Effector stage CC chemokine receptor-1 selective antagonism reduces multiple sclerosis-like rat disease  Sana Eltayeb, Dan Sunnemark, Anna-Lena Berg, Gunnar Nordvall, Åsa Malmberg, Hans Lassmann, Erik Wallström, Tomas Olsson, Anders Ericsson-Dahlstrand  Journal of Neuroimmunology  Volume 142, Issue 1, Pages 75-85 (September 2003) DOI: 10.1016/S0165-5728(03)00264-9

Fig. 1 Structure of the CCR1 selective antagonist. Journal of Neuroimmunology 2003 142, 75-85DOI: (10.1016/S0165-5728(03)00264-9)

Fig. 2 Clinical effects of CCR1 antagonist. Clinical EAE disease courses of CCR1 antagonist (drug)-treated rats and vehicle-injected controls. Rats were injected with MOG and treated from day 8 post immunization (group A, n=20), or from individual onset of disease (group B, n=16) with either drug or with vehicle (cyclodextrin/H2O). Rats were treated twice daily for five consecutive days and scored on a daily basis. The data are combined from two independent experiments and expressed as mean±2 S.E.M. The graph in group B is normalized to individual onset of disease (d0). Black bar below each graph indicates period for treatment. ∗p<0.05, ∗∗p<0.01. ∗∗∗p<0.001, ∗∗∗∗p<0.0001, Mann–Whitney U-test. Journal of Neuroimmunology 2003 142, 75-85DOI: (10.1016/S0165-5728(03)00264-9)

Fig. 3 Histopathological evaluation of drug- and vehicle-treated EAE rats. Effects on histopathological manifestations of EAE following treatment with the CCR1 antagonist or vehicle was evaluated on tissues obtained from group A rats (n=8, perfusion fixation 16 h after last injection). Spinal cord sections from drug-treated rats (b, d, f, h) demonstrate a complete blockade of inflammatory cell infiltration as assessed by HTX eosin staining (b). In vehicle-treated rats (a, c, e, g), extensive inflammation was evident (a). In vehicle-treated animals, many ED-1 positive cells were present thus demonstrating that those cells were in a state of active phagocytosis (c). Sections from drug-treated animals did not contain any ED-1 positive cells at all (d). LFB myelin stain reveals demyelination (e) in the areas corresponding to massive ED-1 positive macrophage infiltration (c). Bielschowsky silver impregnation shows reduction of axonal density in demyelinated area rectangle (g). The loss of myelin stain and the axonal pathology demonstrated in the vehicle animals are in sharp contrast to the drug-treated rats in which no damage to myelin nor pathological changes of axons were found (f, h). Squares in (e) and (f) correspond to the area depicted in (g) and (h), respectively. Journal of Neuroimmunology 2003 142, 75-85DOI: (10.1016/S0165-5728(03)00264-9)

Fig. 4 Expression of receptors and ligands in CNS of vehicle- and drug-treated rats. Darkfield microphotograps demonstrating radioactive in situ hybridization histochemical detection of CCR1, CCL3, CCR5 and CX3CR1 mRNA. Intensive expression of these mRNAs was present in all vehicle-treated animals (left panel) while in the drug-treated rats (right panel) no mRNA was detected except for the CX3CR1 which was expressed at levels equal to those in healthy control rats (data not shown). Journal of Neuroimmunology 2003 142, 75-85DOI: (10.1016/S0165-5728(03)00264-9)

Fig. 5 Serum levels of anti-MOG antibodies in vehicle- and drug-treated rats. CCR1 antagonist treatment did not result in reduced antibody reactivity against MOG. Instead, recorded antibody levels were in fact higher in drug-treated group B rats. Box plots of MOG-specific IgG and IgG isotype specific responses at a 1/200–1/2000 dilution. Rat sera were collected from group A rats on day 13 post immunization (n=4) and group B rats 4 days after individual onset of disease (n=12). The box plot depicts 10th and 90th percentile (whiskers), 25th and 75th percentile as well as median. Pairwise comparisons were performed by Mann–Whitney U-test. *<0.05, **<0.01, ***<0.001. Journal of Neuroimmunology 2003 142, 75-85DOI: (10.1016/S0165-5728(03)00264-9)