Densitometry

analysis was conducted using ImageJ software (NIH). Student’s unpaired t-test was used to measure statistical significance between two groups and one-way ANOVA with Dunnet’s multiple comparison test was used to determine statistical significance between multiple groups against WT control. All statistical analyses were performed by Prism 5 (Graphpad Software). We thank Dr. Clifford Lowell for providing Itgb2−/− mice and Dr. Hua Gu and Dr. Phil Greenberg for providing Cblb−/− mice. We would also like to acknowledge Dr. Amy Weinmann for advice on chromatin immunoprecipitation MK-8669 solubility dmso and thank members of our laboratory for helpful discussions and review of the manuscript. This work was supported by NIH grants R01AI073441 and R01AI081948, an Investigator Award from the Cancer Research Institute, a pilot award from the Alliance for Lupus Research and DOD grant W81XWH-10-1-0149 (to J.A.H). N.Y. was supported in part by NIH training grant 5T32CA09537. The authors declare no financial or commercial conflict of interest. As a service to our authors and readers, this journal provides supporting information supplied by the authors. Such materials are peer reviewed and may be re-organized for online delivery, but are not copy-edited or typeset. Technical support issues arising from supporting information (other than missing files) should be addressed to the authors.

Figure S1. Phenotypic characterization of Itgb2-/- macrophages. (A) The expression of integrin alpha subunits, CD11a, CD11b, CD11c and F4/80 was determined on bone marrow-derived macrophages by flow cytometry. Selleck AZD9291 (B) Macrophages were stimulated with the indicated concentrations of IFNγ for 48 hours and MHC II expression was assessed by flow cytometry. (C) Macrophage surface expression of TLR4, TLR2 and Dectin-1 was determined by flow cytometry. (D) TLR9 mRNA expression was determined

by qPCR, with levels normalized to GAPDH. The data are shown as mean +/- SD of triplicate wells and representative of 3 experiments.! Figure S2. Itgb2-/- macrophages are GNA12 hypersensitive to TLR stimulation. (A) Representative data of the results shown in Fig. 1A. Macrophages were stimulated with the indicated TLR agonists and supernatant cytokine concentrations were determined by ELISA 24 hours later. Results are displayed as mean +/- SD of independently stimulated wells from one experiment. (B) Expression of IL-23 p19 and IL-12 p35 was determined by qPCR, with values normalized to GAPDH. Results are representative of 2 experiments and shown as mean +/- SD of triplicate wells. (C) Representative data of the results shown in Fig. 1B. Kinetics of cytokine secretion as assessed by ELISA. Results are shown as mean +/- SD independently stimulated triplicate wells from one experiment. * p < 0.05, ** p < 0.01, *** p < 0.001! Figure S3. Isolation of thioglycollate-elicited macrophages (A) Mice were injected i.p.