Our present study identifies IFN- as a targetable molecule in the mouse model of PCD and, possibly, in the human disease. Human PCD, which is likely mediated by autoimmune response of T cells against Purkinje cell antigens CDR2/CDR2L, is usually characterized by development of severe cerebellar dysfunction (5). (left, WT; not expressing HA in Purkinje cells) and L7-HA-PCD mice (middle). USP7-IN-1 Level bar: 10 m. Staining for Calbindin (green) and pSTAT1 (reddish) shows upregulation of pSTAT1 in the nucleus of a Purkinje cell (right). Scale bar: 7.5 m. (E) Left: ex vivo cerebellar slices from L7-HA mice treated or not with USP7-IN-1 IFN- (100 U/ml) for 24 hours and stained with an anti-calbindin antibody (green) and an antiCH2-Kd antibody (reddish). Scale bar: 20 m. Right: densitometric analysis of calbindin and H2-Kd staining of Purkinje cells. Yellow lines: segments of Purkinje cells submitted to densitometric analysis of calbindin and H2-Kd staining (right panels). (F) Top left: pSTAT1 staining in the cerebellum of an anti-Ma2 case. Bottom left: enlargement of top left. pSTAT1 upregulation in the nuclei of microglial cells, astrocytes, and some of the granular neurons. Top right: local pSTAT1 in the cerebellar peduncle of an anti-Yo case. Bottom right: pSTAT1 is usually upregulated in various glial cells. In addition, pSTAT1 can be seen in the nucleus of a neuron (arrowhead). Level bar: 50 m (top left and right) and 20 m (bottom left and right). Altogether, our findings recognized 4 integrin and IFN- as potential therapeutic targets in our mouse model, and we, thus, designed in vivo experiments to validate or invalidate them. = 0.27) decreased in mice treated with the anti-4 integrin mAb TPOR (324.4 125.5/mm2; = 8 mice) vs. mice treated with the control antibody (511.3 134.8/mm2; = 8 mice). Using an established and efficient treatment regimen (13), these data show that this anti-4 integrin therapeutic approach is not efficient at blocking ongoing disease, at least in our model of PCD. The CCR5 chemokine receptor has been implicated in immune cell recruitment in several inflammatory diseases of the CNS (14). Although it was not highly expressed on cerebellum-infiltrating T cells at day 16 (Supplemental Physique 2A), we investigated whether CCR5 signaling was required for disease development. Treatment of L7-HA-PCD mice with a CCR5 antagonist (5P12-RANTES) starting on day 10 after disease induction did not block development of PCD and Purkinje neuron destruction, as shown in Supplemental Physique 2, BCE. This antagonist was, however, efficient to restrict brain inflammation but not spinal cord inflammation in a model of experimental autoimmune encephalomyelitis (EAE) following juvenile virus contamination in mice (D. Merkler [Department of Pathology and Immunology, University or college of Geneva, Switzerland], personal communication). Open in a separate window Physique 2 Treatment with an antiC4 integrin mAb shows no efficacy in the PCD model.(A) USP7-IN-1 Tumor size, (B) mouse excess weight loss, and (C) rotarod cumulative score (left) and performance on day 20 (right) of L7-HA-PCD mice treated with PS/2 (antiC4 integrin mAb) or control IgG2b from day 10 after the induction of disease onward (= 8/group, 2 impartial experiments). (D) Left: representative staining for calbindin (violet) and nuclear counterstaining with hematoxylin (blue) on cerebellar sections from a control WT mouse, and from L7-HA-PCD mice treated with isotype control or PS/2. Scale bar: 100 m. Right: quantitative assessment of Purkinje cell density in L7-HA-PCD mice treated with IgG2b or PS/2 (= 8/group, 2 impartial experiments). The shaded area represents the normal range in WT mice, meaning mean 2SD. = USP7-IN-1 23 per group, 4 impartial experiments; 2-way ANOVA post-hoc Sidaks multiple comparison test, **< 0.01). (C) Rotarod overall performance of L7-HA-PCD mice treated with USP7-IN-1 XMG1.2 or control IgG1 (= 16 mice per group, 3 indie experiments; Mann-Whitney test, *< 0.05; **< 0.01). (D) Histological analysis at day 20 of the cerebellum of L7-HA-PCD mice treated with XMG1.2 or IgG1, from day 10 onward. Left: representative staining for calbindin (brown) and nuclear counterstaining with hematoxylin (blue). Level bar: 100 m. Right: quantitative assessment of Purkinje cell density in L7-HA-PCD mice treated with XMG1.2 or IgG1 (= 23 mice per group from 4 indie experiments; Mann-Whitney test, ***< 0.01). (E) IFN- deficiency in Purkinje cellCspecific CD8 T cells does not prevent the development of the.
Our present study identifies IFN- as a targetable molecule in the mouse model of PCD and, possibly, in the human disease