SummaryImmunohistochemical methods were used to search for Fas receptor/Fas ligand system involvement in multiple sclerosis (MS) white matter brain lesions. We found large numbers of Fas ligand (Fas-L)-bearing cells present in two acute lesions and 12 of 16 chronic MS lesions, and very few positive cells in non-inflammatory controls. Four of six brains from non-MS neuropathologic conditions associated with inflammation and white matter disease were, however, also positive for Fas-L. Double staining with cell-specific markers revealed that the pattern of ligand-positive cells in chronic MS lesions was complex and composed of several different cell types which were primarily resident glial cells with a small overlay ofmacrophages. Fas/APO 1 (CD95) receptor expression in MS tissue was also evaluated and marked upregulation of the receptor was found. In addition, Fas receptor was induced, but to a lesser extent, in numerous control brains. The observations that TUNEL-positive dying cells were present in MS lesions and showed excellent co-localization with Fas-L, indicate that the Fas death system may contribute to plaque pathogenesis and could lead to the development of a new category of therapeutic agents for MS.

BackgroundBeneficial effects of short-term erythropoietin (EPO) therapy have been demonstrated in several animal models of acute neurologic injury, including experimental autoimmune encephalomyelitis (EAE)-the animal model of multiple sclerosis. We have found that EPO treatment substantially reduces the acute clinical paralysis seen in EAE mice and this improvement is accompanied by a large reduction in the mononuclear cell infiltration and downregulation of glial MHC class II expression within the inflamed CNS. Other reports have recently indicated that peripherally generated anti-inflammatory CD4+Foxp3+ regulatory T cells (Tregs) and the IL17-producing CD4+ T helper cell (Th17) subpopulations play key antagonistic roles in EAE pathogenesis. However, no information regarding the effects of EPO therapy on the behavior of the general mononuclear-lymphocyte population, Tregs or Th17 cells in EAE has emerged.Methods and FindingsWe first determined in vivo that EPO therapy markedly suppressed MOG specific T cell proliferation and sharply reduced the number of reactive dendritic cells (CD11c positive) in EAE lymph nodes during both inductive and later symptomatic phases of MOG35–55 induced EAE. We then determined the effect in vivo of EPO on numbers of peripheral Treg cells and Th17 cells. We found that EPO treatment modulated immune balance in both the periphery and the inflamed spinal cord by promoting a large expansion in Treg cells, inhibiting Th17 polarization and abrogating proliferation of the antigen presenting dendritic cell population. Finally we utilized tissue culture assays to show that exposure to EPO in vitro similarly downregulated MOG-specific T cell proliferation and also greatly suppressed T cell production of pro-inflammatory cytokines.ConclusionsTaken together, our findings reveal an important new locus whereby EPO induces substantial long-term tissue protection in the host through signaling to several critical subsets of immune cells that reside in the peripheral lymphatic system.

We have known for a long time that erythropoietin signaling plays a key role in bone marrow erythrocyte proliferation. However, recent studies have indicated that erythropoietin also may have protective effects on the nervous system. This unexpected role remains incompletely characterized. To investigate the potential neuroprotective role of erythropoietin in the central nervous system, we assessed its effects on a well-characterized autoimmune demyelinating model of multiple sclerosis-myelin oligodendrocyte glycoprotein-induced experimental autoimmune encephalomyelitis (EAE) in the mouse. We found that erythropoietin administered intravenously for 14 days after the onset of symptoms reduced both disease severity and duration of maximum impairment at dose levels as low as 50U/kg (p < 0.001). We assessed the neuropathology of diseased spinal cords and found that erythropoietin-treated EAE animals had reduced axonal damage, inflammatory cell infiltration and demyelination, and diminished blood-brain barrier leakage when compared with saline-treated EAE controls. Moreover, the pronounced upregulation of spinal cord major histocompatibility complex (MHC) class II expression found in saline-treated EAE was significantly reduced in erythropoietin-treated animals, a finding we replicated in vitro, using microglial cultures. The notion that short-term erythropoietin therapy might be of clinical benefit in human autoimmune demyelinating diseases needs investigation.

Measles virus (MV) mRNA transcription and replication are thought to be controlled by cis-acting sequence elements contained within the terminal MV genomic noncoding nucleotides. To validate these promoter and regulatory signal assignments, cDNAs were constructed allowing synthesis of RNAs corresponding to a MV genome in which all coding and intercistronic regions were replaced by the chloramphenicol acetyl transferase (CAT) coding sequence. Transcript production by T7 polymerase starting and ending precisely with the MV genome terminal residues was achieved by fusing the T7 polymerase promoter and the hepatitis delta virus genome ribozyme followed by tandem T7 polymerase termination sequences to the MV genomic 5' and 3' ends, respectively. Transfection of these negative polarity transcripts, mimicking natural defective interfering RNAs of the internal deletion type, into MV-infected 293 cells gave rise to CAT activity which could be serially transferred and massively amplified together with progeny helper virus in fresh cells. Transfer was blocked only by antibodies able to neutralize MV infectivity, indicating that the chimeric RNA not only was encapsidated, transcribed, and replicated, but also packaged into virions. Sequence analyses confirmed that both the expected chimeric antigenome and mRNA products were transcribed and replicated with fidelity during serial passage. Minor changes introduced in the transcription promoter markedly compromised function. This system now can be exploited to examine MV genomic cis-acting regulatory elements and extended to the development of full-length MV cDNAs.

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Platelet-derived growth factor (PDGF) ligand is a potent glial cell mitogen. When its cognate receptor (PDGF-alphaR) is expressed on oligodendroglial lineage cells, such cells are considered capable of division, and the receptor thus serves as a phenotypic marker for oligodendrocyte precursor cells. Here we identify using immunohistochemistry a considerably enlarged, PDGF-alphaR-expressing oligodendrocyte cell population within multiple sclerosis (MS) white matter lesions compared to control brains. Numerous PDGF-alphaR-positive oligodendroglia also colabel heavily with the nuclear cell proliferation marker antibody Ki-67. Our finding of large numbers of proliferating oligodendroglia in MS brains expressing up-regulated PDGF-alphaR suggests that these progenitor-like cells represent an important source of regenerating cells for the healing MS lesion.

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