Oligodendrocytes in eae and multiple sclerosis

91 37 OLIGODENDROCYTES IN EAE AND MULTIPLE SCLEROSIS H.Lassmann Research Unit for Experimental Neuropathology, Austrian Academy of Sciences. The destruction of myelin sheaths is one of the hallmarks of the pathology of multiple sclerosis and chronic EAE. Although the patterns of myelin destruction have been extensively studied, rather little is known about the fate of oligodendrocytes in these conditions. In a systematic study of oligodendrocyte pathology in EAE, we found that - depending upon the immunopathogenesis - different degrees and patterns of oligodendroglia damage occur. In pure models of T-cell mediated EAE oligodendrocytes are generally spared. Yet in rare occasions death of oligodendrocytes by apoptosis may occur. In contrast, when demyelinating antibodies are involved in the pathogenesis of the lesions, extensive destruction of oligodendrocytes by necrosis is encountered. In different multiple sclerosis cases a wide spectrum of oligodendrocyte pathology can be found, ranging from complete destruction to demyelination in the absence of oligodendrocyte damage. In some cases oligodendrocyte appear to be the primary target, whereas in others the cells are destroyed in parallel with the myelin sheaths. Our data suggest that the pathogenetic mechanisms, that lead to demyelination in different multiple sclerosis patients, may be variable in different patients and/or in different stages of the disease of an individual patient. 39 MACROPHAGES AND MICROGLIAL CELLS IN THE PATHOGENESIS OF EXPERIMENTAL ALLERGIC ENCEPHALOMYELITIS IlW Free University, Amsterdam, The Netherlands Experimental allergic encephalomyelitis (EAE) is an immune mediated inflammatory disease of the central nervous system. In particular in the chronic form of EAE, the inflammatory perivascular lesions are accompanied by demyelination. EAE can be induced in Lewis rats by immunization with myelin components or by transfer of myelin-sensitized T lymphocytes. In addition to T cells, macrophages are found in the perivascular infiltrates in course of EAE. These macophages are crucial for the development of clinical signs. In addition microglial cells undergo phenotypical changes in course of EAE, suggestive for an active role in the local immune response and/or inflammation. This activation of microglial cells

38 MACROPHAGES IN DEMYELINATION AND REMtiLINATION M.L. Cuzner. A.J. Louehlin & J. Newcombe, Institute of Neurology. London,

UK. Lipid-tilled

or circulating have screened obse~wJ of

normal

cithcr

suggesting

ester m the wy

that

from

arcas ofwlute

melter

We and

in the presence

to myelin

may rcllcct

microglia

MS plaques. in MS brain

in mncrophages

in addition

early lcsmn

resident

of dempclinnting

cstcr and I.DL colccnlized

myelin,

cholesterol

originating

we charnctcristic

microscopically

cholcstnol

nomxd

mncrophnges

monoqcs

breakdown

cnrly Bl3R

the

breakdown.

When the cytokinc spectrum of actively demyelinaling MS lesions is investigntcd,inllammato~yplaques arc found to be rich m IFNy and IL-2, while plnqucs

with

phogocytic

macrophnges

have higher

levels of TNF

and IL-I

as

MMP-9 Protein and mRNA are detectable by immunohistochenls~~ and Titus hybridization, in the macmpbages and lymphocytes of inflammatory MS plaques and in microglia well

ns matrix

in the adjacent

metolloproteinoses

white

matter.

Enrichment rewlts

of myelinaling

m a sv#icnnt

producmg

growth

demyrlination

incrensc factors

shmulnte

content,

stimultltc

wilh

m Idcntlljwg

with

indicative

myclin

the cyrokines

is greater dum in contml

pokntinl

cdhres

agpre@te

III myclin

!\‘hicII

of rhese cultures

extent of remyelinntion hc therapeutic

(MMR).

mncrophrges

of macrophnges

synthesis. TNF,

CIIIhIres.

