PGRN restrains macrophage inflammatory responses by synergizing with LPS for induction of IL-10 transcription. (A) Effect of anti–IL-10 antibody on TNF and MCP-1 production from BMDMs. 1 µg/ml anti–IL-10 or control IgG was added together with LPS and cytokines quantitated by ELISA as in Fig. 2 B. Results are expressed as means ± SEM of triplicate samples from one out of four experiments. (B) PGRN regulates IL-10 expression at the transcriptional level. Quantitative RT-PCR for nascent IL-10 transcripts (left) or spliced IL-10 transcripts (right) was measured 1 h after exposure of BMDMs to 100 ng/ml LPS. Results are expressed as means ± SEM of triplicate samples from one out of two independent experiments. (C) Recombinant PGRN synergized with LPS in IL-10 induction. RAW264.7 cells were incubated with the indicated concentrations of LPS and PGRN for 20 h. IL-10 in the conditioned media was measured by ELISA. Results are expressed as means ± SEM of three independent experiments. *, P < 0.05 using the Student’s t test.