Biol. 72, 1075 – 1083. Aucouturier, P., Geissmann, F., Damotte, D., Saborio, G.P., Meeker, H.C., Kascsak, R., Kascsak, R., Carp, R.I., Wisniewski, T., 2001. Infected splenic dendritic cells are sufficient of prion transmission to the CNS in mouse scrapie. J. Clin. Invest. 108, 703 – 708. Aguzzi, A., 2003. Prions and the immune system: a journey through gut, spleen, and nerves. Adv. Immunol. 81, 123 – 171.

119

Aguzzi, A., Heikenwalder, M., Miele, G., 2004. Progress and problems in the biology, diagnostics, and therapeutics of prion diseases. J. Clin. Invest. 114 (2), 153 – 160. Bolton, D.C., McKinley, M.P., Prusiner, S.B., 1982. Identification of a protein that purifies with the scrapie prion. Science 218, 1309 – 1311. Caux, C., Ait-Yahia, S., Chemin, K., 2000. Dendritic cell biology and regulation of dendritic cell trafficking by chemokines. Springer Semin. Immunopathol. 22, 345 – 369. Fischer, H.G., Bonifas, U., Reichmann, G., 2000. Phenotype and functions of brain dendritic cells emerging during chronic infection of mice with Toxoplasma gondii. J. Immunol. 164, 4826 – 4834. Fraser, H., Dickinson, A.G., 1968. The sequential development of the brain lesion of scrapie in three strains of mice. J. Comp. Pathol. 78, 301 – 311. Greter, M., Heppner, F.L., Lemos, M.P., Odermatt, B.M., Goebels, N., Laufer, T., Noelle, R.J., Becher, B., 2005. Dendritic cells permit immune invasion of the CNS in an animal model of multiple sclerosis. Nat. Med. 11, 328 – 334. Henkel, J.S., Engelhardt, J.I., Siklos, L., Simpson, E.P., Kim, S.H., Pan, T., Goodman, J.C., Siddique, T., Beers, D.R., Appel, S.H., 2004. Presence of dendritic cells, MCP-1, and activated microglia/macrophages in amyotrophic lateral sclerosis spinal cord tissue. Ann. Neurol. 55, 221 – 235. Huang, F.P., Mac Pherson, G.G., 2004. Dendritic cells and oral transmission of prion diseases. Adv. Drug Deliv. Rev. 56, 901 – 913. Huang, F.P., Farquhar, C.F., Mabbott, N.A., Bruce, M.E., MacPherson, G.G., 2002. Migrating intestinal dendritic cells transport PrPSc from the gut. J. Gen. Virol. 83, 267 – 271. Inaba, K., Swiggard, W.J., Inaba, M., Meltzer, J., Mirza, A., Sasagawa, T., Nussenzweig, M.C., Steinman, R.M., 1995. Tissue distribution of the DEC-205 protein that is detected by the monoclonal antibody NLDC145. Cell. Immunol. 163, 148 – 156. Jiang, W., Swiggard, W.J., Heufler, C., Peng, M., Mirza, A., Steinman, R.M., Nussenzweig, M.C., 1995. The receptor DEC-205 expressed by dendritic cells and thymic epithelial cells is involved in antigen processing. Nature 375, 151 – 155. Kaneider, N.C., Kesr, A., Tilg, H., Ricevuti, G., Wiedermann, C.J., 2003. CD40 ligand-dependent maturation of human monocyte-derived dendritic cells by activated platelets. Int. J. Immunopathol. Pharmacol. 16, 225 – 231. Kivisa¨kk, P., Mahad, D.J., Callahan, M.K., Sikora, K., Trebs, C., Tucky, B., Wujek, J., Ravid, R., Staugaitis, S.M., Lassmann, H., Ransohoff, R.M., 2004. Expression of CCR7 in multiple sclerosis: implications for CNS immunity. Ann. Neurol. 55, 627 – 638. Kordek, R., Hainfellner, J.A., Liberski, P.P., Budka, H., 1999. Deposition of the prion protein (PrP) during the evolution of experimental Creutzfeldt – Jakob disease. Acta Neuropathol. 98, 597 – 602. Kostulas, N., Li, H.L., Xiao, B.G., Huang, Y.M., Kostulas, V., Link, H., 2002. Dendritic cells are present in ischemic brain after permanent middle cerebral artery occlusion in the rat. Stroke 33, 1129 – 1134. Le, Y., Yazawa, H., Gong, W., Yu, Z., Ferrans, V.J., Murphy, P.M., Wang, J.M., 2001. The neurotoxic prion peptide fragment PrP(106 – 126) is a chemotactic agonist for the G protein-coupled receptor formyl peptide receptor-like 1. J. Immunol. 166, 1448 – 1451. Lee, H.P., Jun, Y.C., Choi, J.K., Kim, J.I., Carp, R.I., Kim, Y.S., 2005. The expression of RANTES and chemokine receptors in the brains of scrapie-infected mouse. J. Neuroimmunol. 158, 26 – 33. Luhr, K.M., Wallin, R.P.A., Ljunggren, H.-G., Low, P., Taraboulos, A., Kristensson, K., 2002. Processing and degradation of exogenous prion protein by CD11c+ myeloid dendritic cells in vitro. J. Virol. 76, 12259 – 12264. Mabbott, N.A., Mackay, F., Minns, F., Bruce, M.E., 2000. Temporary inactivation of follicular dendritic cells delays neuroinvasion of scrapie. Nat. Med. 6, 719 – 720. McMahon, E.J., Bailey, S.L., Castenada, C.V., Waldner, H., Miller, S.D., 2005. Epitope spreading initiates in the CNS in two mouse models of multiple sclerosis. Nat. Med. 11, 335 – 339.

120

B. Rosicarelli et al. / Journal of Neuroimmunology 165 (2005) 114 – 120

McMenamin, P.G., 1999. Distribution and phenotype of dendritic cells and resident tissue macrophages in the dura mater, leptomeninges, and choroids plexus of the rat brain as demonstrated in wholemount preparations. J. Comp. Neurol. 405, 553 – 562. Montrasio, F., Frigg, R., Glatzel, M., Klein, M.A., Mackay, F., Aguzzi, A., Weissmann, C., 2000. Impaired prion replication in spleens of mice lacking functional follicular dendritic cells. Science 288, 1257 – 1259. Reichmann, G., Schroeter, M., Jander, S., Fischer, H.G., 2002. Dendritic cells and dendritic-like microglia in focal cortical ischemia of the mouse brain. J. Neuroimmunol. 129, 125 – 132. Serafini, B., Columba-Cabezas, S., Di Rosa, F., Aloisi, F., 2000. Intracerebral recruitment and maturation of dendritic cells in the onset and progression of experimental autoimmune encephalomyelitis. Am. J. Pathol. 157, 1991 – 2002.

Serafini, B., Rosicarelli, B., Magliozzi, R., Capello, E., Mancardi, G.L., Aloisi, F., submitted for publication. Localization of immature and mature dendritic cells and relationship to T cells in multiple sclerosis lesions. Suter, T., Malipiero, U., Otten, L., Ludewig, B., Muelethaler-Motter, A., Mach, B., Reith, W., Fontana, A., 2000. Dendritic cells and differential usage of the MHC class II transactivator promoters in the central nervous system in experimental autoimmune encephalitis. Eur. J. Immunol. 30, 794 – 802. Tateishi, J., Ohta, M., Koga, M., Sato, Y., Kuroiwa, Y., 1979. Transmission of chronic spongiform encephalopathy with kuru plaques from humans to small rodents. Ann. Neurol. 5, 581 – 584.