The administration of rabies virus neutralizing antibodies fails to clear CVS-F3 from the CNS of JHD^−/− mice.

<p>JHD ^−/− mice were infected i.n. with CVS-F3 and treated at 7, 9, 11, 13, and 15 days post-infection with saline vehicle alone or the rabies-specific antibody 1112. Three weeks after the final treatment mice were transcardially perfused, cerebellar tissues were removed, and virus replication was estimated by quantifying nucleoprotein mRNA levels. Virus replication was similarly estimated in cerebellar tissues from wild-type mice infected with CVS-F3 24 days previously. Virus replication is expressed as the mean±S.E.M. copies of rabies virus nucleoprotein mRNA per copy of the housekeeping gene L13 mRNA in the tissue sample.</p

Rabies virus-specific antibody secreting B cells accumulate in the CVS-F3-infected CNS.

<p>Mononuclear cells isolated from peripheral blood lymphocytes (PBL) and brain (CNS) were assessed for surface phenotype using flow cytometry and for the numbers of rabies virus-specific antibody secreting cells by ELISPOT. The percentage of B cells, identified as positive for both CD19 and MHC class II, among total mononuclear cells is presented on the left side of the figure while the proportion of the total mononuclear cells producing rabies virus-specific antibodies is presented on the right side of the figure.</p

Survival of mice immunized or infected with CVS-F3 prior to infection with CVS-N2c.

<p>Survival of mice immunized or infected with CVS-F3 prior to infection with CVS-N2c.</p

The T cell response to CVS-F3 infection in the absence of B cells mediates enhanced BBB permeability and long-term survival.

<p>The extent of BBB permeability in the cerebellum of wild-type, JHD^−/− and RAG-2^−/− mice, infected i.n. with CVS-F3 10 days previously, is shown in panel A. Weight loss over the course of infection in JHD^−/− and RAG-2^−/− mice is shown in panel B, while the number of copies of rabies virus nucleoprotein mRNA at different times of infection in these animals is presented in panel C. BBB permeability changes are shown as the mean±S.E.M. fold increase in Na-Fluorescein uptake in the tissues with the levels from uninfected mice taken as 1. Weight is expressed as the mean±S.E.M. percent body weight with the weight on day 0 being taken as 100% and copies of nucleoprotein mRNA in the tissues are expressed per copy of L13 mRNA in the same sample.</p

IFN-γ Signaling to Astrocytes Protects from Autoimmune Mediated Neurological Disability

<div><p>Demyelination and axonal degeneration are determinants of progressive neurological disability in patients with multiple sclerosis (MS). Cells resident within the central nervous system (CNS) are active participants in development, progression and subsequent control of autoimmune disease; however, their individual contributions are not well understood. Astrocytes, the most abundant CNS cell type, are highly sensitive to environmental cues and are implicated in both detrimental and protective outcomes during autoimmune demyelination. Experimental autoimmune encephalomyelitis (EAE) was induced in transgenic mice expressing signaling defective dominant-negative interferon gamma (IFN-γ) receptors on astrocytes to determine the influence of inflammation on astrocyte activity. Inhibition of IFN-γ signaling to astrocytes did not influence disease incidence, onset, initial progression of symptoms, blood brain barrier (BBB) integrity or the composition of the acute CNS inflammatory response. Nevertheless, increased demyelination at peak acute disease in the absence of IFN-γ signaling to astrocytes correlated with sustained clinical symptoms. Following peak disease, diminished clinical remission, increased mortality and sustained astrocyte activation within the gray matter demonstrate a critical role of IFN-γ signaling to astrocytes in neuroprotection. Diminished disease remission was associated with escalating demyelination, axonal degeneration and sustained inflammation. The CNS infiltrating leukocyte composition was not altered; however, decreased IL-10 and IL-27 correlated with sustained disease. These data indicate that astrocytes play a critical role in limiting CNS autoimmune disease dependent upon a neuroprotective signaling pathway mediated by engagement of IFN-γ receptors.</p> </div

Astrocytes regulate IL-1, CCL5 and TNF mRNA during acute EAE.

<p>Relative gene expression in spinal cord from GFAPγR1Δ (average clinical score = 3.9) and wt mice (average clinical score = 3.1) during acute disease (day 18 p.i.) determined by qRT-PCR. Representative of 2 experiments (n = 3 – 4 mice/group) analyzed in triplicate. *p<0.05 comparing wt and GFAPγR1Δ mice.</p

IFN-γ signaling to astrocytes does not influence brain inflammation.

<p>(<b>A</b>) CD45^hi bone marrow derived inflammatory cells and CD4⁺ T cells in brains (day 16 p.i.) analyzed by flow cytometry following isolation by Percoll gradients. R1 gate = CD45^hi inflammatory cells; R2 gate = CD4⁺ T cells within R1. Representative of 4 separate experiments at days 14 – 16 p.i. (<b>B</b>) IFN-γ and IL-17 secreting MOG-specific CD4⁺ T cells during acute EAE. Brain derived cells enriched by Percoll gradients at day 16 p.i. stimulated with MOG^35–55 peptide for 4 hrs in the presence of GolgiStop. p>0.05 comparing the frequency of MOG-specific CD4⁺ T cells secreting IFN-γ; p<0.05 comparing IL-17 in wt and GFAPγR1Δ mice. CNS cells pooled from 4 – 5 mice per experiment. Data are representative of 3 – 4 separate experiments at days 14 – 16 p.i. (<b>C</b>) Foxp3⁺ regulatory T cells within CD4⁺ T cells enriched from brain at day 16 p.i. CNS cells pooled from 4 – 5 mice per experiment. p<0.05 comparing wt and GFAPγR1Δ mice. Data are representative of 3 separate experiments at days 14 – 16 p.i.</p

IFN-γ signaling regulates astrocyte activation within grey matter during chronic EAE.

<p>Cross section of spinal cord grey matter from wt (upper panel) and GFAPγR1Δ mice (lower panel) during chronic disease (day 35 p.i.). Grey matter regions shown are distinct from areas of demyelination. Data are representative of 3 separate experiments with 3 – 4 individuals per experiment and 6 cross sections per spinal cord and 2 separate experiments in which the spinal cords were sectioned longitudinally.</p

EAE in GFAPγR1Δ tg mice.

a<p>Peak disease score determined at day 18 post immunization.</p>b<p>Mortality determined at day 50 post immunization.</p>*<p>p≤0.05.</p

Sustained demyelination, axonal loss and astrocyte activation during chronic disease.

<p>Cross section of spinal cord from wt (upper panels) and GFAPγR1Δ mice (lower panels) during chronic disease (day 35 p.i.). Inflammation (HE), demyelination (LFB), axonal damage and loss following visualization with mAb SMI31 and SMI32 (SMI), GFAP expression and hypertrophy of astrocytes (GFAP) in wt and GFAPγR1Δ mice. Data are representative of 3 separate experiments with 3 – 4 individuals per experiment and 6 cross sections per spinal cord.</p