Macrophages.

Extracellular stimuli may influence the M1/M2 phenotypic polarization of
macrophages. We examined M1/M2 biomarkers, phagocytic activity, and tumoricidal
activity in RAW 264.7 mouse macrophages. Macrophages were treated with conditioned
media (CM) from 4T1 breast cancer cells, curcumin, 22-oxacalcitriol, LPS, or a
combination of the previously listed. Arginase activity, a M2 phenotypic biomarker, was
upregulated by the treatment of macrophages with conditioned media. Curcumin, 22-
oxacalcitriol, and LPS partially inhibited RAW 264.7 arginase activity in the presence of
4T1 breast cancer media. 22-oxacalcitriol increased the phagocytic ability of RAW 264.7
macrophages in the presence of M2 polarizing substances produced by the 4T1 breast
cancer cells. Also, LPS increased RAW 264.7 phagocytic ability in the presence of 4T1
breast cancer CM. This study looked at the potential substances that would possibly reverse the M2 tumor promoting macrophage phenotype seen in the breast cancer tumor
environment.

Chitin Microparticles (CMPs, 1-10um), a special form of the ubiquitous and nontoxic
polysaccharide Chitin (GlcNAc), is capable of inducing a switch in macrophages
from the wound-healing M2 phenotype to the classically activated pro-inflammatory M1
phenotype; which has therapeutic implications in allergy and cancer. We hypothesized
that TLR2 forms a complex with CMPs and Chitin-Binding Proteins (CBPs) at the
surface of peritoneal macrophages and remains with that complex after internalization to
initiate downstream signaling events, leading to the production of the M1 cytokine, TNFalpha.
Our results from experiments performed in RAW 264.7 cells show that TLR2 and
TLR1, but not TLR6, are associated with the CMP binding fraction, and that both TLR1
and TLR2 might be important for M1 activation as a result of CMP phagocytosis. This
project sheds light on CMP as a potential therapeutic agent and provides more evidence
for a phagocytosis-dependent TLR2 signaling pathway.