In vitro evaluation of inhibitory effect of nuclear factor-kappaB activity by small interfering RNA on pro-tumor characteristics of M2-like macrophages

Biol Pharm Bull. 2014;37(1):137-44. doi: 10.1248/bpb.b13-00659. Epub 2013 Oct 19.


Tumor-associated macrophages (TAMs) have an alternatively activated macrophage phenotype (M2) and promote cancer cell proliferation, angiogenesis and metastasis. Nuclear factor-kappaB (NF-κB) is one of the master regulators of macrophage polarization. Here, we investigated the effect of inhibition of NF-κB activity by small interfering RNA (siRNA) on the pro-tumor response of macrophages located in the tumor microenvironment in vitro. We used mouse peritoneal macrophages cultured in conditioned medium from colon-26 cancer cells as an in vitro TAM model (M2-like macrophages). Transfection of NF-κB (p50) siRNA into M2-like macrophages resulted in a significant decrease in the secretion of interleukin (IL)-10 (a T helper 2 (Th2) cytokine) and a significant increase of T helper 1 (Th1) cytokine production (IL-12, tumor necrosis factor-α, and IL-6). Furthermore, vascular endothelial growth factor production and matrix metalloproteinase-9 mRNA expression in M2-like macrophages were suppressed by inhibition of NF-κB expression with NF-κB (p50) siRNA. In addition, there was a reduction of arginase mRNA expression and increase in nitric oxide production. The cytokine secretion profiles of macrophages cultured in conditioned medium from either B16BL6 or PAN-02 cancer cells were also converted from M2 to classically activated (M1) macrophages by transfection of NF-κB (p50) siRNA. These results suggest that inhibition of NF-κB activity in M2-like macrophages alters their phenotype toward M1.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Arginase / genetics
  • Arginase / metabolism
  • Cell Line, Tumor
  • Colonic Neoplasms / genetics
  • Colonic Neoplasms / metabolism*
  • Cytokines / metabolism*
  • Interleukins / metabolism
  • Lipopolysaccharides
  • Macrophage Activation
  • Macrophages / metabolism*
  • Macrophages, Peritoneal / metabolism*
  • Matrix Metalloproteinase 9 / metabolism
  • Mice
  • NF-kappa B / antagonists & inhibitors*
  • Neovascularization, Pathologic*
  • Nitric Oxide / metabolism
  • Phenotype
  • RNA, Messenger / metabolism
  • RNA, Small Interfering / metabolism*
  • Signal Transduction
  • Th1 Cells / metabolism
  • Transfection
  • Tumor Necrosis Factor-alpha / metabolism
  • Vascular Endothelial Growth Factor A / metabolism


  • Cytokines
  • Interleukins
  • Lipopolysaccharides
  • NF-kappa B
  • RNA, Messenger
  • RNA, Small Interfering
  • Tumor Necrosis Factor-alpha
  • Vascular Endothelial Growth Factor A
  • vascular endothelial growth factor A, mouse
  • Nitric Oxide
  • Matrix Metalloproteinase 9
  • Arginase