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cei12219-sup-0001-fig_s1.tif4097K

Fig. S1. Purity of the primary neural cell cultures. (a) Purity of the primary cultured astrocytes and cerebellar granule neurones (CGNs) were confirmed by immunofluorescent microscopy with the anti-glial fibrillary acidic protein (GFAP) antibody or a neurone-specific class III beta-tubulin antibody (Tuj1), respectively. (b) Detection of the microglias with anti-CD11b antibodies in the primary astrocyte culture by immunofluorescent microscopy. Similar results were obtained from three independent experiments.

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Fig. S2. Selection of the dilution factor for the inflammatory media (Infl. Med.) that applied to the primary CGN cultures. The half-diluted Infl. Med. induced increased neuronal apoptosis over time and allowed further experiments, whereas a full Infl. Med. induced neuronal cell death quickly and thoroughly within 6 h of incubation. Similar results were obtained from three independent experiments.

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Fig. S3. Reduction of the level of interleukin (IL)-17 in the inflammatory media (Infl. Med.) over time at a 37°C incubator. The peripheral blood mononuclear cells (PBMC) were stimulated with 50 ng/ml phorbol myristate acetate (PMA) plus 1 μg/ml ionomycin for 6 h, the supernatant (as the Infl. Med.) were collected and left in the 37°C incubator for 2, 4 and 6 h; the levels of IL-17 of these supernatants were then examined by enzyme-linked immunosorbent assay (ELISA). All data are means ± standard error of the mean of at least three experiments. **P < 0·01; ***P < 0·001.

cei12219-sup-0004-fig_s4.tif263K

Fig. S4. The reduced level of interleukin (IL)-17 in the inflammatory media (Infl. Med.) by the neutralizing anti-IL-17 antibody. The peripheral blood mononuclear cells (PBMC) were stimulated with 50 ng/ml phorbol myristate acetate (PMA) plus 1 μg/ml ionomycin for 6 h; the supernatant was collected as the phorbol myristate acetate (PMA) (Infl. Med.). The respective neutralizing antibodies against IL-17 or interferon (IFN)-γ were applied at 4 h of stimulation, resulting in the conditional Infl. Med. with either reduced levels of IL-17 or IFN-γ, compared to the original inflammatory media. The level of IL-17 was measured by enzyme-linked immunosorbent assay (ELISA). All data are means ± standard error of the mean of at least three experiments. *** P < 0·001.

cei12219-sup-0005-fig_s5.tif4134K

Fig. S5. The involvement of microglias during the development of experimental autoimmune uveitis (EAU) in rats. Immunofluorescent microscopy was used to detect microglias with anti-CD11b antibodies on the retina of EAU rats across the disease. The positive-staining was shown only at the peak of disease at 14 dpi. Similar results were obtained from three independent experiments.

cei12219-sup-0006-table_s1.doc41K

Table S1. Relative expression of T cell-associated molecules from phorbol myristate acetate (PMA)/ionomycin-stimulated astrocytes at the mRNA level by real-time reverse transcription–polymerase chain reaction (qRT-PCR).

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