Skip to main content
. Author manuscript; available in PMC: 2017 Jul 28.
Published in final edited form as: Prostaglandins Other Lipid Mediat. 2015 Feb 14;116-117:64–75. doi: 10.1016/j.prostaglandins.2015.01.004

Fig. 4.

Fig. 4

Pro-inflammation transfers from cardiomyocytes to macrophages. (A) Scheme of macrophage culture in inflammation responded cardiomyocytes-conditioned medium. Briefly, cardiomyocytes were treated as indicated, conditioned medium harvested and transferred to macrophages for the indicated times, and inflammatory cytokine production in macrophages assayed by RT-qPCR and ELISA. (B) The nuclear translocation of NF-κB p65 was induced in RAW 264.7 culturing in Iso or Ang II stimulated H9c2 cell-conditioned medium, respectively. 14,15-EET pretreatment suppressed the activation of NF-κB p65 in RAW 264.7. Proteins were normalized to β-actin or LaminB. *P < 0.05 vs. H9c2 control c.m., #P < 0.05 vs. H9c2 Iso.c.m. or H9c2 Ang II c.m. (C) and (D) NF-κB p65 activity of DNA binding detected by EMSA. (E)–(J) Inflammation and profibrotic response were induced in RAW 264.7 culture in Iso or Ang II stimulated H9c2 cell-conditioned medium, and 14,15-EET pretreatment inhibited the effects above. Inflammatory factors IL-1β, IL-6, IL-10 and MCP-1 were detected by ELISA, profibrotic factor TGF-β1 was detected by RT-qPCR and ELISA. n = 3 to 5 for each experiment. *P < 0.05 vs. H9c2 Control c.m., #P < 0.05 vs. H9c2 Iso or Ang II c.m., &P < 0.05 vs. H9c2 Iso or Ang II + 14,15-EET c.m. (K) and (L) The nuclear translocation of NF-κB p65 was induced in primary macrophages culturing in Iso or Ang II stimulated primary cardiomyocytes-conditioned medium, respectively. 14,15-EET pretreatment suppressed the activation of NF-κB p65 in primary macrophages. Proteins were normalized to β-actin or LaminB. (M) and (N) Expression of inflammatory factors (IL-1β, IL-6, IL-10 and MCP-1) in the conditioned supernate from primary macrophages. n = 3–5 for each experiment. CM: primary cardiomyocytes; *P < 0.05 vs. CM Control c.m., #P < 0.05 vs. CM Iso or Ang II c.m., &P < 0.05 vs. CM Iso or Ang II + 14,15-EET c.m. Differences between data groups were evaluated for significance using ANOVAs.