Erythropoietin exerts an anti-inflammatory effect on the CNS in a model of experimental autoimmune encephalomyelitis

doi:10.1016/S0006-8993(02)03239-0

Brain Research

Volume 952, Issue 1, 11 October 2002, Pages 128-134

https://doi.org/10.1016/S0006-8993(02)03239-0 Get rights and content

Abstract

In recent work we reported that systemically administered erythropoietin (EPO) crosses the blood–brain barrier and has protective effects in animal models of cerebral ischemia, brain trauma and in a rat model of experimental autoimmune encephalomyelitis (EAE). Here we characterize the effect of systemic EPO on the inflammatory component of actively induced, acute EAE in Lewis rats. Administration of EPO at doses of 500–5000 U/kg bw i.p., daily from day 3 after immunization with myelin basic protein (MBP), delayed the onset of EAE and decreased its clinical score at peak time (days 12–13). Immunohistochemical analysis of the spinal cord using anti-glial fibrillary acidic protein (GFAP) and anti-CD11b antibodies showed that EPO markedly diminished inflammation and glial activation/proliferation. EAE induced significant levels of TNF and IL-6 in the spinal cord, where IL-6 was maximum at the onset of the disease (day 10) and TNF at its peak (day 12). EPO delayed the increase of TNF levels, without altering their peak levels, and markedly reduced those of IL-6 suggesting that the decreased inflammation and clinical score may be in part upon attenuation of IL-6. On the other hand, EPO was without effect in a model of adjuvant-induced arthritis in Lewis rats, suggesting a specificity towards autoimmune demyelinating diseases. These data suggest that EPO might act as a protective cytokine in inflammatory pathologies of the CNS.

Introduction

CNS diseases with an inflammatory component include cerebral ischemia, brain trauma, multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). In these diseases, various experimental strategies have been utilized to inhibit inflammatory cytokines, and anti-TNF antibodies, IL-1ra, or IL-10 are protective in animal models of these diseases. The extrapolation of these data to humans is particularly difficult for autoimmune diseases. For instance, although anti-TNF antibodies, or knocking out the TNF gene, are protective in animal models of EAE [3], [15] anti-TNF antibodies produced a clinical deterioration in MS [10]. It seems therefore important to identify complementary strategies for inhibiting inflammation in these diseases.

In recent publications [4], [23], we reported that systemically administered EPO crosses the blood–brain barrier to protect mice and rats from cerebral ischemia (middle cerebral artery occlusion), traumatic brain injury, kainic acid toxicity, and EAE in Lewis rats. In rats with cerebral ischemia, we observed that EPO decreases the inflammatory infiltrate and inflammatory cytokine production (P. Villa, manuscript in preparation).

In the present study, we evaluated the effect of systemic EPO administration on inflammation in the spinal cord of Lewis rats where EAE is induced by immunization with MBP in complete Freund’s adjuvant (CFA). In particular, we measured the levels of TNF and IL-6 in the brain and spinal cord, and performed a histological analysis of the inflammatory infiltrate and of estimated glial activation/proliferation by use of immunohistochemical staining for glial fibrillary acidic protein (GFAP) and CD11b. To investigate whether the beneficial effect of EPO was specific for EAE or was common to other autoimmune diseases, we tested the effect of EPO treatment in a model of adjuvant-induced arthritis in Lewis rats. Our findings indicate that EPO has an anti-inflammatory effect only in EAE.

Section snippets

Acute EAE in Lewis rats

Female Lewis rats, 6–8 weeks of age, were purchased from Charles River (Calco, Italy). Procedures involving animals and their care were conducted in conformity with the institutional guidelines that are in compliance with national (D.L. no. 116, G.U., Suppl. 40, 18 Febbraio 1992, Circolare no. 8, G.U., 14 Luglio 1994) and international (EEC Council Directive 86/609, OJL 358, 1, 12 December 1987; Guide for the Care and Use of Laboratory Animals, US National Research Council, 1996) laws and

EPO ameliorates the clinical signs of EAE in Lewis rats

Fig. 1 shows the protective effect on the clinical signs of EAE of different doses of EPO, given from day 3 after immunization with MBP, until day 18. EPO, in a dose-dependent fashion, delayed the onset of disease and decreased disease severity, as summarized in Table 1. As shown in this table, EPO at the doses of 500, 2500 or 5000 U/kg bw significantly decreased the mean cumulative score.

In experiments where treatment of EPO was discontinued after the disease regressed, and the rats were

Discussion

Results of the present study demonstrate an ameliorating effect of EPO in actively induced EAE in rats, which is maintained without further EPO administration for at least 2 months. Further, EPO decreased the peak clinical severity of the disease in a dose-dependent manner and this effect is paralleled by a marked decrease in inflammatory infiltrate and glial activation/proliferation, as assessed by anti-GFAP and anti-CD11b immunohistochemistry. Since there is evidence that TNF inhibition

Acknowledgements

Davide Agnello is a fellow of the Alfredo Leonardi Fund and G.L. Pfeiffer Foundation. P.B. is a recipient of a Fondazione Monzino fellowship.

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¹: D.A. and P.B. contributed equally to this work.

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Research reportErythropoietin exerts an anti-inflammatory effect on the CNS in a model of experimental autoimmune encephalomyelitis

Abstract

Introduction

Section snippets

Acute EAE in Lewis rats

EPO ameliorates the clinical signs of EAE in Lewis rats

Discussion

Acknowledgements

J. Neuroimmunol.

Am. J. Kidney Dis.

Biochem. Biophys. Res. Commun.

J. Neuroimmunol.

Erythrocyte catalase. A somatic oxidant defense?

J. Clin. Invest.

Control of established experimental allergic encephalomyelitis by inhibition of tumor necrosis factor (TNF) activity within the central nervous system using monoclonal antibodies and TNF receptor-immunoglobulin fusion proteins

Eur. J. Immunol.

Erythropoietin crosses the blood–brain barrier to protect against experimental brain injury

Proc. Natl. Acad. Sci. USA

Erythrocytes decrease myocardial hydrogen peroxide levels and reperfusion injury

Am. J. Physiol.

Recombinant human erythropoietin stimulates production of interleukin 2 by whole blood cell cultures of hemodialysis patients

Artif. Organs

IL-6-deficient mice resist myelin oligodendrocyte glycoprotein-induced autoimmune encephalomyelitis