Following

IL-l, or IFNv the ‘There may therefore

mnc~ophoge-dc~i\‘ed

factors

which

can

rcmyelination

40

THE REPAIR OF TRANSPLANTED

DAMAGED

GLW,

ENVIRONMENTS

BY

ASTBOCYTES

W.F.Blakemore, University of Cambridge, Cambridge CB3 OES, UK Glial cell transplantation provides a tool to explore cell-cell interaction and lineage in the CNS. Using this techniques we have compared the ability of tramplanted astrocytes derived from clonal bipotential gIiaI prcgenitor cell lines (“type-2 astrocytes”) to that of tissue culture aatmc@s (“type-1 astrccytes) in reconstNcti ng an ashuqte environment around dmnyulinated axons in the presence and absence of Schwann cells and meningeal ceils. Tissue c&are astroc@a, but not progenitor derived astmqtes, are strongly influenced by the pnsence of meningeal cells. Thus, when tissue culture astrocytes are transplanted without O2A lineage calls, they form cords of cells joined by gap junctions and show little ability to coot4 Schwaon cell invasion into gliel free areas of domyelinatioo created by the injection of ethidium bromide in white matter tracts of the spinal cord. However, when the normeI repair reqonsee of the Ieeioo are qpmeeed by xirradiation and Schwann cells and meningeal cells are mmoved fkom the transplant, they can form an integrakd .&al environment. Rcgenitor derived astrocytes show a greater ability to reform a glial environment, forming a loose cellular matrix around the demyelinated axons even in the presence of

meningeal cells.

42 ON

OUOODEIiDllOCVlE MOUFERAltONl

AND DiFFEREMltANON INVtlRO C.&rest6 0. D’uno. 6. Levi.lstitutoSupwiorn diSanita’,Rome,Italy Purifiedoligodsndroqtes and rd@kndmqto pmcursars obtainedfromneonatalrat brainprimarycultureswornsubcultured in a defined.-free medium and exposad10 IFN? (l-100 Ulmll.TNFa 05.100 t@ml)or both(100 Ulmland 50 nghd, respectively) fromday1 103 or day3 106. Whilecallsurvival was notaffectedin anyof the conditions tested,IFNr dase&psadanntly inhibited 13Hlthymidii@ or BrdUincorporation (upto 50%) andMTT cellular reduction I+ to 33KI. TNF-x syoqimd with IFN-y, but was ineffective by itself. Moreover. IFNy totally antagonized the bF6F. and PO6F4tduced proliferation of the oligodandrocyte population odor study. IFNy also blocked the differentiation of oligodmdrocyte precursors, as evidanmd by mll motphology, imunostaining for GalC and M6P and activity of CGdT. Again, th8 effect of IFNy was potentiated by TNFa, which was ineffective wk~ tested done. The inhibitory activity of 1FN.y was rapidly reversible: 3 days after rcrnev~l of the cytokii, odmbds&red from day 1 to 3, a complete recovery of cell proliintion md diffemntiation could be documented. The cytokirm4nduod arrest in the sxpressim~ of Mfmwtiatiw, antigens was accoqnnial by perturbatmns in the axprassion of the corrqowkg mRNAs, as renakd by semiquantitatiw RT.PCR. In particular, the mesqes for M6P and, in tbs casa of treatment from days 3 to 6, alsothat for MAGwnradscnasadin adturasexposed10 1FN.y.andfurtherdepressad in cultures treatedwithIFNy andTNFq whii TNFQ don was imfktivs. Thaabove0bseNations may hetp explairdng the mle of lFN.7 and TNF.a in the pathogatwsis 01 inflarmMory demyalinating diseases. in which local increases in the cerebral levels of these substances Cave bsen described.

INDUCIBLE NITRIC OXIDE SYNTHASE IN MS LESIONS.-‘, SC1&Bat&&iz~CF_lAlbert Einstein College of Medicine, Bronx,NY, USA, ‘Universita La Sapienza, Rome, Italy Nitric oxide (NO) is generated by at least three isofonns of nitric oxide synthase. cNOS and eNOS (nos I and nos III) are constitutively expressed at low levels. In contrast, iNOS (nos II) transcription is induced by cytokines. In humans the presence of iNOS in peripheral blood macrophages is controversial. Furthermore, it has not been observed in microglia in vtio or in vitro. iNOS mRNA and nitric oxide synthase enzyme activity have been shown in activated fetal astrocytes in vitro. In addition, NADPH diaphorase, a histochemical marker, demonstrates nitric oxide synthase activity in hypertrophic astrocytes in acute MS lesions. Using non-radioactive in situ hybridization, we detected mRNA for the inducible form of nitric oxide synthase in these astrocytes. In acute MS lesions, cytoplasm of hypertrophic astrocytes was intensely stained. Staining was also observed in astrocytic foot processes as well as in about 50% of endothelial cells. In subacute lesions, astrocyte staining was less intense than in acute, and fewer endothelial cells were positive for iNOS mRNA. In chronic active lesions only astrocytes showed expression of INOS, however, it was at a lower level than those in acute lesions. No iNOS mRNA was detected in macrophages or microalia in lesions at any stage. Normal appearing white matter was negative aswell. (Supported by NIH T32 GM07288,NIMH