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. Author manuscript; available in PMC: 2015 Jul 15.
Published in final edited form as: J Immunol. 2014 Jun 13;193(2):597–609. doi: 10.4049/jimmunol.1303048

Figure 5. PIM1 and NFATC2 signaling is promoted in fibroblast-activated macrophages.

Figure 5

(A) PIM1 and NFATC2 expression in laser-capture micro-dissected PA tissue from humans with iPAH compared to controls (both n= 8) expressed as δCt values normalized to expression of porphobilinogen deaminase (PBGD). *P < 0.05 by unpaired two-tailed Student’s t-test. (B) Soluble factors from the dPA adventitia (using dPA explants and 0.4-µm Transwells as in Fig. 2) induce gene expression of Pim1 and Nfatc2 in mouse BMDMs. Displayed is the fold-induction (normalized to basal expression) of a representative PCR triplicate (average +/− SEM) from one of two calves. Three dPA segments were tested from each animal; gene expression oafter incubation for 16hrs is shown. *P < 0.05 by one-way ANOVA. (C) Bovine PH-Fib CM induces gene expression of PIM1 and NFATC2 in bovine (16hr time point shown), mouse (D, time course is shown), and rat (E; 16hr time point is shown) BMDMs. (C–E) Displayed are PCR triplicates of a representative experiment with one CO-Fib and one PH-Fib CM (mean ± SEM) normalized to Hprt1 expression and expressed relative to basal (untreated) gene expression. These are representative of 3 experiments with CM from 3 separate CO-Fibs and 3 separate PH-Fibs cell populations on BMDMs from 3 different animals. *P < 0.05 by unpaired two-tailed Student’s t-test. (F) siRNA-mediated suppression of IL6 gene transcription in bovine PH-Fibs limits the ability of CM to induce transcription of Pim1 and Nfatc2 in WT mouse BMDMs. Gene expression is normalized to expression of Hprt1 and relative to that in macrophages exposed to CM from CO-Fibs treated with control siRNA. Data are mean ± SEM from triplicate determinations and representative of two separate experiments. (G) Expression of Pim1 and Nfatc2 in WT and Il6−/− BMDMs exposed for 16 hrs to bovine PH-Fib CM. *P < 0.05 by unpaired two-tailed Student’s t-test of triplicate PCR analysis; one representative experiment with CM from one of three PH-Fib populations was tested on BMDMs from 3 different animals of each genotype.