2 and supplementary Fig. S2). Up-regulation of Gr-1 is not part of the maturation process demonstrated in Fig. 7, and although a role in limiting T-cell proliferation is not ruled out by this experiment, the soluble mediators NO and PGE2 together are sufficient to restrict T-cell proliferation. Finally,
PLX4032 chemical structure it was striking that despite the strong phenotype of TNFR1−/− Mϕin vitro and in vivo, we could restore near normal inhibitory function by treating with a combination of soluble mediators (Fig. 7). This result illustrates on the one hand the emergent properties that multiple signals can have on cell function and on the other hand the many levels of redundancy that are inherent in Mϕ responses. This redundancy complicates the analysis of function BGJ398 cell line in vitro and in vivo, but it is also likely to produce immune responses that in the wild are more robust and less
susceptible to a single targeted attack by a pathogen. This work was carried out with support from the National Eye Research Centre. The authors declare no competing interests. Figure S1. WT BM-Mφ were prepared as described in the methods section. Cells were then stained with antibodies against CD11b, CD31, F4/80, and Gr-1. Plot A shows F4/80 and CD11b expression by naïve BM-Mφ. Plots B and C show CD31 and Gr- 1 expression naïve BM-Mφ (black lines) compared with isotype controls (grey filled). Figure S2. WT or TNFR1−/− BM-Mφ were co-cultured with OT-II T cells in the lower
chambers of 0.22 μm transwells in the presence of 100 μg/ml OVA peptide. Equal numbers of either WT or TNFR1−/− BM-Mφ were added to the top chamber. After 72 hr, from the both chambers were harvested separately and stained with antibodies against CD11b and Gr-1 for flow cytometric analysis. Plots show Gr-1 expression of CD11b+ cells, with Mφ from the top chamber (red lines) and those from the lower chamber (blue lines). Figure S3. WT or TNFR1−/− BM-Mφ were co-cultured with OT-II T cells in the presence of 100 μg/ml OVA peptide for 72 hr. L-NMMA or SNAP was added at the indicated concentrations. T cell proliferation was measured over the final 8 hr of culture. NO production was measured in the culture Glutamate dehydrogenase supernatant. Plots show the effect of addition of L-NMMA or SNAP on NO production and proliferation on cocultures containing WT (black lines) or TNFR1−/− (grey lines) Mφ, as compared with control co-cultures in which Mφ alone were cultured. “
“M protein is an important virulence determinant in Streptococcus pyogenes, but the amounts of M protein in various strains of the species remain to be elucidated. To assess the amount of M protein in strains of each emm genotype, dot blot analysis was performed on 141 clinically isolated strains. Among the cell membrane-associated proteins, M protein was present in greater quantities in the emm1, 3, and 6 strains than in the other emm strains.