Suppressors of Cytokine Signaling (SOCS)1 and SOCS3 Proteins Are Mediators of Interleukin-10 Modulation of Inflammatory Responses Induced by Chlamydia muridarum and Its Major Outer Membrane Protein (MOMP) in Mouse J774 Macrophages

Figure 2

SOCS1 and SOCS3 transcriptional and translational expressions are differentially induced and regulated by chlamydial stimulants and exogenous IL-10 in macrophages. Macrophages were exposed to dose-dependent concentrations of IL-10 (0.1, 1 and 10 ng/mL) in the presence and absence of rMOMP (10 μg/mL). RNA samples were collected at 0, 0.5, 1, 2, and 24 h post-stimulation to quantify the mRNA gene transcript of SOCS1 (A) and SOCS3 (B). RNA samples were collected at 24 h from macrophages stimulated with Cm (MOI of 2) with and without dose-dependent concentrations of IL-10 to quantify SOCS1 and SOCS3 transcripts (C). Macrophages were stimulated with rMOMP (0.1, 1 and 10 μg/mL) or Cm (MOI of 0.5, 1 and 2) with and without IL-10 (10 ng/mL) and SOCS1 and SOCS3 transcripts were quantified at 24 h post-stimulation (D-E). For TaqMan qRT-PCR, all values were normalized with respect to the mRNA levels of the “housekeeping” gene that codes for GAPDH. Results are presented as fold increase over the control (i.e., the level in unstimulated cells. Calculations of SOCS1/SOCS3 (F) and SOCS3/SOCS1 (G) ratios in macrophages exposed to rMOMP (10 μg/mL) or Cm (MOI of 2) in the presence and absence of IL-10 (10 ng/mL). Macrophages were exposed to rMOMP (1 μg/mL) or Cm (MOI of 2) in the presence and absence of IL-10 to evaluate SOCS1 and SOCS3 expression by flow cytometry (H-M) and SOCS3 by immunofluorescence microscopy (N) 24 h post-incubation. An asterisk indicates significant differences (P <0.05), and P values were calculated as described in Figure 1 legend. Each bar represents the mean ± SD of samples run in triplicates, and each experiment was repeated at least 4 times with a representative experiment shown.

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