PGE₂ is increased in human lung tumor tissue and PGE₂ increases the expression of TREM-1 in macrophages.

<p>A) PGE₂ levels measured from the human lung tumor tissue showed an increase in PGE₂ message from cancer tissue with no significant detection in the normal lung (n = 3, p<0.05). B) Bone marrow derived macrophages from wild type mice were treated with recombinant PGE₂ or PGD₂ (10 µmol) for 12 hours and TREM-1 expression was detected. BMDM treated with PGE₂ showed an increase in TREM-1 message in response to PGE₂ treatment (n = 3–4, p<0.05) however TREM-1 message was not detected in cells that were treated with PGD₂.</p

TREM-1 expression is abrogated in macrophages with COX-2 siRNA.

<p>A) Western blot analysis from human macrophages with COX-2 siRNA confirmed knock down of COX-2 protein in cells with siCOX-2 compared to the mock (control) siRNA. B) FACS images demonstrating TREM-1 expression from human macrophages expressing control siRNA or COX-2 siRNA co-cultured with NL-20 cells or cancer cells (A549, H23 or H 838 cells). The expression of TREM-1 was increased in macrophages expressing control siRNA co-cultured with cancer cells whereas macrophages with siCOX-2 showed an attenuated expression of TREM-1. C) Percentage of cells that stained positive for TREM-1 staining n = 3, p<0.05). D) Representative RT-PCR from macrophages with siCOX-2 show a decreased TREM-1 message compared to macrophages with control siRNA.</p

Proposed mechanism of TREM-1 expression in TAMs by lung cancer cells.

<p>COX-2 induction in lung cancer cells leads to production of PGE2 which then causes expression of TREM-1 in tumor associated macrophages. These effects are medicated through EP2 and EP4 receptors and driven through cAMP. Expression of TREM-1 in macrophages increases production of mediators that propagate tumor growth, migration and invasion.</p

TREM-1 is upregulated in lung cancer tissue and tumor associated macrophages in human non-small cell lung carcinoma.

<p>A) RNA extracted from human lung cancer samples showed an increase in TREM-1 message. B) Real time RT PCR confirmed the increase in TREM-1 message (n = 3, *p<0.05). C) Representative immunohistochemistry image demonstrating an increased expression of TREM-1 protein in macrophages in tumor stroma D) Confocal microscopy with CD68 a specific macrophage marker and TREM-1 confirmed that cells expressing TREM are TAMs.</p

TREM-1 is induced in human macrophages co-cultured with lung adenocarcinoma cells.

<p>A) Macrophages were co-cultured with NL-20 (normal epithelial cells) or lung cancer cells (A549, H23 or H838) for 48 hours. TREM-1 expression was increased in macrophages that were co-cultured with cancer cells as demonstrated by FACS analysis B) percentage of cells that stained positive for TREM-1, n = 3–4, * p<0.05) C) Representative RT-PCR from macrophage co-cultured with tumor cells showed an showed an increased expression of TREM-1 message.</p

TREM-1 expression is attenuated in monocytes co-cultured with tumor cells that are treated with COX-2 inhibitors.

<p>A) Representative image of FACS analysis-U937 monocytes were co-ultured with NL-20 or A549 cells in the presence of absence of NS-398 (100 µmol) (specific COX-2 inhibitor). The expression of TREM-1 was increased in U937 cells co-cultured with A549 cells. Treatment with NS-398 inhibited this increased expression B) Percentage cells that stained positive with TREM-1 (n = 4–5, p<0.5).</p

Figure 6

<p>A) FACS images demonstrating that TREM-1 expression is attenuated in macrophages that are co-cultured with tumor cells in the presence of EP2 and EP4 antagonist (AH6809 and AH23848) whereas it is increased in the presence of forskolin. B) Percentage cells that stained positive with TREM-1 (n = 4–5, *p<0.05).</p

An example of spatial representation of post-stimulus interactions after targets between all investigated brain sites in one subject (No.7).

<p>Correlation results are arranged in the triangular matrices into groups according to brain structures (delimited by black lines)(D) and in graphic form of “glass brains” with linked pairs of investigated electrode contacts (A – Coronal, B – Sagittal, C – Axial). Matrix values and links are colored according to the percentage of duration of the increase (red) or decrease (blue) in cross-correlations within time window 250–750 ms after stimulation. Three selected frequency bands – δ (left panel), β (middle panel), and upper γ (right panel).</p

Relative incidences of significant post-stimulus changes in inter-areal cross-correlations (A – monopolar; B – bipolar) and post-stimulus power changes (C – monopolar; D – bipolar) across all investigated subjects.

<p>Statistical significance of target/frequent differences is indicated by asterisks.</p

Timing of significant post-target cross-correlation increases across all subjects and areas.

<p>The duration was determined as the mean level of corresponding changes over contact pairs, and the occurrence was given as the center of changes.</p